Results: A significant group x time interaction was determined for aerobic capacity  at week 3 and for skinfolds at week 3  and at week 5 , in favour of the HIGH sleep group. No differences were evident between groups for strength or speed measures (p ≥ 0.05). Conclusion: This study highlights that longer sleep duration during the preseason may assist in enhancing physical qualities including aerobic capacity and body composition in elite rugby union athletes.Background: Preseason training optimises adaptations in the physical qualities required in rugby union athletes. Sleep can be compromised during periods of intensified training. Therefore, we investigated the relationship between sleep quantity and changes in physical performance over a preseason phase in professional rugby union athletes. Methods: Twenty-nine professional rugby union athletes  had their sleep duration monitored for 3 weeks using wrist actigraphy. Strength and speed were assessed at baseline and at week 3. Aerobic capacity and body composition were assessed at baseline, at week 3 and at week 5. Participants were stratified into 2 groups for analysis: <7 h 30 min sleep per night (LOW,  Repeated performance is underpinned by an athlete’s ability to recover from multiple physical and psychological stressors ,2. Thus,In rugby union, preseason training aims to enhance physical capacities, including power, speed, strength, aerobic and anaerobic fitness and body composition ,13,14. TA reduction in sleep efficiency and duration seen during intensified training periods may be detrimental to maximising physiological adaptation and muscle recovery . ResearcIn contrast to the reported adverse effects of a reduction in sleep, increasing sleep duration may be beneficial for moderating stress responses and performance during periods of intensified training . SwinbouTo the best of the authors’ knowledge, there are no studies investigating the relationship between sleep and changes in physical performance markers, including strength, speed, fitness and body composition over an intensified micro-cycle of training  in elite rugby union athletes. Therefore, the current study aimed to examine the relationship between sleep and physical performance changes throughout a preseason training phase in a group of professional rugby union athletes.A total of 29 professional rugby union athletes  from a New Zealand Super Rugby team volunteered to participate in the current study. The Super Rugby competition is the premier professional club rugby competition in the southern hemisphere and involves 15 teams from New Zealand, Australia, South Africa, Argentina and Japan. All athletes undertook a 3-week off-season training program prior to the study to ensure athletes arrived at the start of the preseason in appropriate physical condition to meet the subsequent training demands. Written informed consent was obtained from participants, and ethical approval (Ethics number: FEDU066/16) was obtained from the Institutions Research Ethics committee.A pre-post observational study design was utilised for the current investigation. Markers of strength, aerobic fitness, speed and body composition were collected across several sessions throughout the study period. Their relationship to sleep metrics was examined across a five-week training period. Sleep groups were split retrospectively based on average nightly sleep duration. The LOW sleep group displayed an average sleep duration of 6 h 55 min, less than the 7 h, which has been deemed to demonstrate “sleep loss” . ConversTM, Fatigue Science, Vancouver, BC, Canada). Athletes were required to wear a wrist actigraph on either wrist [TM, Fatigue Science, Vancouver, BC, Canada). Athletes were blinded to their sleep results throughout the duration of the study. The raw activity data were translated into sleep–wake indices including total sleep time (TST), sleep efficiency (SE%), sleep latency (SL), wake after sleep onset (WASO) and wake episodes (WE). The ReadiBand has been validated against polysomnography (PSG) and has been deemed to be acceptable with an approximately 90% agreement in total sleep duration when compared to PSG [Quantitative sleep measures were collected throughout the study via the use of wrist actigraphy  and their sleep quality on a scale of 1 to 7 . These data were collected via the wellness questionnaire described below.The Athlete Sleep Behaviour Questionnaire (ASBQ) and the Pittsburgh Sleep Quality Index (PSQI) were collected to assess various aspects of sleep behaviour. The ASBQ is an 18-item survey that includes specific questions about sleeping habits and behaviours common for elite athletes . Each rer = 0.993) and back squat (r = 0.969) [Maximal strength was assessed from one-repetition maximum (1RM) testing of the back squat, bench press and weighted chin-up exercises. Prior to testing, athletes undertook a generalised warm-up that consisted of foam rolling, static and dynamic stretching and basic locomotion. Testing protocols used for bench press and squat exercises have been previously described by Schoenfeld . Each pa= 0.969)  exerciseAthletes self-selected their hand position and were required to lower the bar to the chest lightly touching before vertically pushing the bar until the arms were fully extended. Athletes could not bounce the bar off their chest, display excessive back arching or receive any help from spotters .Athletes self-selected their hand and foot position and were required to descend until the upper thigh was below parallel with the floor (below 90°). Once the athlete had achieved adequate depth, they were required to ascend to the standing position without any assistance. Depth of the squat was visually assessed by the strength and conditioning staff, and athletes were allowed to wear weightlifting belts, but no other supportive equipment was allowed .Athletes were required to pull vertically until their chin was over the height of the bar and were required to display a controlled return to the starting position. Athletes were not permitted to utilise their legs or any swinging motion, push off the floor to assist or create any elastic energy to utilise. External weight was added by attaching weight plates to the athletes via a weight belt and chain secured around the hips . The extSpeed was measured over both 5 m and 10 m from a standing start position. The athletes were required to perform 2 to 3 trials of maximal effort sprints. Athletes were instructed to stand 50 cm behind the first timing gate prior to starting . Sprint Aerobic fitness was assessed at three time points of the study using the Bronco shuttle test. The Bronco test is widely used in rugby environments and consists of running 1200 m in a shuttle (out and back) manner . Each atBodyweight and skinfold measurements were obtained as measures of body composition for the current study. Body weight was measured to the nearest 0.1 kg using digital scales . Skinfolds were assessed at eight sites following the protocols of the International Society of the Advancement of Kinanthropometry and summed for the analyses . BodyweiA wellness questionnaire based on the recommendations of Hooper and Mackinnon  was compLocomotion activity was measured during all on-field team training, units and speed sessions throughout the 3-week preseason phase using an 18 Hz GPS unit . Each unit was worn on the upper back between the scapulae. To decrease variability, the same GPS unit was used by each participant for all sessions. After the completion of each session, the raw data were downloaded and analysed using the company’s software .trivial; 0.1–0.3, small; 0.3–0.5, moderate; 0.5–0.7, large; 0.7–0.9, very large; and 0.9–1.0, almost perfect. The data were then stratified into two groups based on the midpoint/median for total sleep time (7.5 h). For the purpose of analysis, total sleep time was selected due to being the most accurate and important measure of sleep that was collected via wrist actigraphy [n = 15), and participants who slept greater than 7.5 h per night on average were classed as the high sleep group . Statistical analysis was performed using the Statistical Package for Social Sciences , with statistical significance set at p < 0.05 for all analyses. A two-way mixed ANOVA, with 2  × 3  factors was performed for all physical performance measures and sleep questionnaires. Analysis of the studentised residuals was verified visually with histograms and by the Shapiro–Wilk test of normality. The presence of outliers was assessed via inspection of boxplots. Mauchly’s test of sphericity was used to establish equal variance within-subjects for all variables (p > 0.05). The main effects were run to identify where statistically significant differences existed. When the main effects were found, post-hoc analysis was performed. Additionally, a Wilcoxon signed-rank test was performed to determine if there were differences in individual scores for sleep questionnaires. Effect-size statistics were calculated using Cohen’s d and interpreted using thresholds of 0.2, 0.5 and 0.8 for small, moderate and large, respectively [trivial, and the effect was deemed unclear if the 95% confidence interval overlapped the thresholds for both positive and negative effects [All descriptive statistics are reported as means ± 95% confidence intervals unless otherwise stated. The data for sleep measures, changes in performance, wellness scores and training load were pooled, and Pearson’s correlations were used to determine if any relationships were present between sleep, changes in performance, markers of subjective wellness and training load. Correlations were interpreted using thresholds of <0.1, tigraphy . Particiectively . An effe effects .p ≥ 0.05) between sleep and wellness measures or performance markers, with all r-values in the trivial–moderate range, as seen in n = 15) and HIGH: >7 h 30 min sleep per night, n = 14) for all remaining analyses. Differences observed between HIGH and LOW sleep groups for average nightly sleep duration, weekly total sleep duration and overall total sleep duration are shown in The Pearson’s correlation correlational analyses revealed no significant correlations  = 7.89, p < 0.01 and Bronco performance, F = 7.60, p = 0.01, but not for any other performance measure (p > 0.05). The HIGH group displayed significantly greater reductions in skinfold measurements at Week 3  = 6.39, p = 0.01 (p = 0.03) and Q15  of the ASBQ in the HIGH sleep group. No significant group x time interactions were observed between groups for any wellness measures. However, a significant increase (p < 0.01) in fatigue was observed from Week 1 to Week 3 across the entire cohort.Sleep questionnaire analysis revealed a significant group x time interaction for ASBQ global scores, p = 0.01  with thep = 0.01 .. The WiThe aim of this study was to investigate the relationship between sleep duration and changes in physical performance throughout a 5-week preseason training phase in elite rugby union athletes. Key findings indicate that when the group stratified into two quantile groups, those obtaining greater amounts of sleep (>7 h 30 min) resulted in positive changes in aerobic fitness and body composition compared to those who obtained < 7 h 30 min of sleep per night. Additionally, we observed a decrease in weekly sleep duration and increased fatigue from Week 1 to Week 3 for both groups, suggesting sleep duration is negatively impacted during a preseason phase of training.small improvement of 4.3% in reaction time performance in elite rugby union athletes. Lastly, Mah and colleagues [While this is the first study to evaluate the link between sleep duration and performance metrics during a preseason phase in elite rugby union athletes, previous research has evaluated the efficiency of sleep extension on athletic performance ,45,46,472 max and work rate [Although speculative, the positive effects observed in Bronco performance may be related to an increased amount of slow-wave sleep (SWS). Increased sleep duration results in an increased amount of SWS ; this isork rate ,51. GiveFurther, higher growth hormone levels have been linked to elevated levels of resting metabolic rate (RMR) . RMR is No differences were observed between groups for strength or speed throughout the 3-week training phase. Additionally, results revealed that both groups displayed decreases in strength markers at week 3. The observed lack of change in speed performance is contrary to previous results by Mah and colleagues , who demThe decrease in sleep duration of 1 h 13 min less sleep per night from Week 1 to Week 3 observed for both groups is likely due to increased fatigue. Thornton et al. . previouThe current study highlighted a moderate improvement in the HIGH sleep group for the ASBQ questionnaire global score, which suggests that the HIGH sleep group displayed better sleep behaviours during the 3 weeks than the month leading up to the study. No significant differences were seen for either sleep group for PSQI global scores. It should be noted that the current study’s sleep groups displayed global scores of 5 and above at baseline and after Week 3. A global score of 5 or greater has been suggested to indicate that an individual has moderate sleep difficulties in at least 3 areas or severe difficulties in at least 2 areas . TherefoGiven the restraints of conducting research on professional athletes, we acknowledge several limitations in the current study. The study was designed to have minimal impact on training, performance and each athlete’s regular routines; therefore, we could only employ an observational study design, with retrospective analysis. Thus, numerous areas were difficult to control. Firstly, we were unable to control training during the 2-week maintenance period , nor were we able to monitor sleep during this period. Athletes were provided with a training program to complete throughout the 2-weeks; however, we had no control over how strictly the athletes adhered to the program. Secondly, we were unable to control numerous aspects of nutrition. Whilst athletes were provided with the same breakfast and lunch during the preseason phase (Weeks 1–3), we were unable to control caloric intake for all meals. An addition to strengthen the study design would have been to measure all physical performance measures at every time point , allowing comparisons to be drawn across the 5 weeks for all physical performance measures. Future research should employ an experimental design investigating the differences in physical performance outcomes between a short sleep duration group  and a long sleep duration group  during a period of intensified training. Additionally, researchers should investigate ways of preventing the decrement in sleep duration seen during intensified periods of training .The current investigation is the first to show a relationship between sleep duration and changes in physical performance in an elite rugby union environment during a preseason phase of training. The results suggest that, when grouped for sleep duration, athletes who obtained more sleep displayed greater positive changes in aerobic fitness and body composition than athletes who slept less. Additionally, results showed that sleep duration can be negatively affected during the preseason, which is likely due to an increase in training volume and an increase in fatigue. The current study provides some promising results concerning the effects of sleep duration on physical performance throughout the preseason in elite rugby union athletes and therefore warrants further investigation.


Background: Physiologically based pharmacokinetic (PBPK) modeling and simulating may be a powerful tool in predicting drug behaviors in specific populations. It is a mathematical model that relates the pharmacokinetic (PK) profile of a compound with human anatomical characteristics, physiological characteristics, and biochemical parameters. Predictions using PBPK models offer a promising way to guide drug development and can be used to optimize clinical dosing regimens. However, PK data of new drugs in the pediatric population are too limited to guide clinical therapy, which may lead to frequent adverse events or insufficient efficacy for pediatric patients, particularly in neonates and infants.Objective: The objective of this study was to establish a virtual Chinese pediatric population based on the physiological parameters of Chinese children that could be utilized in PBPK models.Methods: A Chinese pediatric PBPK model was developed in Simcyp Simulator by collecting published Chinese pediatric physiological and anthropometric data to use as system parameters. This pediatric population model was then evaluated in the Chinese pediatric population by predicting the pharmacokinetic characteristics of four probe drugs: theophylline (major CYP1A2 substrate), fentanyl (major CYP3A4 substrate), vancomycin, and ceftazidime .Results: The predicted maximum concentration (Cmax), area under the curve of concentration-time (AUC), and clearance (CL) for theophylline (CYP1A2 metabolism pathway) and fentanyl (CYP3A4 metabolism pathway) were within two folds of the observed data. For drugs mainly eliminated by renal clearance (vancomycin and ceftazidime) in the Chinese pediatric population, the ratio of prediction to observation for major PK parameters was within a 2-fold error range.Conclusion: The model is a supplement to the previous Chinese population PBPK model. We anticipate the model to be a better representative of the pediatric Chinese population for drugs PK, offering greater clinical precision for medication given to the pediatric population, ultimately advancing clinical development of pediatric drugs. We can refine this model further by collecting more physiological parameters of Chinese children. In this study, we developed a Chinese pediatric PBPK model to predict in vivo drug behavior in the Chinese pediatric population. We anticipate the model to assist in pediatric drug development and rational clinical medication.Pediatric rational medication and drug development have received much attention these years. However, the The rate of adverse drug reactions (ADR) reported in Chinese children and newborns are two and four times higher than that of adults . One of The physiologically based pharmacokinetic (PBPK) model could integrate drug and physiological information and clinical trial design to make predictions on PK in special populations, including pediatrics. For example, Abduljalil et al. developed a preterm PBPK model by collecting published information on preterm developmental physiology including demographics, hematocrit, protein binding data, tissue volume, and enzyme ontogeny data . The modin vivo PK data. We anticipate that this model can assist in Chinese pediatric drug development and rational clinical medication.Therefore, the objective of this study is to develop a Chinese pediatric population model by incorporating the physiological parameters specific to the Chinese pediatric population with reference to the Caucasian population model. The new Chinese pediatric population model will be used in Simcyp Simulator to predict PK for CYP1A2 probe substrate (theophylline), CYP3A4 probe substrate (fentanyl) and renal-eliminated drugs (vancomycin and ceftazidime) in Chinese ®  to generate a Chinese pediatric population model. Next, a literature search for clinical PK data in the Chinese pediatric population was carried out with specific inclusion and exclusion criteria. Based on the availability of the PK data and specificity of the substrate, four probe drugs, predominantly eliminated by three different pathways , were selected. Drug models were then developed for these probe drugs in Simcyp and were verified using PK data from the Caucasian adults, Caucasian children, and Chinese adults. Finally, the drug models of these probe drugs, in combination with the newly developed Chinese pediatric population model, were applied to simulate PK in the Chinese pediatric population to evaluate the population model. The prediction performance of the models was evaluated by comparing the simulated maximum concentration (Cmax), clearance (CL), and area under the concentration-time curve (AUC) with the observed value.https://www.ncbi.nlm.nih.gov/pubmed/). Keywords used in the search included “Chinese” in all combinations with “demographic”, “height”, “weight”, “organ weight ”, “cardiac output”, “body surface area (BSA)”, “yellow marrow and blood volume”, “tissue compositions ”, “glomerular filtration rate (GFR)”, “biologic parameters ”, “ontogeny ”, and “skin parameters ”. Data were included in our study if they met the following criteria: 1) the data came from Chinese children aged 0–18 years old; 2) male and female statistics were separate (in specific parameters).A systematic literature search was carried out for various physiological and demographic data of Chinese children using PubMed (The Chinese pediatric population model was developed in Simcyp based on the inbuilt Northern European Caucasian (NEC) pediatric population model. The physiological and demographic data in the Chinese pediatric population collected and included in our study were used to recalibrate the corresponding parameters in the NEC pediatric population model. All the parameters were modeled against age, height, weight, or BSA using inbuilt equations of the NEC pediatric population. By integrating the equations of physiological and anthropometric parameters into the Simcyp simulator, the equations of height and weight were set as inputs in “Population-Demographic-User-defined HT and WT relationship”. Equations of organ weight were modified in “Population-Paediatric-Tissue Volume-Define Tissue Volume”. The cardiac output function was defined in “Population-Paediatric-Blood Flows-Define Blood Flow”. For the rest of the physiological parameters, because there was very limited Chinese pediatric data available, the default value of the inbuilt NEC pediatric population model was used with the assumption that they were the same between Chinese and Caucasians.https://www.ncbi.nlm.nih.gov/pubmed/), Embase (https://www.embase.com), and CNKI (https://www.cnki.net/). The literature search strategy was illustrated in The candidate probe drugs were selected based on the following considerations: a sensitive substrate of CYP1A2 or CYP3A4, namely more than 80% of the drug was metabolized and more than 70% metabolism was contributed by CYP1A2 or CYP3A4; or more than 70% of the drug was eliminated by renal filtration. A literature search was then conducted to collect PK data of candidate probe drugs in the Chinese pediatrics population at different ages using PubMed , and the keyword related to the targeted race and region . Deadline for inclusion in the literature is 2019. The related drugs were included in the probe drug candidate list.in vivo  that were mainly eliminated by three respective pathways  were selected. Theophylline is one of the substrate drugs of CYP1A2, and CYP1A2 contributed 75% of its metabolism . Theophyin vivo . Fentanyin vivo . Ceftaziin vivo . The clemax, CL, AUC, and the concentration-time profile were included in our study. Inclusion criteria were: 1) study subjects were of the Chinese pediatric population, included neonates, infants, children, and adolescents with normal cardiac, hepatic and renal functions 2) sample size was larger than three 3) dosing regimen was reported 4) the drug was administered intravenously 5) there was no comedication in the study and 6) drug concentration-time profiles or at least one of the PK parameter was reported. Exclusion criteria were: 1) participants included preterm infants 2) the age range of subjects was not reported clearly 3) dosing regimen varied from individual to individual and the mean dosage was not reported.There were 57 studies related to these four probe drugs. If the study met the following inclusion criteria but not exclusion criteria, the PK data such as CSeventeen studies were included according to inclusion criteria and nine studies were excluded. Four of them were excluded for meeting criteria one, two of them were excluded for criteria two, and three of them were excluded for criteria three. Finally, eight studies met our criteria and were included in our research.https://www.drugbank.ca/), SciFinder (https://scifinder.cas.org), and literature. The collected data were then integrated into a PBPK model using Simcyp Simulator.Theophylline compound file from Simcyp V20 library was used to carry out simulations. Fentanyl, ceftazidime and vancomycin PBPK models were self-developed compound files. The physicochemical properties, as well as the drug metabolism and pharmacokinetics (DMPK) characteristics of the probe drugs that need to be self-developed, were obtained from the Drug Bank  model was used in oral administration cases. The PBPK model parameters of probe drugs were presented in max, CL, AUC) and the concentration-time profile were included in drug model verification if the study met the following inclusion criteria: 1) study subjects were of the Chinese/Caucasian adult population or the Caucasian pediatric population with normal cardiac, hepatic and renal functions; 2) sample size was larger than three; 3) dosing regimen was reported; 4) there was no comedication in the study and 5) drug concentration-time profiles or at least one of the PK parameters was reported. If the ratio of predicted and observed mean values for major PK parameters was within the range of 0.5–2.0, these drug models were deemed fit to be verified.The drug models for all four probe drugs were verified in a stepwise manner using clinical PK data from Caucasian adults, Caucasian children, and Chinese adults. The major PK parameters  of each probe drug were compared with the observed values. If the ratio of the predicted value was within a 2 folds difference to the observed one, the population model was considered acceptable.The verified drug models of four probe drugs in combination with the newly developed Chinese pediatric population model were applied to predict drug PK in the Chinese pediatric population. The simulation was conducted by mimicking trial designs in the literature. The predicted drug concentrations vs. the observed ones were examined. Furthermore, the predicted PK parameters  in the Chinese pediatric population were collected in our study but there is still a lack of information on enzyme and transporter’s ontogeny, GFR, some of the tissue volumes , tissue compositions, biologic parameters , and skin parameters .R2) > 0.98. The height and weight data were collected from two pieces of literature covering Chinese children aged 0–18 years old . The carTo evaluate the capability of the developed pediatric population model to describe the ontogeny of height, weight, cardiac output and organ size of Chinese children, a simulation of 4000 virtual Chinese pediatric population was performed using Simcyp simulator . As shown in The drug model for all four probe drugs was verified with PK data of Caucasian adults, the Caucasian pediatric population, and the Chinese adult population from various studies. As shown in max, CL, AUC) from various studies were within the range of 0.5–2.0. For ceftazidime and vancomycin, which were eliminated via renal elimination pathway, the ratio of predicted vs. observed CL from the single study of ceftazidime was 0.88, and the ratio of vacomycin was 1.12 and 0.65. The ratio of predicted vs. observed AUC for vancomycin was 1.76 and 1.17, respectively. No Cmax was reported for both drugs. The verified drug model of these four probe drugs in combination with the developed Chinese pediatric population model was applied to predict drug concentration in the Chinese pediatric population. As shown in In the past decades, the PBPK model has been applied to predict pediatric drug concentrations during drug development to support dosing rationalization in the pediatric clinical trial. For example, the PBPK model was used to set a starting dose for Eribulin in children and adolescents aged 6–18 years old . HoweverIn our study, depending on the availability of the Chinese pediatric PK data, four probe drugs that were eliminated either by CYP1A2 metabolism (theophylline), CYP3A4 metabolism (fentanyl) or by renal filtration (ceftazidime and vancomycin) were selected to evaluate the prediction performance of this pediatric population model. It should be noted that CYP1A2 and CYP3A4 are the major P450 enzyme in eliminating several drugs in clinical  ChoonaraOur population model was developed based on the inbuilt NEC pediatric population model in Simcyp, with height, weight, cardiac output, and tissue volumes  recalibrated with Chinese pediatrics data. To confirm whether there was an ethnic difference between Chinese children and Caucasian children, we performed the same simulation using the NEC pediatric population model. However, the Chinese pediatric model showed no significant improvement in predicting the CL of CYP3A4-metabolized and renal-eliminated drugs, as shown in Although great efforts have been made to search for physiological data extensively in Chinese pediatric patients, there is still a lack of information on many physiological parameters such as body surface area (BSA), glomerular filtration rate (GFR), and especially on the ontogeny of a cytochrome P450 enzyme. Microsomal protein per gram of liver (MPPGL) and CYP enzyme ontogeny is critical in order to correctly predict the exposure of drug with metabolism as the major elimination pathway. Hence in our study, we assumed the Chinese pediatric population shares the same CYP1A2 and CYP3A4 ontogeny as the Caucasian population, which may not necessarily true. More clinical trials need to be carried out in order to prove our assumption. In addition, all the PK data of the Chinese pediatric population were from patients. Pathological factors would influence the accuracy of a prediction.In this study, a preliminarily physiologically based pharmacokinetic model based on Chinese pediatric physiological characteristics was developed. The model was verified in the Chinese pediatric population by predicting PK behavior of theophylline, fentanyl, ceftazidime, and vancomycin which are eliminated either through CYP1A2, CYP3A4 and renal pathway. In comparison of the observed PK data, our model successfully predicted the PK profile of these drugs within a 2-fold error range. However, it warrants further study to improve this population model.


Halophila stipulacea, by investigating the effect of in situ nutrient addition during an unusually warm summer over a 6-month period. We measured a suite of different morphological and biochemical community metrics and individual plant traits from two different sites with contrasting levels of eutrophication history before and after in situ fertilization in the Gulf of Aqaba. Nutrient stress combined with summer temperatures that surpassed the threshold for optimal growth negatively affected seagrass plants from South Beach (SB), an oligotrophic marine protected area, while H. stipulacea populations from North Beach (NB), a eutrophic and anthropogenically impacted area, benefited from the additional nutrient input. Lower aboveground (AG) and belowground (BG) biomass, reduced Leaf Area Index (LAI), smaller internodal distances, high sexual reproductive effort and the increasing occurrence of apical shoots in seagrasses from SB sites indicated that the plants were under stress and not growing under optimal conditions. Moreover, AG and BG biomass and internodal distances decreased further with the addition of fertilizer in SB sites. Results presented here highlight the fact that H. stipulacea is one of the most tolerant and plastic seagrass species. Our study further demonstrates that the effects of fertilization differ significantly between meadows that are growing exposed to different levels of anthropogenic pressures. Thus, the meadow’s “history” affects it resilience and response to further stress. Our results suggest that monitoring efforts on H. stipulacea populations in its native range should focus especially on carbohydrate reserves in leaves and rhizomes, LAI, internodal length and percentage of apical shoots as suitable warning indicators for nutrient stress in this seagrass species to minimize future impacts on these valuable ecosystems.Eutrophication is one of the main threats to seagrass meadows, but there is limited knowledge on the interactive effects of nutrients under a changing climate, particularly for tropical seagrass species. This study aimed to detect the onset of stress in the tropical seagrass,  Seagrass meadows are becoming increasingly threatened, as their distribution along coastal areas makes them especially vulnerable toward local anthropogenic pressures . SeagrasHalophila stipulacea (Forsskål) Ascherson is a tropical seagrass species native to the Indian Ocean, Red Sea and Persian Gulf , shows a high photo-physiological plasticity and has been found to thrive under eutrophic conditions, even in sulfidic sediments , northern Red Sea, have mainly focused on local coral reefs e.g., , 2019, e meadows . H. stip the GoA , where i the GoA . These s the GoA . However the GoA , 2020.–1  alone has already been shown to be insufficient for protecting seagrass meadows  in the northern GoA are increasing at rates of 0.254 ± 0.058°C decade–1 , slightl–1  and morphological  and biochemical (tissue nutrient content and carbohydrates) plant traits identified as potential stress response indicators  and two sites in a marine protected area with relatively low anthropogenic pressures (South Beach – SB) along the western coast of the northern Gulf of Aqaba , entailing a relatively high Secchi disk depth and relatively small seagrass leaves (Kd) determined by measurements of PAR at the water surface and sediment depth, is much higher than in the South Beach site (Sparus aurata) were cultivated in cages in this area for approximately 20 years, which led to nutrient enrichment of the water column and underlying sediments, and a die-off of the underlying seagrass meadows (15N) and higher levels of NH4+ in seawater in the NB than in the SB alongside a higher annual signal of δ15N in H. stipulacea plants growing in the more impacted site (NB) compared with the more pristine site (SB). These results indicate a potential anthropogenic input of N in the seawater, which is reflected in the plant content in the impacted site (NB), and together with the light measurements and leaf morphometrics . The fertilizer contained nitrogenous compounds that were composed of nitrate (6.3%), ammonia (8.2%) and urea (4.5%). Fertilizer pellets were filled in 0.5 m long PVC tubes, which were hammered approximately 20 cm into the sediment, delivering nutrients to the below-ground and above-ground tissues (Six circular plots with a diameter of 2m were established at each one of the four sites (S1–S4), at least 10 m apart from each other. Three of these plots were used as control, while the other three were enriched with slow-release fertilizer pellets  and stored on ice. Samples were frozen at −20°C upon arrival to the laboratory on the same day of sampling and stored for further analyses. Nutrient measurements for samples taken in July were performed spectrophotometrically with a TECAN plate reader  according to 2–, NOx (NO3– and NO2–), NH4+, PO43–, respectively. The NOx (NO3– and NO2–) and NH4+ concentrations sum up to dissolved inorganic nitrogen (DIN). The coefficient of variation was always <3.4%. The lower nutrient concentrations from samples taken during December 2019 were analyzed using a continuous flow injection analyzing system . The detection limits were 0.043, 0.094, 0.290, and 0.042 μM for NO2–, NOx (NO3– and NO2–), NH4+, and PO43–, respectively. The coefficient of variation was always <3.4%.Water samples were taken with acid rinsed plastic syringes (approximately 30 mL) at around 10 m depth above the seagrass canopy of each plot on each site for nutrient analysis of the surrounding water. Porewater samples were taken by placing a syringe 5 cm deep into the sediment and carefully drawing out water from interstitial spaces of the sediment to determine porewater nutrient concentrations. Once on shore, water samples were immediately filtered into HDPE vials using sterile syringe filters  bottle. Collected seawater was immediately stored in the dark on ice. Environmental parameters, such as salinity, temperature, oxygen and pH were measured directly after collection with a multi parameter probe (WTW Multiprobe). Moreover, temperature loggers (HOBO Water Temp Pro v2) were fixed to the main pole of plots 1 and 6 in the seagrass meadows (ca. 10 m water depth) to record the water temperatures hourly during the entire time of the experiment.® GF/F filters on return to the laboratories of the Interuniversity Institute for Marine Sciences in Eilat (IUI). Filters for chlorophyll a (Chl a) analysis were immediately stored at −80°C, filters for organic carbon (Corg), total carbon (C) and nitrogen (N) content were frozen at −20°C until further analysis at the Leibniz Centre for Tropical Marine Research (ZMT) in Germany. Filters for suspended particulate matter (SPM) determination were weighed prior to filtering. Subsequently, filters were dried at 40°C for 48 h and the dried filters were weighed again. Filters for Corg, C and N content were divided in 4 equal pieces and one quarter was used for each analysis. For Chl a analysis, filters were analyzed and extracted in 80°C hot ethanol, following a concentrations were determined with a TD10AU-Fluorometer (Turner Designs). The detection limit of this method is 0.002 μg l–1.A defined volume of water was vacuum-filtered onto pre-combusted  WhatmanPercent cover was measured by randomly placing a 25 cm × 25 cm quadrat in each plot and taking a picture with an underwater camera. Pictures were analyzed with Coral Point Count for Excel CPCe; . 100 poi2. Shoots that were examined for the presence of male or female reproductive structures were counted separately and the flowering percentage  was calculated (–2) and the above/below-ground biomass ratios (AG:BG).A biomass corer was used to sample above ground (AG) and below ground (BG) seagrass biomass in July and December 2019 at SB and NB sites. Samples were taken with an open- ended, standard PVC drainpipe with a diameter of 12.5 cm and a length of 0.5 m. In each plot, biomass cores were taken by twisting and hammering the corer approximately 10–15 cm into the sediment. The contents of the biomass cores were transferred underwater into diving mesh-bags and sediment was removed carefully underwater. Upon arrival at the laboratory facilities of the IUI in Eilat, biomass samples were carefully cleaned and all sand, sediment particles and epiphytes were carefully removed. Plant material from the core samples was separated into AG (leaves) and BG (rhizomes and roots) plant compartments. Shoot density was determined by counting the total number of shoots in each biomass core sample and then normalizing these counts per mlculated . AG and Additionally, leaf and rhizome samples were taken from each biomass core to evaluate shoot morphology. Internodal distance of rhizome was measured for each plot (9 subsamples per plot). From each subsample, intact and representative leaf samples including the smaller, youngest shoot (3–14) were used to measure leaf length, leaf width, leaf area and leaf area index (LAI). The leaves were either scanned  or photographed. Leaf descriptors were measured with ImageJ software , as well as total non-structural carbohydrate (NSC) reserves such as starch and sugars.Several ramets were randomly collected at different locations within each plot. AG and BG tissues were separated and immediately frozen at −20°C until further analysis for AG and BG nutrient content, stable carbon and nitrogen isotope analyses  with Acetanilid 5 (Hexatech) used as standard. Measurement accuracy was ensured by repeatedly measuring standards with known C and N concentrations . Carbon and nitrogen contents were expressed as a percentage of dry weight and the values were used to calculate the 13C/δ 12C) and nitrogen (δ15N/δ14N) in samples were analyzed using a Finnigan Delta Plus mass spectrometer coupled with a Flash EA 1112 elemental analyzer. Results of isotopic composition in samples were expressed as following:Stable isotopic ratios of carbon  and USGS 24 and NBS22 (carbon) from the International Atomic Energy Agency were used for calibration. The precision of the measurements was <0.06‰ for both C and N isotopes. All δ13C and δ15N values were normalized to the international standards of wheat flour  and high organic sediment .where 2SO4) for 1.5 h at 100°C, the remaining polysaccharides were broken down to glucose molecules. Starch and sugar contents in the extracts were determined spectrophotometrically (620 nm) using an anthrone-sulphuric acid assay with a F200-Pro TECAN plate reader. Carbohydrate concentrations were quantified as sucrose equivalents  was suspended in 1.5 mL Milli-Q water and soluble sugars were extracted from the ground dry tissues by vortexing and shaking for 15 min. Samples were centrifuged  and the supernatant was used for soluble sugar determination via the Anthrone Assay . Starch ivalents  using suFrom each plot of the four sites, surface layer sediments were collected in July and December 2019 in polystyrene Biological Specimen Containers. After collection, samples were stored at −20°C until further analysis.Upon arrival at the ZMT, samples were dried at 60°C for 48 h and dry sediments were subsequently ground with a ball mill. Ground samples were then acidified with 37% HCl vapor to remove carbonates and stable isotopic composition measured as described under Section 2.7.in situ fertilization treatment started in July, it was assumed there was no nutrient treatment effect in July. We used two linear mixed effects models (LMM). One to determine if there were seasonal effects over the time of the experiment (1). For this analysis, we only looked for changes in the control plots over time and added site as a random effect. The second linear mixed effects model was performed to look at the differences between the conditions of the sites (high [NB] and low [SB] impacted sites) in July (2) and December (3), the effect of treatment- differences between fertilized and control plots (4) as well as their possible interactions (5). We performed both linear mixed effects models using the lme4 function in R  was performed to look for significant differences between low (S1 + S2) and high (S3 + S4) impacted sites.Environmental parameters were analyzed separately for July and December 2019 with the ANOVA aov or Kruskal–Wallis function in R, and a subsequent paired Salinity ranged between 40.4 and 40.6 psu and pH ranged between 8.22 and 8.23 with no significant differences between sites . Daily aa and SPM concentrations were higher in the NB sites  and N compounds  as well as P in the water column , confirming that our fertilization was successful.There were no significant differences in nutrient concentrations in the water column and the porewater between SB and NB sites . However13C isotopic signatures of sediment contents between low (SB) and high (NB) impacted sites in July and December 2019  were even two to three times higher than the SB sites  biomass was significantly higher in NB meadows in July 2019 . However2 was not significantly different in seagrass meadows from SB and NB sites in July 2019, but decreased significantly in both locations by December 2019 (Low impacted sites (SB) had a higher percentage of apical shoots in their shoot population in July 2019 . The number 2019 . Howeverber 2019 .Seagrass plants growing in meadows at SB had significantly smaller and thinner leaves as well as a smaller leaf area and LAI than plants growing in NB in July and in December 2019 . The addin situ fertilization treatment  and sugar concentrations in their rhizomes and roots. Moreover, plants in the NB sites were related with increased AG and BG biomass, higher AG:BG ratios and were enriched in δ15N isotopic contents in their rhizomes as well as in their sediments.Principal component analysis were performed for the two different sampling months, July and December. In July, the first two components of the PCA explained roughly 50% of the variability . Seagras13C isotopic contents in their sediments. SB plants from control plots were also characterized by higher starch concentrations in their rhizomes and higher δ15N isotopic contents in their rhizomes, while SB plants in fertilized plots were associated with a higher percentage of apical shoots in their shoot populations and higher porewater P and N compounds.In December, SB and NB sites were clearly separated as well as the control and the fertilized plots, although these latter plots showed some slight overlap . The firOn the other hand, plants from fertilized plots in the NB sites, were positively correlated with increased AG and BG biomass and higher P content in their rhizome and roots as well as larger leaf length, width and LAI. Seagrasses from fertilized plots in the impacted NB sites were positively correlated with higher AG:BG ratios as well as increased shoot densities and percent cover. Seagrasses from control plots in both sites were positively correlated with higher rhizome and root tissue nutrient content (%C and %N).H. stipulacea in its native range (northern GoA). It is likely that the effects of fertilization would have resulted in more significant changes in our chosen seagrass indicators if further temporal and spatial replication would have been possible. Despite these limitations, our approach was successful for highlighting the different responses to eutrophication among H. stipulacea meadows growing under different levels of anthropogenic pressures . The in situ fertilization treatment, simulating chronic eutrophication had mainly negative effects on seagrass plants from the SB sites, a marine protected area, while H. stipulacea populations from the anthropogenically impacted NB sites benefited for the most part from the additional nutrient input .The highest temperatures were observed in August with daily average values of 27.88 ± 0.07°C SE at the NB sites and 28.33 ± 0.07°C SE at the SB sites at around 10 m water depth. 2) compared with the leaf sizes of plants growing in the clearer waters of the SB sites (1.19 ± 0.05 cm2)  . Seagras.05 cm2) . This ad.05 cm2) . Indeed,The addition of fertilizer had different effects on seagrass meadows from low and high impacted sites. Percent cover and shoot density significantly increased in both sites as a response to the additional nutrient input compared to control plots. However, while the additional nutrient input resulted in a significantly enhanced biomass production in plants from the NB, there was only a small trend of an increase in biomass of seagrasses growing in fertilized plots in the SB sites.Internodal distances of plants growing in the SB sites were smaller than the ones of seagrasses from the NB, and decreased further with fertilization, probably as a response to nutrient stress. Shorter internodal distances have been shown to be a common response to environmental stress and might serve as a proxy of increased shoot and/or root densities . This isMoreover, seagrasses from SB had smaller and thinner leaves and a smaller LAI. LAI combines individual plant traits (leaf area and number of leaves) with population level metrics (shoot density) that have been shown to decrease in response to anthropogenic pressures . LAI was15N in both their leaf and rhizome tissues  5 months after the summer heat peak in urbidity  and the measured . Under lmeasured . Additiomeasured  will likmeasured , therebyreserves . In addireserves . These preserves  and our H. stipulacea populations from NB experienced temperature stress and reduced fitness after a 2 weeks exposure to temperatures of beyond the possible thermal threshold of 29°C, resulting in reduced growth rates, leaf sizes and the loss of shoots , changes in structural and demographic/population-level traits occurred also as a response to different seasons. Nutrient enrichment, as shown in our study, combined with summer temperatures that surpassed the threshold for optimal growth, had only negative effects on performance of plants from the more pristine SB sites, located within a marine protected area. Seagrass plants growing in the protected SB sites showed first signs of stress under the simulated eutrophication and high summer temperatures by reduction in NSC, sugar (decline of 37%) and starch (decline of 15%) contents of their rhizomes. Furthermore, plants in SB demonstrated declining internodal distances and an increase in the frequency of their apical shoots within their shoot population, which are confirmed indicators for stress in this and other seagrass species . The fact that NB populations perform even better under the additional nutrient inputs further supports previous suggestions of H. stipulacea living in these vast NB meadows is actually N limited.Known for its ability to grow in a wide range of light conditions , salinities, temperatures, and substrates [reviewed by asticity . This pl species . Moreovea concentrations in comparison to SB sites  vs. invasive H. stipulacea populations from Greece and Cyprus  in both its historical (Mediterranean) and the new invasive (Caribbean) habitats.Thus, while future ocean warming and the rising frequency of heat waves and high cultural eutrophication will most likely lead to further losses in  meadows , emerginipulacea . Indeed,ipulacea . The resoceanica . Our resThe original contributions generated for this study are included in the article/SH, GW, HR, and MT conceived and designed the experiments. EB and MiS contributed in designing the experiments. MiS, MaS, and SB assisted with the fieldwork and during sampling campaigns. SH performed the field experiments, developed methodologies, and performed all the other laboratory analyses together with SB. EB did the statistical analysis. EB and MaS did the graphs and worked on the manuscript. All authors wrote and reviewed the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Postoperative cognitive dysfunction (POCD) is one of the most common. Neuroprotective effects of dexmedetomidine (DEX) are reported in previous studies but evidence regarding the POCD is still unclear. In order to gain latest evidence, the present study analyzes the outcomes of randomized controlled trials (RCTs) which utilized DEX with general anaesthesia perioperatively.Four online databases  were used to find relevant RCTs to conduct systematic analysis. All studies comparing the incidence of POCD or MMSE score between the DEX group and the placebo or comparator group in patients undergoing general anaesthetic surgery were eligible for inclusion. Based on the inclusion and exclusion criteria, the studies were selected. This meta-analysis was performed using odds ratios (ORs) with 95% confidence intervals (CIs) for dichotomous data and standardized mean difference (SMD) and 95% CIs for continuous data as effective measures.In total of 21 studies were included in this meta-analysis. The results showed that the incidence of POCD in DEX group was significantly lower than the control group on the first ,third  and seventh  postoperative days; the MMSE scores in DEX group were higher than the control group on the first , third and seventh  postoperative days.Intraoperative DEX use can ameliorate the POCD of patients who received surgical operations under general anesthesia, and effectively reduce the incidence of POCD and improve MMSE score. Postoperative cognitive dysfunction (POCD) is one of the most common complications affecting the central nervous system after general anaesthesia and surgery, especially in elderly patients, which is characterized by short-term cognitive decline and includes memory, mood, confusion, and sleep disorders . Its cliDexmedetomidine (DEX) is a effective and highly selective α2-adrenergic receptor agonist and acts as a multifunctional drug in the treatment of various human diseases , 18. A pSeveral basic researches with animal models have reported that DEX provide neuroprotective effects and improve cognitive function following surgery , 23. In Two review authors (Jianli Liand Qifan Yin) independently searched PubMed, Embase, the Cochrane Library, China Academic Journals full-text database (CNKI) to find relevant RCTs without any language restrictions. All studies comparing the incidence of POCD or MMSE score between the DEX group and the placebo or comparator group in patients undergoing general anaesthetic surgery were eligible for inclusion. The search was updated to Dec 31, 2020. The main search terms included: (“dexmedetomidine” OR “Dex”) AND (“postoperative cognitive dysfunction” OR “POCD” OR “postoperative cognitive impairment”). The reference list was also checked for relevant articles.Studies were included according to the following criteria: (1) all of the enrolled studies were randomized controlled trials (RCTs); (2) adult patients undergoing surgery under general anaesthesia; (3) the experimental group received a single or continuously-administered intravenous dose of intraoperative DEX; the control groups received an intravenous injection of placebo or comparator; (4) the main outcomes in enrolled studies were the incidence of POCD, postoperative MMSE score; (5) all of included studies contained original data sufficient for meta-analysis.The exclusion criteria were: (1) abstracts, letters, case reports, reviews, duplicates or nonclinical studies; (2) studied patients who underwent local or spinal-epidural anesthesia; (3) patients administered DEX compared with other sedative agents  were excluded; (4) articles with poor quality, Jadad score ≤ 3 points, were excluded; (5) studies with incomplete information or data, and articles for which we could not obtain the full text.Two researchers (Xuejiao Xun and Jinhua He) independently screened the literature, extracted data and evaluated the methodologic quality of the studies identified. If disagreement occurred, a consensus was reached after discussion with a third author. All the corresponding data including the name of first author, publication year, country, age, number of participants, type of surgery, administrations for patients, incidences of POCD, MMSE score were extracted from each selected study. We evaluated methodologic quality of the RCTs enrolled using a Jadad scale. Evaluation included randomization, allocation concealment, and blinding of implementers and participants. Studies awarded with greater than three scores were considered to be of acceptable quality. The information and data of enrolled studies were entered in a standard data extraction form. Table 2 test. A fixed effects model was performed to conduct the meta-analysis if no heterogeneity (P > 0.1 or I2 < 50.0%) was found among the studies. If significant heterogeneity (P < 0.1 or I2 ≥ 50.0%) was found, a random effects model was applied for the meta-analysis. Bias or publication bias was evaluated as quality using funnel plots, Egger’s and Begg’s tests. A value of P < 0.05 was considered statistically significant.We used Review Manager 5.3 to conduct the meta-analysis. For dichotomous data, odds ratios (ORs) with 95% confidence intervals (CIs) were used to express effect-size, while standardized mean difference (SMD) and 95% CIs were used for continuous data. First, we conducted a heterogeneity test on included studies using the IIn the initial search, a total of 1,527 potential articles were identified. After the initial screening of abstracts and titles, 1,448 articles were excluded based on the inclusion and exclusion criteria. The remaining 78 articles were collected for full text review, after the secondary screening and carefully inspection of these articles, 21 studies including 2,902 patients published between 2012 and 2020 were eventually selected for final analysis. Another 57 articles were excluded; 14 studies reported animal trials or basic experiments, 13 studies demonstrated DEX combined with other sedative agents , 8 studies described POCD in patients with local or spinal anesthesia, the quality of 8 studies with Jadad score ≤ 3 points were poor, 5 articles described POCD in adolescents, 5 studies did not provide complete data, 4 studies were reviews and meta-analyses. The detail processes of study screening and selection were shown in the flow diagram  and saline/comparator (1552). All included studies published between 2012 and 2020. Among them, 19 studies were from China, one study was performed in The United States and Africa, respectively. With regard to the incidence of POCD in our meta-analysis, 14 studies with 1,639 patients demonstrated the relationship between DEX and the incidence of POCD on the first day after surgery. 9 articles with 1,328 patients illustrated the association between DEX and the incidence of POCD on the third day after surgery. 5 literatures with 968 patients shown the relation between DEX and the incidence of POCD on the seventh day after surgery. Concerning the relationship between DEX and MMSE score, 9 studies with 800 patients described the relationship between DEX and the MMSE score on the first day after surgery. 8 articles with 976 patients reported the association between DEX and the MMSE score on the third day after surgery. 4 researches with 628 patients reported the association between DEX and the MMSE score on the seventh day after surgery. Dosage of DEX was in the range of 0.5 to 1.5 µg/kg followed by continuous infusion at a rate of 0.15 to 0.80 µg/kg/h. The majority of included studies investigated the elderly. The basic characteristics of each of the enrolled studies, including publication year, country, average age, number of cases, surgical type, administrations for patients, incidence of POCD, MMSE and Jadad score, were shown in Table P = 0.009, I2 = 52%, Fig. 14 studies including 1639 patients reported the relationship between DEX and the incidence of POCD on the first day after surgery. There was an obvious heterogeneity between the studies (2 = 0%, P = 0.49, Fig. 9 studies including 1328 patients elaborated the association between DEX and the incidence of POCD on the third postoperative day. Considering no obvious heterogeneity between the studies (I2 = 47%, P = 0.11, Fig. 5literatures including 968 patients reported the incidence of POCD on the seventh day after surgery. On account of the little heterogeneity (IP < 0.00001, I2 = 96%, Fig. 9 studies with 800 participants reported the relationship between DEX and the postoperative MMSE score on the first day after surgery. As we all know, MMSE is commonly used to assess cognitive function, a higher MMSE score indicates better cognitive function for postoperative patients. There was an obvious heterogeneity between the studies (P < 0.00001, I2 = 98%, Fig. 8studies involving 976 patients reported the MMSE score on the third postoperative day. A random effect model was adopted for meta-analysis considering the apparent heterogeneity (P < 0.00001, I2 = 98%, Fig. 4 articles including 628 patients reported the association between DEX and the MMSE score on the seventh day after surgery. Due to a large heterogeneity (A funnel plot of the 9 studies reporting the incidence of POCD on the third postoperative day was symmetrical, which indicated no publication bias Fig. , with alAt the present study, our meta-analysis clearly showed that the intraoperative use of DEX could obviously alleviate cognitive dysfunction, lower the incidence of POCD, and improve MMSE score compared to saline treated controls on the first, third, and seventh day following surgery. We found that DEX did reduce the incidence of POCD over saline placebo. Our results similar to previous reports in which sedation with intraoperative DEX was associated with a lower incidence of POCD compared with placebo.As the development of society and the trend of aging population aggravates, the increasing number of elderly patients are undergoing surgery, the elderly are more vulnerable to POCD. The incidence of POCD in the elderly is high, appealing to increasing interest in identifying strategies to prevent POCD. However, given its consequences, the concrete cause of POCD is still uncertain , 25. It Although POCD is common in older patients, the concrete pathophysiologic mechanism is still poorly understood. Our meta-analysis clearly reported that DEX can significantly reduce the incidence of POCD in patients and improve postoperative MMSE score. DEX, an imidazole derivative and a dextrorotatory isomer of medetomidine, is a novel adrenal α2 receptor agonist with high selectivity. DEX acts at the locus coeruleus in the brain stem, which contains the highest concentration ofα2adrenoceptors. It can activate the G protein on the α2 adrenergic receptor in the brain and spinal cord, and inhibit the excretion of norepinephrine and neuronal discharge, which affects the sympathetic nervous system and exerts its sedative, anti-anxiety and analgesic effects , 29. TheOverall, the previous meta-analyses published by Yang et al. and Lei et al. have also found that intraoperative DEX use was associated with reduction in the incidence of POCD and increased the MMSE score compared to saline group , 40. HowSome limitations were associated with our study. First, the number of studies and the corresponding sample size were relatively limited, and the doses and methods of administration of DEX given to patients varied substantially. Secondly, the enrolled studies’ inclusion and exclusion criteria, body weight, anesthetic doses, duration of surgery, surgical blood loss and consequently the characteristics of the patient cohorts, were also varied, which might have led to heterogeneity. Clinical heterogeneity of the participants can also influence the results of a meta-analysis. Finally, we have to admit that the result of POCD was lack of strength due to the only MMSE test. Different studies had different definition of POCD, which might lead to high heterogeneity in current study.In conclusion, DEX can ameliorate the POCD of patients who received surgical operations under general anesthesia, and effectively reduce the incidence of POCD and improve MMSE score. Thus, it can serve as a kind of adjuvant drug for general anesthesia in clinical practice.


Plasma extraction from blood is essential for diagnosis of many diseases. The critical process of plasma extraction requires removal of blood cells from whole blood. Fluid viscoelasticity promotes cell migration towards the central axis of flow due to differences in normal stress and physical properties of cells. We investigated the effects of altering fluid viscoelasticity on blood plasma extraction in a serpentine microchannel. Poly (ethylene oxide) (PEO) was dissolved into blood to increase its viscoelasticity. The influences of PEO concentration, blood dilution, and flow rate on the performance of cell focusing were examined. We found that focusing performance can be significantly enhanced by adding PEO into blood. The optimal PEO concentration ranged from 100 to 200 ppm with respect to effective blood cell focusing. An optimal flow rate from 1 to 15 µL/min was determined, at least for our experimental setup. Given less than 1% haemolysis was detected at the outlets in all experimental combinations, the proposed microfluidic methodology appears suitable for applications sensitive to haemocompatibility. Blood is a two-phase suspension of red blood cells (RBCs), white blood cells (WBCs), and platelets suspended in plasma. Given the high protein content, including serum albumin, globulins, and fibrinogen, within plasma, this fluid is often separated from whole blood and used for analytical purposes, including in vitro diagnostics. Blood plasma also contains glucose, electrolytes, hormones, carbon dioxide, and oxygen, so it is crucial for the intravascular osmotic effect which maintains the balance of electrolyte concentration and protects the body from infection and other blood diseases. However, the presence of cellular components inhibits diagnostic sensitivity, accuracy, and reproducibility. Hence, it is of great importance to remove blood cells from the whole blood and extract plasma for medical applications. Two conventional methods, namely centrifugal separation and membrane filtration, have become standard practice over the last century. Although the centrifugal technique is widely used in biological labs, such a device is bulky and involves a batch-based separation technique, thus it is limited for integration with other units to form a fully automated system for point of care diagnosis. Moreover, inappropriate centrifugation, despite the low likelihood, may cause haemolysis of RBCs, leading to contamination risks and misdiagnosis due to handling errors . On the Microfluidic techniques refer to the systems that process fluids in channels with typical dimensions (width and height) in the order of 10 to 100 µm. The corresponding characteristic length scale enables precise manipulation of fluid and particles, such as cellular components of blood, and thus facilitates the purity and efficiency requirements for plasma separation. Microfluidic technology has consequently become a promising candidate for plasma separation in recent years ,4,5. Based on the origin of manipulating forces, microfluidics can be classified as either active or passive techniques. The former utilises external active forces, such as dielectrophoresis , magnetoE. coli bacteria from RBCs  = (167.2A415 − 83.6A450 − 83.6A380). Haemolysis was determined for central (A) and side (B) outlets relative to each respective total haemoglobin concentration.To ensure the microfluidic device did not damage RBCs in the process of cell focusing and phase separation, samples collected from all outlets were assessed for haemolysis using the Harboe spectrophotometric method (PMID: 13646603) . BrieflyBlood samples diluted in PEO PBS were infused into the microfluidic device at specific flow rates by a syringe pump . To avoid the presence of air bubbles inside the microchannel, which could adversely interrupt the flow characteristics, the microchannel as well as the connected tubing were fully filled with PBS before each experiment. In experiments, to reduce sedimentation of blood cells , the syringe pump was regularly arranged from vertical to horizontal orientation, and vice versa. However, a mixing device should be integrated inline in practical applications, such as placing a magnetic stirrer bar in the sample chamber and activating it by a rotating magnetic field. The outputs from Outlet A and Outlet B were collected into two 2 mL tubes for each case. An inverted microscope (Nikon Eclipse Ti) equipped with a high-speed camera  at an ultra-short (~20 µs) exposure time was utilised to monitor and record the flow of samples inside the microchannels. The open-source software ImageJ  was adopted to analyse the captured videos. The images of each video were stacked so that the statistical distribution of blood cells could be visualised. The polynomial curve fitting method was adopted to fit the data of focusing performance through MATLAB.When blood samples diluted in PEO PBS solution flow through the serpentine channel, three forces exert on the cellular components, including an inertial lift force induced by the flow inertia , an elasCPEO = 0 ppm. We first compare it with whole blood diluted in 500 ppm PEO PBS solution. CPEO = 0 ppm, even with 10 times dilution, the cell-free layers are very narrow. However, the cell-free area is increased by the viscoelastic effect at certain range of flow rates for blood samples of CPEO = 500 ppm. Furthermore, the total width of free cell layers first increases and then decreases with the increase of flow rate , and then the total width of cell-free layers on two sides Wf can be estimated, so as δ.To quantify the focusing performance, a cell-free area ratio δ can be obtained (δ of 10 times diluted blood in 500 ppm PEO PBS is significantly higher than that in only PBS at low to moderate flow rates (1~30 µL/min), while a narrowed cell-free area ratio δ was found at high flow rates (40 and 50 µL/min). Importantly, a peak value of δ was identified for the focusing performance of 10 times diluted blood in 500 ppm PEO PBS, as demonstrated by the fitting curve. Specifically, when the flow rate was 10 µL/min, δ of 10 times diluted blood in 500 ppm PEO PBS was at its highest value of approximately 0.56, which is more than two times of the peak δ value (around 0.23) of PBS.By repeating the procedures above, the effect of flow rate on the cell-free area ratio obtained . As presδ generally shows an initial increase and subsequent decrease when flow rate increased, except for the case with the highest PEO concentration (CPEO = 2000 ppm) which demonstrates a consistently decreasing trend. Additionally, low PEO concentrations  demonstrated superior performance compared to counterparts with high PEO concentration . Specifically, the highest δ was approximately 0.44 of CPEO = 100 ppm at 15 µL/min, 0.58 of CPEO = 200 ppm at 15 µL/min, and 0.56 of CPEO = 500 ppm at 10 µL/min. In general, optimal δ were found within low to moderate flow rates, and this optimised flow rate decreased from 15 to 1 µL/min when CPEO increased from 100 to 2000 ppm  in the RBCs region increased for samples collected from the side outlet for the blood sample with PEO concentration ≥500 ppm.After determining the optimal viscoelastic focusing at 15 µL/min, the blood plasma separation was conducted for diluted blood samples (1/10) with different PEO concentrations and the collected plasma from the different outlets was examined. To determine an index of filtration efficiency for plasma extraction, absolute cell counts in the side outlet were presented relative to total cell counts in the central channel for each PEO concentration A. The fiWhile this study mainly focused on RBCs movement and blood plasma extraction for haemoglobin detection, movement of other blood cells (WBCs and platelets) must also be considered. Since particle size dictates cell centreline focusing or margination, WBCs  should focus towards the centreline, and platelets  should have less alignment. Therefore, there are likely fewer WBCs and more platelets in the separated plasma, evidenced by the flow cytometric data in The processing throughput of the proposed device is far less than standard centrifugation. However, many analytical methods only require small volumes of isolated plasma . While increasing the throughput of the device would not be required for future microfluidic devices with embedded inline analytics, one potential solution for increasing throughput would be to implement multiple parallel channels to amplify the output. The proposed plasma extraction microfluidic technology in the present study is part of a staged theme of works seeking to develop a fully-automated bedside point-of-care free-haemoglobin quantification system. Given the developments towards inline analytics, we believe that the relatively low throughput may not be a significant issue. The advantages of microfluidic plasma extraction technology and potential integration with other functional units and automation will likely increase clinical efficiency and reduce human intervention.Finally, it is plausible that the current microfluidic device could be deployed to ascertain blood phase separation for cell focusing and plasma isolation, with prospects to integrate inline plasma analytics at side outlets for development into a future stand-alone device for blood analytics.The current study investigated viscoelastic blood plasma extraction in a sheathless serpentine microchannel. The influences of PEO solution concentration, blood dilution, and flow rate on the cell focusing performance were tested. The experimental data demonstrated that the cell focusing performance, in general, shows a biphasic trend (first increasing and then decreasing) when flow rate increases, at least for the measured PEO concentrations and blood dilutions used. Hence, the optimal cell focusing performance for each case was determined with the flow rate ranging from 1 to 15 µL/min. Furthermore, the highest cell focusing performance among all the cases was found to be approximately 0.58 with 200 ppm PEO concentration and 10 times diluted blood at 15 µL/min flow rate. Finally, the haemocompatibility results showed that only <1% haemolysis was detected in the outlet region of any condition, indicating that the microfluidic device does not rupture RBCs in the process of viscoelastic focusing and plasma extraction.


MD = 4.16, p < 0.001. There was significant improvement at 12 weeks, compared to baseline, MD = 0.59, p = 0.009 for PA knowledge, and MD = 0.68, p = 0.003 for heart disease knowledge. There were reductions from baseline to 24 weeks in diastolic blood pressure , waist circumference  and BMI . Significant heart disease knowledge improvements were found at 24 weeks compared to baseline . Supervised exercise positively influenced Black African females′ health behaviours by understanding cultural perceptions of modifiable risk factors for NCDs.We employed the Health Belief Model (HBM) as a theoretical lens to explore the influence of an exercise intervention on the perceptions and knowledge of modifiable risk factors for non-communicable diseases (NCDs) among women from a low-resource setting in South Africa. We used a mixed-methods design, gathering qualitative and quantitative data at baseline (n = 95) and again after 12 weeks (n = 55) and 24 weeks (n = 44) of an exercise intervention. Qualitative data consisted of focus group discussions exploring the knowledge and perceptions of modifiable risk factors for NCDs at the three time points. We collected quantitative measurements of modifiable risk factors for NCDs  as well as objective physical activity (PA) data over seven consecutive days. Surveys on coronary heart disease and PA knowledge were conducted at all three time points. Qualitative findings indicated that health exposures and cultural traditions influenced the participant’s perceptions about PA and NCDs. Waist circumference significantly decreased at 12 weeks compared to baseline  Many people do not engage in sufficient physical activity (PA) ,2 and arWomen in sub-Saharan Africa engage in less leisure-time PA than their male counterparts. South African women also report lower PA levels than their male counterparts ,13 and aStudies to describe knowledge and perceptions of NCDs and PA in SA have previously been conducted ,21,22. AThe Health Belief Model (HBM) is one of the widely used frameworks to describe health behaviours. The HBM includes six constructs, namely, (1) perceived susceptibility, (2) perceived severity, (3) perceived benefits, (4) perceived barriers, (5) cues to action and (6) self-efficacy. The HBM was utilised as a theoretical lens  through We conducted this study to explore, describe and understand whether engaging women from a low-resourced community in supervised exercise could change their perceptions and knowledge of NCD risk factors and PA. By improving our understanding of the link between knowledge and perceptions and the role of supervised exercise, it will be possible to develop more sustainable PA interventions to manage NCD risk factors.This study is shaped by a pragmatic philosophy, which explores and describes the phenomenon. A mixed-methods study was conducted using the strength of both the qualitative and quantitative methods to yield a complete and sound analysis . A conveThe participants were volunteers recruited in 2015 via a convenience sampling technique from the Steve Tshwete Clinic catchment area in Ikageng, a dormitory town in the Dr Kenneth Kaunda District, near Potchefstroom North West Province, South Africa. The residents of the area are dependent on the public health system, report high levels of unemployment and live in informal housing with limited access to water and electricity. Based on the limited access to services, the setting is defined as low resourced. Recruitment resulted in 110 participants being assessed for eligibility for participation, of which 95 participants were included in this study. The research process flow is indicated in Participants were requested to attend supervised exercise sessions once a week for approximately sixty minutes at the Ikageng community hall. The exercise intervention was guided by the American College of Sports Medicine  and infoQualitative and quantitative data were collected simultaneously, with qualitative data given dominance at three time points. Baseline measurements were taken and repeated after 12 weeks of supervised exercise and again after 24 weeks of supervised exercise, as shown in The same researchers conducted four focus group discussions (FGDs) at each measurement point. FGDs were guided using an open-ended questionnaire and based on previous research until data saturation was reached and lasted approximately an hour. Discussions included four to 10 participants per group and were conducted in English with a Tswana translator. All FGDs were audio-recorded for analyses. Baseline FGDs had a total of 34 participants. Focus group discussions consisted of 25 participants at 12 weeks and 22 participants at 24 weeks of the intervention. Qualitative data aimed to explore and describe perceptions and knowledge of NCD risk factors and PA.®, CamNtech Ltd., Cambridge, UK) ) compared to baseline (43.97 min/day [0–437.0]) and 24 weeks (44.84 min/day [0–175.0]) see .2(2) = 14.56, p < 0.001, W = 0.18). Pairwise comparison with Dunn–Bonferroni correction shows significant differences between baseline and 12 weeks p = 0.003 and baseline and 24 weeks p < 0.001. There was no significant difference between 12 and 24 weeks, p = 1.00. The findings indicate an improvement in heart disease knowledge at 12 weeks and 24 weeks after baseline testing, as shown in Friedman’s test shows that heart disease knowledge significantly affected the time points ꭓ2(2) = 9.62, p = 0.010, W = 0.12). Post hoc analyses with Dunn–Bonferroni correction show a significant difference between baseline knowledge of PA compared to 12 weeks (p = 0.009). There was no significant difference between baseline and 24 weeks or between 12 and 24 weeks after baseline testing.Friedman’s test shows that PA knowledge scores differed significantly across the time points (ꭓAt baseline, participants lacked knowledge about NCD risk factors, scoring an average of 39% for the coronary heart disease knowledge survey, as shown in The PA knowledge survey results indicated that most of the participants knew the effect of PA on health, scoring high on PA knowledge, as shown in Qualitative and quantitative results are integrated and presented in This mixed-methods study used the HBM as the lens to explore and describe perceptions and knowledge of NCD risk factors and PA among women from a low-resourced community. Black South Africans tend to lack knowledge and have negative perceptions about NCDs and PA ,22,25; TIn this study, participants initially perceived an increased risk of NCDs was caused by non-modifiable risk factors, including family history and age. According to Sallis , family Though participants were aware, through media, of various types of PA, including sporting activities, they did not see themselves engaging in such activities because of fear of injury. Participants were generally knowledgeable that PA is good for health. However, they lacked knowledge about appropriate PA activities they could engage in. This finding is consistent with an observational study in a similar setting where participants reported enjoying walking and house chores as their preferred form of PA . ParticiConsistent with the findings by Sehole and Van der Heever , particiDuring the 24 weeks of supervised exercise interventions, a positive effect was found on the NCD risk factors and PA knowledge. Although physical improvements were observed during the 12 weeks of the supervised exercise intervention, perceptions concerning risk factors of NCD and PA changed only after 24 weeks of the exercise intervention. A supervised exercise intervention seems to positively influence women’s health behaviours from a low-resourced community through a deeper understanding of the influence of cultural perceptions on modifiable risk factors for NCDs. Interventions should preferably be longer than 12 weeks to change perceptions and planned outside working hours in a safe environment to reduce program attrition.


Progressive familial intrahepatic cholestasis (PFIC) is a spectrum of rare genetic diseases characterized by inadequate bile secretion that requires substantial ongoing care, though little research is published in this area. We report health-related quality of life (HRQoL) and work productivity outcomes from the retrospective, cross-sectional PICTURE study investigating the burden of PFIC on caregivers. Information from caregivers of patients with PFIC 1 or 2 in Germany, the United Kingdom and the United States from September 2020 to March 2021 was included.The PICTURE study sample comprised HRQoL responses from 22 PFIC caregivers. Patients were on average 8.2 years old; most caregivers were 30–49 years old (68%) and mothers (77%). Median CarerQoL-7D score was 67.7/100; mean CarerQoL-VAS score for general happiness was 5.7/10 (SD 2.1). Most caregivers reported fulfilment in their caregiving responsibilities, but problems with mental and physical health, finances, and relationships. When stratified by patient’s PFIC type, mean CarerQoL-7D and CarerQoL-VAS scores suggested worse HRQoL outcomes with PFIC2 versus PFIC1 . Additionally, more caregivers reported impact on sleep in the PFIC2 versus PFIC1 subgroup (93% vs. 75%). When stratified by history of PFIC-related surgeries, mean CarerQoL-7D and VAS scores were higher among those whose children had no specified surgeries . Nearly all caregivers reported an impact of caregiving responsibilities on sleeping (86%) and on personal relationships (82%). No caregivers reported having formal care support. Most caregivers were employed (73%); a third reported mean productivity loss of 12.9 days (SD 19.3) over the last 3 months, and a mean of 2.8 (SD 9.5) missed years of employment during their career. A higher number of workdays were missed by PFIC 2 caregivers compared to PFIC1 over last 3 months (16 days vs. 3 days).The PICTURE study has demonstrated the prevalent, comprehensive, and meaningful burden that caring for an individual with PFIC has on caregivers. Despite fulfilment from caregiving, the breadth and depth of these responsibilities reduced caregiver reported HRQoL including mental and physical health, productivity, career prospects, sleep, relationships and finances.The online version contains supplementary material available at 10.1186/s13023-022-02177-0. Progressive familial intrahepatic cholestasis (PFIC) is a spectrum of rare genetic diseases of cholestasis, characterized by inadequate bile secretion, often requiring liver transplantation and leading to liver failure and early death –7. The rThe most prevalent PFIC types within the patient population are PFIC type 1 and type 2. Type 1 (PFIC1) presents in infancy or early childhood and is caused by mutations in the ATP8B1 (or FIC1) gene. PFIC1 typically manifests with a moderate to severe intensity: children may have severe itching (pruritus), an enlarged liver  and may also have extrahepatic symptoms, including watery diarrhea, pancreatitis, deafness and short stature or failure to thrive . ChildreType 2 (PFIC2) is caused by a malfunctioning copy of the gene ABCB11 (encoding the BSEP export pump). Also typically diagnosed in early childhood, PFIC2 shows rapid disease progression, with severe itching, hepatomegaly, and a risk for hepatocellular carcinoma and cholesterol stones; extrahepatic manifestations are more rarely encountered in PFIC2 patients .The rare nature of PFIC has presented challenges to understanding its impact on the daily lives of patients and their caregivers. Recent systematic reviews have identified few studies investigating the health-related quality of life (HRQoL) of patients with PFIC , 12. HRQIn the context of extremely limited information on the humanistic burden of PFIC on patients directly, there is no identifiable published work on the burden of PFIC on their caregivers. Considering the early onset, severe nature, and certain morbidity associated with PFIC through the lifespan, the care requirements of these patients are necessarily substantial and persistent. Analogous work has shown significant caregiver burden including reduced HRQoL and work productivity among parents of children and adolescents with cystic fibrosis  –17. To tA retrospective analysis of data from the PICTURE study was conducted, the design and methods of which have been reported previously . The PICThe PICTURE study design has two main sources of data: physicians and caregivers. Physicians provided clinical and resource use data of PFIC patients at the time of consultation, via an electronic Case Report Form (eCRF). Caregivers of PFIC patients, recruited by the physician as they attended a clinical appointment with the patient, completed online specific self-completion questionnaires about the impact of the disease on their lives.The study was conducted under the guidance and expertise of an Expert Reference Group (ERG), consisting of a representative of academia as principal investigator (University of Chester), partnering charity and advocacy representatives (PFIC Network and Children’s Liver Disease Foundation), as well as experts in the field of liver diseases: Prof. Karen Murray, Prof. Jorn Schattenberg, Prof. Anil Dhawan and Jose Willemse.The study protocol and materials were approved by the Research Ethics Sub-Committee of the Faculty of Health and Social Care at the University of Chester (no. RESC0820-1042). The study was conducted in accordance with all relevant ethical guidelines; all caregivers provided informed consent.Participant outreach and recruitment was conducted via electronic media, using both a standard fieldwork approach and a hybrid approach. The standard fieldwork approach was conducted from September 2020 through to March 2021 and aimed to recruit representative samples of physicians treating PFIC in each country . Eligible physicians had to have ≥ 2 years of practice experience and have treated ≥ 1 patient with PFIC in the last 12 months. Participating physicians recruited consecutive eligible patients/caregivers (maximum of 10) during routine clinical visits for any reason, including adult caregivers/guardians of patients (any age) with genetic diagnosis of PFIC  ≥ 12 months prior to study entry. Patients/caregivers were excluded for current or recent (≤ 12 months) clinical trial participation. Physicians completed electronic case report forms (eCRFs) with clinical and economic information based on the patient’s medical history. Caregivers completed online questionnaires regarding the impact of PFIC on their lives and productivity. Physicians performed all activities outside of professional practice parameters and received remuneration based on fair market value; caregivers also received remuneration for completion of questionnaires.The hybrid recruitment approach was conducted through February and March 2021 to bolster caregiver participation, as their involvement was likely affected by the COVID-19 pandemic over 2020 fieldwork months. Partnerships with the PFIC Network (pfic.org) and the Children’s Liver Disease Foundation  facilitated direct outreach to families of patients with PFIC. Participants recruited through the hybrid approach completed the same online questionnaire on the impact of PFIC as in the fieldwork approach, with 5 additional questions related to clinical outcomes and resource use, to facilitate analyses of clinical and economic burden in the absence of eCRFs completed by the physicians.Given that PFIC is a rare disease, measures were taken to solidify the investigators’ confidence that the study sample was based on patients identified with PFIC. An interim quality-control analysis of incoming data revealed that a high percentage of cases reported were adults, with no surgical history . This suggested that non-progressive cholestasis cases were included for participation in the study. Therefore, a clinical algorithm was developed and applied post-hoc, to select children and young adults most likely to exhibit PFIC. The requirements were patients < 5 years old at the time point of PFIC diagnosis and < 18 years old at the time of study entry. Exceptions were made for those aged 18–35 at study entry and ≤ 5 years old at diagnosis if they had a record of a PFIC-related surgery such as PBD or liver transplantation  or caregivers only (hybrid approach), including age, sex, country, PFIC subtype, caregiver type , highest caregiver education level, marital status, and urbanicity . Caregivers provided information about the impact of PFIC on HRQoL, work productivity, their professional career, sleep, and relationships. A supplemental question regarding the impact of the COVID-19 pandemic on PFIC caregiving was also included.www.imta.nl/carerqol/) [Caregiver-reported HRQoL was captured using the validated 7-dimension CarerQoL-7D and the caregiver HRQoL visual analog scale  , 20. Thererqol/)  (Additiowww.eprovide.mapi-trust.org). The WPAI captures the percentage of impairment in the past 7 days, in terms of work-related absenteeism (not being present at work), presenteeism , work productivity loss (based on absenteeism and presenteeism), and impairment of regular activities due to health-related caregiver duties, where higher scores indicate greater impairment, or worse outcomes [Work productivity was assessed using the Work Productivity and Activity Impairment questionnaire Adapted for Companion or Caregiving v2.0  and relationships . Results were examined overall, by PFIC type (1 or 2), by history of surgery (with PBD/Liver transplant vs. without PBD/Liver transplant), and by history of specific procedures .Caregivers also indicated their ability or lack thereof to access supplementary professional care. The impact of the COVID-19 pandemic was also assessed, in relation to the impact on resource use, caregiver mental health, and caregiver physical health.www.stata.com). No imputation of missing values was performed.All data were summarized using descriptive statistics including frequencies, proportions, and measures of central tendency using STATA 16  were excluded by the clinical algorithm exercise to identify the most probable PFIC cases (n = 106). HRQoL responses were available from a total of 22 caregivers . Caregivers were evenly distributed across Germany, the UK and US. Most caregivers were 30–49 years old (68%) and mothers  out of 100, with a median score of 67.7. The mean CarerQoL-VAS score of general happiness was 5.7/10 . For the positive dimensions of the CarerQoL-7D, most caregivers appeared to indicate fulfilment in their caregiving responsibilities and having support available to help when needed . Both CarerQoL-7D and CarerQoL-VAS scores were lower for patients with PFIC 2 when compared with PFIC 1, suggesting worse HRQoL outcomes for this subgroup , and without formal statistical comparisons, CarerQoL-7D and CarerQoL-VAS scores both appeared higher among those whose children had none of these surgeries .Additionally, more caregivers of children with PBD/liver transplant surgeries appeared to report difficulty with partner relationships; other HRQoL measures were generally comparable between subgroups . Based on responses collected between September 2020 and March 2021, approximately half of caregivers reported an impact of the COVID-19 pandemic on resource use , and on their own mental health  and physical health .The majority of caregivers indicated current employment . During the 7 days prior to data collection, mean impact on productivity due to absenteeism was reported as 27%  reported suffering an impact on their employment, with a mean loss of 12.9 days (SD 19.3) within the last 3 months. Additionally, 36% of caregivers reported missing years of employment as a result of their caring duties, with a mean of 2.8 years (SD 9.5) during their career , as well as daily activity impairment (mean impact of 54% in PFIC 2 vs. 40% in PFIC 1). On the other hand, scores for presenteeism and overall work productivity impairment were comparable across subgroups.When compared to PFIC 1, a larger proportion of PFIC 2 caregivers reported missing workdays, and a substantially higher number of workdays were missed by PFIC 2 caregivers in the 3 months prior to data collection (16 days vs. 3 days in PFIC 1 group). A higher proportion in this subgroup also reported missing work years due to caregiving duties . The mean number of years missed was similar across subgroups .History of surgical procedures was also found to yield differences across strata in the assessment of productivity and activity impairment. Caregivers of patients with PBD/liver transplant surgeries reported greater impact on productivity due to absenteeism when compared to caregivers with no history of these surgeries (mean impact of 48.3% vs. 5.5%). Additionally, more caregivers in the PBD/liver transplant subgroup reported missing workdays 60% vs. 17%), and they appeared to miss substantially more workdays in the 3 months prior to data collection . Caregivers in this subgroup often reported missing years of work due to caregiving duties , nearly all also reported caregiving-related mental (73%) and physical (64%) health problems, and relational problems with their child (95%). Despite having general support for caregiving tasks when needed (82%), caregivers reported disruption of close personal relationships (82%), particularly with their partner (50%). All caregivers reported impediments to accomplishing regular daily activities; on average, a level of impairment of 49% was reported, with nearly all caregivers (86%) suffering from some level of sleep difficulty attributable to the burden of caregiving. The majority of respondents were mothers aged 30–49 years—prime working age adults—nearly all of whom had a minimum of high school-level education (82%). While most caregivers were employed in some capacity (73%), only one-third were employed full-time  with half of the working caregivers reporting having missed an average of 13 workdays in the past 3 months, and losing, on average, 2.8 work years of their career. Absenteeism and presenteeism were high, with the majority of employed caregivers reporting limitations to career choice and progression (both 73%).When considering outcomes from the perspective of presence/absence of surgeries in patient sample, it appears HRQoL scores of parents with surgically treated children are worse . An observation by the clinical expert group overseeing the PICTURE study suggests this may be correlated with children in the ‘surgeries’ group having a more severe or more progressive form of PFIC 1 or 2, warranting a higher and potentially earlier need for life-saving surgeries. Furthermore, by looking at the PFIC type stratum, caregivers of PFIC 1 children have better HRQoL than PFIC 2 (CarerQoL mean scores of 71.2 in PFIC 1 group vs. 59 in PFIC 2)—which could possibly be correlated with higher rate and need of liver transplant in PFIC 2 cohorts, as stated by the currently available literature. Taken together, these findings provide a first overview of the burden of caregiving for a child with PFIC, with no other published reports of this burden to the authors’ best knowledge.Given there is no available literature to provide direct context for these findings, the authors suggest cystic fibrosis (CF) may serve as a fair analogous condition for PFIC, as it also presents in early life and children/adolescents with CF also risk serious morbidity with demanding care and treatment regimens that substantially impact their HRQoL .Fitzgerald and colleagues  reportedPseudomonas aeruginosa  [Significant predictors of high caregiver burden in the CF study included older age of the child with CF, being the mother of the child with CF, and having a child with a positive test (ever) for rtality) , 24. ThoThis study also raises research questions which the authors recommend for investigation in future studies; among these, further insights into the potential correlation of the patients’ HRQoL with the caregivers’ HRQoL would likely aid the patient and medical community in identifying any remaining unmet needs.Our findings should be interpreted in the context of the rare nature of PFIC and the consequent small sample size, which constitutes an unpreventable study limitation. While we were able to sample treating physicians and received recruiting support through PFIC-specific organizations, a clinical algorithm applied post-hoc was necessary to increase the investigators’ confidence that included cases were PFIC, rather than another manifestation of cholestasis. Nonetheless, the voluntary nature of participation and consequent potential for selection bias, both among the healthcare providers and caregivers, should be noted. Caregivers completing the questionnaires may have been subject to recall bias, particularly for measures intended to characterize outcomes over longer time periods in the past, though there were few such questions.The PICTURE study data indicated that caring for an individual with PFIC could cause comprehensive and meaningful burden on caregivers. Despite fulfilment from caregiving, the breadth and depth of these responsibilities reduced caregiver reported HRQoL including mental and physical health, productivity, career prospects, sleep, relationships and finances. The nature of PFIC and currently scarce available management approaches are potentially the reasons for the heavy burden on caregivers participating in this study, which may be alleviated by novel transformative therapeutic options. Given the level of unmet need that this study has uncovered, along with the scarcity of developed and published literature in this rare disease field, it is evident that more research is required to support PFIC patients and their caregivers.Additional file 1. An overview of the algorithm applied to the PICTURE patient dataset, the tariffs used to derive caregiver quality of life, and the characteristics and outcomes of the sample analyzed in this paper.