Supplemental Digital Content is available in the text Prediabetes is prevalent and significantly increases lifetime risk of progression to type 2 diabetes mellitus (T2DM). Acupuncture has been increasingly used for prediabetes in China but its effect is unclear. We aim to assess the efficacy and safety of acupuncture in preventing or delaying incident diabetes among individuals with prediabetes.The following 8 databases will be searched from inception to September 1, 2018: the Cochrane Central Register of Controlled Trials (CENTRAL) on the Cochrane Library, PubMed, Embase, Chinese National Knowledge Infrastructure database, Chinese Biomedical Literature database, Chinese Scientific Journal database, Wan Fang database, and Clinical Trials. The incidence of diabetes and regression toward normoglycemia will be accepted as the primary outcomes. The Cochrane Risk of Bias tool will be used to evaluate the methodologic quality of eligible studies. Meta-analysis will be performed by Review Manager 5.3.This study will provide a high-quality synthesis of current evidence of acupuncture in the prevention of T2DM from several aspects including the incidence of diabetes, regression toward normoglycemia, fasting plasma glucose, 2-hour plasma glucose level after a 75-g oral glucose tolerance test, glycosylated hemoglobin level, body mass index, and adverse drug events.The conclusion of this review will provide evidence to judge whether acupuncture is an effective and safe intervention for prediabetes.PROSPERO CRD42018111236. The toll of diabetes and diabetic complications on patients’ health and quality of life is enormous. The burgeoning diabetes pandemic also comes with high personal and financial costs to the individual, society, and the economy. Prediabetes is an intermediate metabolic state between normoglycemia and diabetes, including either IGT, impaired fasting glucose (IFG), or both conditions. IGT and IFG are “high-risk” states for diabetes development.,5 The risk of progression to diabetes is greater for patients with IGT and IFG close to the diagnostic boundary for diabetes. On average, about 15% to 30% of adults with prediabetes are expected to progress to diabetes in 5 years. From a population perspective, this risk is substantial given the high prevalence of prediabetes.Diabetes mellitus is currently a major public health problem. The International Diabetes Federation estimates that 420 million adults worldwide now have diabetes and 318 million have impaired glucose tolerance (IGT), with a projected global prevalence of 642 and 482 million, respectively, by 2040.–9 However, it is not easy for patients with prediabetes to maintain rigorous and sustained LM for the long term. Numerous landmark trials in the past quarter-century have demonstrated that various pharmacologic interventions, including metformin, acarbose, pioglitazone, rosiglitazone, and orlistat, can effectively delay or prevent the development of T2DM in high-risk individuals with IGT.–19 Despite the clarity of these findings, translational prevention programs have faced many real-world impediments, and none of the tested interventions have an approved label for use in the prevention of T2DM and have been widely adopted as components of routine clinical care.Accordingly, many studies have examined the efficacy of lifestyle modification (LM) and pharmacologic interventions in preventing or delaying incident diabetes among patients with prediabetes. LM has been shown to reduce the incidence of type 2 diabetes mellitus (T2DM), and is recommended for prediabetes by American Diabetes Association guidelines.Acupuncture, one of the important treatments used in traditional Chinese medicine (TCM), involves the insertion of fine needles into the skin at specific acupoints. It has a long history of use in China and is increasing accepted by people in the developing and developed world as an alternative therapy to conventional treatments. The selection of acupoints is based on the syndrome differentiation from the diagnosis according to TCM theory. Pattern identification is a unique TCM concept that summarizes and differentiates the nature, location, and pattern of diseases, which is the essential guide for TCM therapy. The precisely tailoring acupuncture acupoints for individuals based on each individual pattern type can maximize its efficacy. TCM emphasizes the idea that health is maintained by a balance of energy within the body. Acupuncture helps to correct imbalances to relieve symptoms and treat the disease by stimulating various acupoints based on individual syndrome differentiation.–22 Mechanistic studies applying acupuncture to the treatment of diabetes suggest that the major effects are associated with the control of blood glucose, body weight loss, protection of pancreas islet function, improvement of insulin resistance, and adjustment of the levels of related hormones, such as melatonin, insulin, glucocorticold, and epinephrine.,24 Moreover, acupuncture treatment combined with hypoglycemic drugs has a synergic effect in lowering blood glucose. Currently, acupuncture has been increasingly used in the prevention of T2DM. Some studies indicated that acupuncture could increase the regression toward normoglycemia, lose body weight, lower fasting plasma glucose (FPG), 2-hour plasma glucose (2-hPG), and glycosylated hemoglobin (HbA1c) in patients with prediabetes.,26 However, few systematic reviews have been conducted to evaluate the effect of acupuncture in the prevention of T2DM. Therefore, we aim to systematically assess the efficacy and safety of acupuncture in preventing or delaying incident diabetes among individuals with prediabetes.Acupuncture has been widely used for the treatment of diabetes and diabetic complications in China. Previous studies indicated that acupuncture therapy might be effective for diabetes and its complications .22.12.1.1All randomized controlled trials (RCTs) of acupuncture for prediabetes will be included. Excluded from the meta-analysis are duplicated publications, studies with unavailable or incorrect data, articles not reporting outcomes of interest. Also excluded are studies enrolling fewer than 10 participants and quasi-randomized controlled clinical trials . The articles will not be restricted based on publication type.2.1.2Participants with prediabetes, including IFG, IGT, or both conditions, will be included irrespective of gender, age, and ethnicity. All patients must be diagnosed with prediabetes by clearly defined or internationally recognized criteria.2.1.3Acupuncture type can include: body acupuncture, electroacupuncture, ear acupuncture, scalp acupuncture, fire needling, plum blossom needle, elongated needle, intradermal needling, or dry needling. The control intervention can include: no treatment, placebo/sham acupuncture, or other interventions . Trials that evaluate acupuncture plus another therapy compared with the same therapy alone will also be included. Trials that only compare different types of acupuncture or different points will be excluded.2.1.42.1.4.1The incidence of diabetes and regression toward normoglycemia will be assessed as the primary outcomes.2.1.4.2The secondary outcomes of this review will include FPG, 2-hPG level after a 75-g oral glucose tolerance test (OGTT), HbA1c level, body mass index (BMI), and adverse drug events.2.22.2.1https://www.Clinicaltrials.gov/). The search will be performed in English and Chinese. The following search terms will be used: acupuncture, acupuncture therapy, acupuncture treatment, acupoints, electroacupuncture, ear acupuncture, auricular acupuncture, scalp acupuncture, fire needling, plum blossom needle, elongated needle, intradermal needling, dry needling, prediabetes, impaired fasting glucose (IFG), impaired glucose tolerance (IGT), impaired glucose regulation (IGR), diabetes prevention, RCT, controlled clinical trial, randomized, placebo, drug therapy, randomly, trial, groups. The search terms will be translated into Chinese when reviewers search the Chinese databases. The search strategy for PubMed will be presented in supplementary file 1 (which describes the PubMed search strategy) and modified by using other databases.The Cochrane Central Register of Controlled Trials (CENTRAL) on the Cochrane Library, PubMed, Embase, Chinese National Knowledge Infrastructure (CNKI) database, Chinese Biomedical Literature database, Chinese Scientific Journal database (VIP), and Wan Fang database will be searched to identify eligible trials published from inception to September 1, 2018. Ongoing registered clinical trials will be searched in the Clinical Trials, gov . The full text of articles potentially suitable for the review will be obtained for further assessing eligibility based on the inclusion criteria or/and exclusion criteria. The studies that do not fulfill the inclusion criteria will be excluded and listed with reasons for their exclusion. Any disagreement will be resolved by consensus or discussion with a 3rd party (QN).2.3.2Two review authors (YY and XL) will independently extract information on patients, methods, interventions, outcomes, and results using a data extraction form designed for this review. Extracted data will be compared by 2 review authors for completeness and accuracy and double-checked by another review author if necessary. Authors will be contacted by e-mail to obtain further data and verify methodologic quality when necessary. Any disagreement will be settled by discussion or by consulting a 3rd author (QN). The data extraction form will include the following items:1.general information: title, authors, year of publication, and source;2.trial characteristics: design, duration of follow-up, method of randomization, allocation concealment, incomplete outcome data, blinding ;3.intervention(s): intervention(s) , comparison intervention(s) ;4.patients: total number and number in both groups, baseline characteristics, diagnostic criteria, withdrawals and losses to follow-up ;5.outcomes: outcomes specified above, length of follow-up, quality of reporting of outcomes.2.3.3 including random sequence generation (selection bias), allocation concealment (selection bias), blinding of participants and personnel (performance bias), blinding of outcome assessment (detection bias), incomplete outcome data (attrition bias), selective reporting (reporting bias), and other bias. For each domain, the following description will be used to assess proper management of the risk of bias: low risk, high risk, or unclear. Each study will be respectively categorized as “low risk of bias” , “high risk of bias” (1 or more items are in high risk), and “unclear risk of bias” (1 or more items are in unclear risk). Low risk of bias represents a good quality. Any disagreements will be resolved by discussion with QN.The methodologic quality of RCTs will be assessed using each item specified by the Cochrane Risk of Bias Tool,2.3.4Dichotomous outcomes will be presented as risk ratio and 95% confidence intervals (CIs), and continuous outcomes will be presented as mean difference/standard mean difference and 95% CIs.2.3.5The unit of analysis will be each patient recruited into the trials.2.3.6For each included study, the number of dropouts, exclusions from the analysis and missing data will be gathered by contacting the study author. If unable to obtain sufficient data, available case analysis will be performed. The potential impact of missing data on the findings of the review will be described in the discussion section if necessary.2.3.7I-square (I2) statistic. The I2 statistic of less than 50% indicates a low level of statistical heterogeneity, and that of 50% or more will be considered substantial statistical heterogeneity. If substantial heterogeneity is identified, we will report it and explore possible causes using sensitivity analysis and subgroup analysis.Statistical heterogeneity will be assessed with the 2.3.8 The results will be interpreted carefully based on several explanations for funnel plot asymmetry.A funnel plot will be constructed and examined to assess publication bias and possible small study biases if the group include more than 10 trials.2.3.9P < .05 is considered statistically significant. Whether a fixed effects model or a random effects model will be used depends on the results of I2 test for heterogeneity. If I2 test <50%, a fixed effects model will be used to pool the data. If I2 test ≥50%, a random effects model will be used for data analysis instead. Subgroup analysis and sensitivity analysis will be performed to explore the causes of heterogeneity. If meta-analysis is not applicable, we will conduct a systematic narrative synthesis providing information to summarize and explain the characteristics and findings of the included studies.If 2 or more eligible RCTs are identified, meta-analysis will be performed with Review Manager 5.3. All tests are 2-tailed, and 2.3.10We plan to carry out the following subgroup analyses if possible: comparison between acupuncture and no treatment, placebo or sham acupuncture, LM, western medicine treatment; comparison between acupuncture plus another therapy and the same therapy alone; different types of prediabetes, including IFG, IGT, or both conditions. We will use the formal test for subgroup interactions in Review Manager 5.3.2.3.11Sensitivity analysis will be performed to explore the effects of trial risk of bias on primary outcomes if possible. In the analysis, we will exclude lower quality trials and repeat the meta-analyses to examine whether the quality of included studies influences the pooled results.2.3.12The Grading of Recommendations Assessment, Development, and Evaluation (GRADE) methodology will be used to assess the quality of the evidence and risk of bias with GRADEprofiler (GRADEpro) V.3.6 software. The assessment will be adjudicated into 4 levels: high, moderate, low, or very low.3,2,4–6 Numerous landmark trials have demonstrated that LM and pharmacologic interventions could delay or prevent incident diabetes among individuals with prediabetes, but it is not easy for patients to maintain rigorous and sustained LM for the long term and none of the tested pharmacologic interventions have been widely adopted as components of routine clinical care.–19 There are an increasingly number of studies on acupuncture for prediabetes published with inconclusive results in recent years. Acupuncture may be a useful treatment for prediabetes. Therefore, it is essential to perform a systematic review and meta-analysis to assess the effect of acupuncture in preventing or delaying the development of T2DM. The flow diagram of this systematic review is shown in Figure Prediabetes is prevalent and significantly increases lifetime risk of progression to T2DM.However, this systematic review will have some limitations. Only studies published in English and Chinese will be searched and included because of language barriers, so a language bias may exist. High heterogeneity may also arise from the various evaluation standards from different acupuncture therapies. Nevertheless, this systematic view should help further expand our understanding of acupuncture treatments in prevention of T2DM.The PRISMA-P  checklist of this protocol is presented in PRISMA-P checklist.QN conceived of the study, supervised LS to perform this review. The manuscript was drafted by LS and LF. LS and LF developed the search strategy. YY and XL will independently screen the potential studies and extract data. MZ and YZ will assess the risk of bias and perform data synthesis. QN will arbitrate any disagreement and ensure that no errors occur during the review. LS and LF contributed equally to this work and are co-first authors. All review authors critically reviewed, revised and approved the subsequent and final version of the protocol.Conceptualization: Qing Ni.Data curation: Yanan Yang, Xiaowen Li, Meizhen Zhang, Yueying Zhang.Methodology: Liwei Shi.Project administration: Liwei Shi.Supervision: Qing Ni.Writing – original draft: Liwei Shi, Ling Feng.Writing – review & editing: Liwei Shi.


Psychological harassment is a serious occupational risk for nurses, but little is known about its related factors and possible predictors. The objective of the present study was to investigate whether nursing students’ neuroticism trait and coping styles can predict psychological harassment at work when they later become nurses. A non-experimental, longitudinal, three-wave prospective study with a time lag of 6 years was carried out, following nursing students from three Spanish universities until they joined the health labor market. The age range of the sample was 20–48 years, and the mean age was 26.99 ± 5.72; the majority of the sample were women (88.5%). Predictor variables were neuroticism and coping styles . The criterion variable was psychological harassment. To examine the model fit between the predictor and criterion variables, we conducted structural equation modelling. Results confirmed a high correlation between neuroticism and psychological harassment. In addition, a direct effect of neuroticism on psychological harassment was found; however, emotional and behavioral coping styles did not show a good fit. Proactive interventions to improve emotional self-control are needed in order to prevent negative effects of psychological harassment at work on nurses. The European Foundation for the Improvement of Living and Working Conditions has shown that workplace violence , which can include verbal and physical abuse, is strongly associated with work-related stress and poor mental health . LeymannResearch has identified factors associated with work-stress, bullying, and psychological harassment in the workplace . HoweverPsychological harassment can be considered a work-related process which is influenced by both environmental and personal factors that can act independently of each other . Among t1The neuroticism personality trait, measured at T1, T2, and T3, positively predicts harassment at T3.2Emotional- and behavioral-focused coping, measured at T1, T2, and T3, predict harassment at T3.Although the effect of personality and environmental factors on workplace violence has been explored, to our knowledge, no longitudinal studies have been published about the influence of the personality on psychological harassment, following a cohort of nurses from the beginning of their studies until their professional role. The present study addresses this gap. Based on the diathesis-stress model, we hypothesized that an emotional dispositional personality trait (high neuroticism) and certain styles of coping with stressful situations could increase vulnerability to harmful stress situations (harassment) in the workplace. More specifically, we hypothesized that:n T1 = 249, T2-2010, end of academic studies n T2 = 199, T3-2013, after three years of work n T3 = 70) was carried out, following Spanish nursing students until they became registered nurses. At T3, the sample consisted of 70 nurses (10 men and 60 women) from three Spanish universities, with a mean age of 26.99 ± 5.72 years. For a more detailed description of the initial sample, see references [A longitudinal, three-wave prospective study  . The iteThe study was conducted in accordance with the Declaration of Helsinki. The procedure was approved by the Research Committee of the University of the Balearic Islands. Participation was voluntary. It was made clear that the participants could withdraw at any time and that the information would remain confidential. A full explanation of the research was provided, and informed consent was obtained. At T1 (start of nursing studies) and T2 (the end of the studies), the questionnaires were filled out in a regular lecture class. At T3 (three years after graduation), T2 participants were contacted by e-mail and encouraged to participate in the last wave of the study.2 statistic [Descriptive analyses were conducted using IBM SPSS v.24 . After finding no attrition bias on the variables and between participants who remained and those who dropped out, we applied structural equation modelling (SEM) to test the hypotheses. SEM was calculated with EQS 6.2 software  , using ttatistic  which shtatistic , Normed tatistic , Non-Nortatistic , whose vDescriptive analysis showed that neuroticism scores remained stable over time, and the use of emotional coping was predominant in all the assessments. In the case of reported psychological harassment measured at T3, participants retrospectively reported its occurrence in the previous two years. Thus, 8.8% of the participants perceived themselves as being harassed at least once a week for more than 6 months, 27.9% had witnessed harassment, and 5.9% felt harassed at the time of the assessment. The harassment could come from co-workers, supervisors, or subordinates, indistinctly.p < 0.01) and coping styles (p < 0.05) in the different waves  = 2.871; CFI = 0.989; NFI = 0.967; NNFI = 0.968; SRMR = 0.035); that is, the neuroticism personality trait (exogenous variable), measured at T1, T2, and T3, positively predicts psychological harassment (endogenous variable) at T3  at T3 . Howevere) at T3 ; as can The results of this study confirm that the neuroticism personality trait predicts psychological harassment in a 6-year longitudinal study from the beginning of nurses’ training until three years after their professional role begins.Based on our findings, 8.8% of Spanish nurses perceived themselves as being harassed at least once a week for six months. A meta-analysis carried out on 86 independent samples found an average prevalence of workplace bullying of 14.6% . This hiNeuroticism was also associated with harassment. Our literature review detected only one similar study in Slovak nurses , but theAs we established in our hypotheses, the neuroticism trait was relevant in explaining nursing harassment six years later. This is a very interesting result with no supporting evidence from longitudinal nursing studies in the literature. The potential explanation for this predisposition to harassment in highly neurotic nurses could be their extreme hyperreactivity to stressors. Based on scientific evidence, highly emotional nurses are more sensitive to signs of punishment, perceive events as more dramatic and uncontrollable, and react in an exaggerated emotional way, making their own problematic situations worse . These eThis study has some limitations. The first is the reduction in the sample size from wave one to wave three. Professional nurses with high levels of harassment would have been more motivated to participate in the last phase of our study. However, the low participation in wave three could be due to feelings of embarrassment and shame produced by the possibility of being labelled a victim of harassment or a highly emotional reactive professional. Nevertheless, the three-wave assessment from the beginning of nursing training to the professional nursing role can be considered an important strength. Follow-up studies are needed with larger professional nursing samples, following them for several years at work, in order to replicate our study and consider the possible reverse causality hypothesis .Psychological harassment is a serious work-stress problem in nursing. Based on our results, this phenomenon can be predicted by emotional instability (neuroticism). These findings have interesting implications for professional practice and nurses’ wellbeing. Based on our findings, and in order to prevent negative effects of harassment and work stress in nursing, proactive interventions are needed, not only in the form of social support from colleagues and staff, but also through training in emotional self-control techniques, such as relaxation or mindfulness, in both nursing degrees and postgraduate studies.


To identify main complications in outpatient follow-up, as well as factorsbefore or during operation that may interfere in patient's evolution.Retrospective study of patients submitted to total cavopulmonary shunt withextracardiac conduit from 2000 to 2014 at the Hospital doCoração  and who underwent clinicalfollow-up at this institution.P=0.04). Between thepatients currently under follow-up, 20 (35%) of them had been evaluated byultrasonography and had some degree of hepatic congestion and/orhepatomegaly. 16.7% of the patients with such alteration had HLHS(P=0.057).One hundred and fifty surgeries were performed and 59 patients maintainedoutpatient follow-up. The mean age of these patients at the time of surgerywas 4.45 years (median of 45 months) and 70.2% of them were males. Among thepatients undergoing outpatient follow-up, postoperative time at evaluationranged from 10 days to 145 months; 30 (50.8%) patients had single leftventricle and 29 (49.2%) had single right ventricle (48.2% of thesepresented with hypoplastic left heart syndrome [HLHS]). Patients with singleleft ventricle had a higher percentage of reintervention-free survival, butwithout statistically significant difference. 40% of the patients had nocomplications and 35% of them presented with thrombosis at some point in thefollow-up period, with ventricular dysfunction being the second mostfrequently found complication (15% of cases), mainly among patients withsingle right ventricle morphology (Except for the right ventricular morphology, no other factor has been shownto interfere in late evolution after total cavopulmonary shunt. Single ventricular hearts correspond to a group of congenital heart diseases, withdifferent anatomical combinations that culminate in a common characteristic: asingle ventricle responds by both the systemic and pulmonary circulations. In mostcases, it is possible to confirm the presence of two distinct ventricles, right andleft, one being hypoplastic and called rudimentary and the other being well formedand called the main chamber.One of the most common anatomic forms is the tricuspid atresia, with differentpresentations, which is responsible for 1.5-3% of congenital heart diseases and ispresent in approximately 0.6 out of every 10,000 live births. Another dominant leftventricular presentation is the pulmonary atresia, with intact interventricularseptum, which accounts for about 1% of congenital heart diseases, occurring in about0.7 out of every 10,000 newborns. Regarding to the dominant right ventricle, themain anatomical representation is the hypoplastic left heart syndrome (HLHS), whichrepresents about 2% of congenital heart diseases, being present in approximatelyfour out of every 10,000 newborns.The natural history of single ventricular hearts is quite unfavorable: 64% of thepatients were diagnosed during childhood and 50% in the neonatal period.[1], as a final stage.Treatment is usually performed through surgical staging, with interventions beginningin the neonatal period and culminating in the completion of a total cavopulmonaryoperation (Fontan)The total cavopulmonary bypass allows close-to-normal saturation to these patients,decreasing the demand of the systemic ventricle. Despite this, it maintains alteredhemodynamics leading to a series of late complications, among them, physicalcapacity reduction, ventricular dysfunction, intra- or extracardiac thrombi,arrhythmias, protein-losing enteropathy (PLE), and cirrhosis.We carried out this study in order to evaluate the complications presented in thepopulation submitted to total cavopulmonary bypass, with late follow-up at ourinstitution, and to assess factors that may predispose to these complications, inaddition to estimate the reintervention-free survival.To evaluate the late evolution of patients submitted to totalcavopulmonary derivation with extracardiac conduit, with respect tocomplications  presented and the need for reintervention.To evaluate factors that may interfere in the occurrence ofcomplications or the need for reintervention: diagnosis, age oftotal cavopulmonary bypass, pulmonary vascular resistance andpressure, use of acetylsalicylic acid (ASA) or oral anticoagulation,and fenestration.To evaluate signs of congestion and/or hepatic parenchyma alterationafter five years of total cavopulmonary shunt through abdominalultrasonography (USG).This is a retrospective study using a chart analysis of 59 patients followed-up atthe Single Ventricle Heart outpatient clinic of the Hospital doCoração . To do so, we considered allpatients undergoing total cavopulmonary bypass between 2000 and 2014. Patients withearly death  and those under ambulatory follow-up at other institutionswere excluded.Anatomical and surgical data were obtained through a medical record survey conductedduring in-hospital period and then outpatient follow-up.Regarding the data of the surgical moment, we evaluated: diagnosis, stagingperformed, age of total cavopulmonary derivation, fenestration, pulmonary arteryangioplasty/stenting, preoperative pulmonary pressure, preoperative pulmonarystrength, and need for reintervention.Concerning the late follow-up, we evaluated the occurrence of main latecomplications, such as: thrombi, arrhythmias, ventricular dysfunction, PLE, plasticbronchitis, and changes in hepatic parenchyma through abdominal USG.Considering main late complications, we tried to identify factors that couldinterfere in the occurrence of complications and reintervention-free survival.[2] or Fisher's exact test was used.Initially all variables were analyzed descriptively. Analysis of quantitativevariables was done by observing the minimum and maximum values and calculatingmeans, standard deviations and median. Absolute and relative frequencies werecalculated for qualitative variables. To test the homogeneity between proportions,chi-square test[2]. The software used for the calculations was SPSS17.0 for Windows(r).The study of time until reintervention was performed through Kaplan-Meyer curve withlog-rank testThe level of significance used for the tests was 5%.During the study (from 2000 to 2014), 150 total cavopulmonary shunt operations wereperformed with an extracardiac conduit. Of these, 59 patients were followed-up atthe Single Ventricular Heart outpatient unit of the Hospital doCoração.These patients' ages varied from 10 to 186 months , being 40 (70.2%) males and 17(29.8%) females.At the time of the study, follow-up time ranged from 10 days to 145 months .The distribution of patients being followed-up at our institution by diagnosis isdetailed in Among the patients with HLHS, 13 were submitted to the hybrid procedure (branchpulmonary arteries banding and stent implantation in ductus arteriosus) and only onewas submitted to Norwood-Sano reconstruction as the first stage.Fenestration between the extracardiac conduit and the right atrium was performed in22 (37.2%) patients and the mean pulmonary artery pressure (MPAP) evaluated bypreoperative catheterization was > 15 mmHg in 20 (33.8%) patients.P=0.31), two had a diagnosis of HLHS(P=1.00), and four had MPAP > 15 mmHg in the catheterizationperformed prior to total cavopulmonary bypass (P=1.00); none ofthese three factors could be related to the need for postoperative reintervention.When comparing it with the intervention-free evolution showed in Reintervention was necessary in 12 patients during follow-up for the reasons listedin Among the patients evaluated, 31 (52%) presented with at least one of thecomplications considered; the complications found in late evolution are distributedas shown in P=0.54), themost common site being the extracardiac tube.Two patients have died during the follow-up. However, among the 20 patients whopresented with thrombosis at sometime of the follow-up, 50% of them were in thegroup of patients treated with anticoagulant therapy (P=1.00); one patient had HLHS(P=1.00) and two patients were in the group with preoperativepulmonary pressure > 15 mmHg (P=0.6). Therefore, none of thesefactors can be related to the PLE occurrence in the postoperative period.Regarding the PLE, it occurred in five patients (8.4%), two of them in the group withfenestrated tube (P=0.49); 77% had aright ventricle (P=0.04) and two had pulmonary pressure > 15mmHg (P=1.00). Right ventricular morphology was the only factorthat alone had statistically significant relationship with ventricular dysfunctionin the outpatient follow-up of these patients.As secondary findings, ventricular dysfunction was present on echocardiogram in nine(15%) patients, five of them had fenestration (P=0.057),60% presented with pulmonary pressure > 15 mmHg and in 54% of them the tube wasnot fenestrated (P=1.00).In our institution, it is part of the postoperative follow-up protocol of patientssubmitted to total cavopulmonary bypass the abdominal USG examination at thepostoperative fifth year to search for parenchymal alterations and/or signs ofcongestion justifying further investigation of possible hepatic cirrhosis. Amongpatients currently undergoing follow-up, 20 (35%) have been evaluated by USG and 10(50%) presented with some degree of hepatic congestion and/or hepatomegaly. Betweenpatients with this alteration, 16.7% of them had HLHS , the difference in morbidity and mortality was mainly due tothe occurrence of arrhythmias: the use of extracardiac tube to derive the flow fromthe inferior vena cava to the right pulmonary artery reduced significantly thisoccurrenceThe use of the extracardiac polytetrafluoroethylene (PTFE) tube in all cavopulmonarybypass surgeries at our service without atrial scarring and without altering theatrium architecture justifies the absence of arrhythmias in the follow-up group atour center.[4], especially in the extracardiac tube and with nodifference between the groups using ASA or oral anticoagulant, similar to a findingin a multicenter study published in 2011, which has found a 21% occurrence ofthrombosis in the ASA group and 24% in the oral anticoagulantgroup[5].Currently, at the Hospital do Coração, all patients undergoing totalcavopulmonary shunt are maintained using subcutaneous enoxaparin sodium, dose of 1mg/kg/dose, 12/12 hours, until stabilization and possibility of oralanticoagulation. Oral anticoagulation is the prophylaxis of choice for thrombosis inthese patients in the first two postoperative years and it is not performed inpatients with social contraindication or drug intolerance. After two years, in theabsence of episodes of thrombosis, all patients started to use ASA as prophylaxis,indefinitely. During follow-up, all patients underwent transesophagealechocardiography to evaluate the possibility of thrombi in the Fontan circuit, infive days, 90 days, 180 days, nine months, one year, 18 months, and two yearspostoperatively; after two years, they were evaluated once a year or whenever it wasnecessary. Thrombosis occurred in 35% of the patients, a higher occurrence than itis usually shown in the literature: 0-16% and 4.6%, according to somereports[6].Another prospective study carried out in our department followed for two years agroup of 40 patients submitted to cavopulmonary bypass, 20 in use of antiplateletagent and 20 in use of oral anticoagulant, evidencing the absence of a statisticallysignificant relation in the occurrence of thrombosis between the two groups, butshowing a trend towards a greater deposit of material on the wall of theextracardiac tube, which was evidenced by computed tomography, in patients takingASA.PLE is a serious event; its occurrence is found in most of the literature rangingfrom 1% to 5% of the patients and it has a 50% mortality rate in 5yearsOf the patients followed-up in our center, 7% developed with such complication. Amongthem, one required surgical intervention for the correction of major mitral valveinsufficiency, with resolution of the condition. In a second case, we chose toperform a fenestration, and the child evolved to death postoperatively. Since thebeginning of the use of oral budesonide in our service for the treatment of PLE, allother patients had this condition resolved, maintaining a serum level of albumin> 3 g/dL.[9].Regarding the ventricular dysfunction, it is a constant concern in the late follow-upof patients with univentricular heart morphology, because in addition to the cardiacinsufficiency manifested in patients with ventricular dysfunction of any nature, itmay compromise the operation of the Fontan type circuit, leading to pulmonaryvenocapilar congestion. Its occurrence was higher in patients with single ventricleof right morphology in our series and because of this the concern to preserve thisventricle is greater since the beginning of the follow-up. A study based on recordsfrom Australia and New Zealand of patients undergoing Fontan surgery (1268 in 2015)showed 7% of patients on (angiotensin converting enzyme) ACE inhibitors in order topreserve normal function and 36% systolic or diastolic dysfunction of the singleventricle. The use prevailed in patients with right ventricular morphology(70%).Among the patients currently under follow-up, 35% have already underwent abdominalUSG and 50% of them have alterations characterized by hepatic congestion. Betweenthose, 16.7% of the alterations are in patients with HLHS (P=0.057). Hepaticimpairment has been widely studied these days, being found from congestion to liverfibrosis and cirrhosis, changes that are directly related to the time since totalcavopulmonary derivation.Current studies have shown a concern in proposing a protocol to evaluate liverchanges in relation to periodicity of computed tomography, magnetic resonanceimaging, and liver biopsy in search for precursor changes ofhepatocarcinomaComplications such as thrombi and PLE were present in the assessed group duringoutpatient follow-up, but no notable relation with any of the pre or intraoperativefactors assessed was found, so it remains a challenge to establish the best way toprevent and treat such complications in patients undergoing total cavopulmonaryshunt.No patient so far has evolved with arrhythmias or plastic bronchitis. Among thepatients in follow-up, two had death as an outcome.Among the evaluated causes that could interfere in the complications found, only theright ventricular morphology was statistically significant with ventriculardysfunction.Regarding the hepatic congestion, the abdominal USG has shown signs of hepaticcongestion in 50% of the evaluated cases, and a significant portion of them has beendiagnosed with HLHS.In order to evaluate parenchymal alterations, the indication of computed tomographyand liver biopsy may be considered in patients with sign of congestion.It is important to consider a better evaluation throughout this follow-up, trying toidentify tissue changes, which are possibly precursors of hepatocellularcarcinoma


Zoster Vaccine Live (ZVL) is marketed in the US since 2008, and a non-live adjuvanted Recombinant Zoster Vaccine (RZV) was approved in 2017. Literature suggests that waning of ZVL efficacy may necessitate additional vaccination. The Advisory Committee on Immunization Practices recommended vaccination with RZV in immunocompetent adults aged 50+ years old, including those previously vaccinated with ZVL. The objective of this study was to determine the cost-effectiveness of vaccinating US adults aged 60+ years old, previously vaccinated with ZVL. The ZOster ecoNomic Analysis (ZONA) model, a deterministic Markov model, was adapted to follow a hypothetical 1 million(M)-person cohort of US adults previously vaccinated with ZVL. Model inputs included demographics, epidemiology, vaccine characteristics, utilities and costs. Costs and quality-adjusted life-years (QALYs) were presented over the lifetimes of the cohort from the year of additional vaccination, discounted 3% annually. The model estimated that, vaccination with RZV 5 years after previous vaccination with ZVL, would reduce disease burden compared with no additional vaccination, resulting in a gain of 1,633 QALYs at a total societal cost of $96M . Compared with revaccinating with ZVL, vaccination with RZV would result in a gain of 1,187 QALYs and societal cost savings of almost $84M. Sensitivity, scenario, and threshold analyses demonstrated robustness of these findings. Vaccination with RZV is predicted to be cost-effective relative to no additional vaccination, assuming a threshold of $100,000/QALY, and cost-saving relative to ZVL revaccination of US adults aged 60+ years old who have been previously vaccinated with ZVL. Additional vaccination with RZV at age 60 YOA would yield an ICER of $44,962, compared with of $44,789, $73,720, $75,783 at the ages 65, 70 and 80 YOA, respectively.In the scenario analyses comparing RZV vaccination with no additional vaccination, when we varied the interval between RZV vaccination and the original ZVL vaccination to 1 year and 10 years, ICERs of $107,300 and $32,945 were observed, respectively. Additionally, when effectiveness results of ZVL against PHN as reported by Baxter,5 vaccine coverage has increased slowly each year, and by 2016, about 33% of US adults aged 60+ years old reported receipt of the vaccine.17 However, as ZVL vaccine efficacy waned, additional vaccination was considered and the cost-effectiveness of ZVL revaccination was studied.8 Then following the approval of RZV, ACIP recommended the use of RZV in immunocompetent adults previously vaccinated with ZVL. This is the first post-RZV approval study examining the cost-effectiveness of vaccinating with RZV, compared with either no additional vaccination or ZVL revaccination, basing the analysis on US adults aged 60+ years old who were previously vaccinated with ZVL 5 years earlier. It demonstrated that the RZV vaccine would be cost-effective compared with a no additional vaccination strategy, assuming a hypothetical threshold of $100,000/QALY, and cost-saving compared with ZVL revaccination.Since 2008, when ACIP recommended the use of ZVL for the prevention of HZ and its sequelae,12 The CDC model demonstrated ICERs of less than $60,000 per QALY for all age groups over 60 years of age independent of the time interval between original vaccination with ZVL and vaccination with RZV (i.e. ranging from immediate vaccination (8 weeks after) to 5 years after primary vaccination with ZVL). Our analysis focused on 5 years between vaccinations in the base-case, with an ICER of $58,793, compared with $107,300 and $32,945 when vaccinating 1 or 10 years apart respectively. When determining the optimum time for vaccinating an individual previously vaccinated with ZVL, the person’s age and time since receipt of ZVL should be considered. Since clinical trials of ZVL indicate lower efficacy in older adults, a shorter interval than studied for the population in our analysis may be considered.17Our conclusions are in line with the model independently developed for the Centers for Disease Control and Prevention (CDC) and presented at the October 2017 ACIP meeting.5 Healthcare professionals may be slower in prescribing anti-viral therapy if a patient were vaccinated, compounding the quality of life detriment.20 Also, to maintain vaccine acceptance, persons should have confidence in a vaccination program.20 It is implicit therefore that patients should be protected when they are at greatest risk, pre-empting in the case of HZ, the need for additional vaccination. Healthcare professionals themselves may feel an obligation to offer additional vaccination so as to not misplace the trust patients placed in them to be protected from HZ and its complications when they first received HZ vaccination. The ACIP recommendations allow for this, stating health professionals can “Administer 2 doses of RZV 2–6 months apart to adults who previously received ZVL at least 2 months after ZVL”.21Continued protection of the early adopters of HZ vaccination is important for several reasons. As a person ages, their risk of HZ as well as PHN and other complications increases. Additionally, older adults with less physiological reserve and already taking multiple medications for pre-existing chronic conditions may be less able to tolerate HZ and its sequelae.11 and, where those trials could not provide sufficient data, the Shingles Prevention Study clinical trial for ZVL.1 The ZOE trials reported efficacy to year 4, therefore because of the lack of long-term data, estimates on waning had to be extrapolated. Although the results were robust to a variety of sensitivity and scenario analyses, real-world RZV effectiveness should be studied further as only in population settings can the long-term duration of protection, adherence to the 2-dose schedule, and the effectiveness and duration of protection in case of non-compliance to the dose series and schedule be elucidated.As with every modeling analysis this study has limitations, however with the model design and inputs, intentional decisions were made to minimize unlikely real-world results. The impact of RZV on HZ incidence in the analyses, both in terms of initial protection and maintenance of that protection over time, were based on efficacy data from the RZV ZOE-50 (NCT01165177) and ZOE-70 (NCT01165229) clinical trialsUsing RZV to vaccinate US adults 60+ years old who were vaccinated 5 years earlier with ZVL, is predicted to be cost-saving compared with ZVL revaccination, and cost-effective when compared with no additional vaccination. Since vaccination with ZVL has been recommended by ACIP for over 10 years in the adults aged 60+ years old, and the effectiveness of vaccination wanes, the aging early adopters of vaccination are becoming increasingly vulnerable to HZ and its complications. This new analysis can be valuable to healthcare professionals, public health decision makers and payers contemplating improving the quality of care of the elderly.22 to include prior vaccination. The adapted ZONA model is a deterministic multi-cohort Markov model including four hypothetical cohorts split into age groups for people aged 60+ years old  with age groupings consistent with those used by the US Census Bureau.23 The model, with annual cycle lengths, follows all subjects within a cohort over their remaining lifetimes from the year of vaccination, with all-cause mortality rates taken from the CDC National Center for Health Statistics.24 Lifetime outcomes are considered to capture the full effects of HZ vaccination through reduced morbidity. Three different HZ vaccination strategies are compared: no additional vaccination, revaccination with ZVL, and vaccination with RZV. In this analysis all individuals in each cohort have been previously vaccinated against HZ using ZVL, as such they enter the model having residual protection based on initial efficacy of ZVL followed by linear waning. HZ incidence, probability of PHN, and initial efficacy of RZV were modeled using linear rather than step functions to increase consistency between vaccination strategies. Otherwise all input values and assumptions were unchanged.For the analysis we adapted the previously published ZOster ecoNomic Analysis (ZONA) model14 As with the CDC model, presented at ACIP in 2017, our model included direct medical cost input parameters based on a population based study, which reported healthcare utilization and costs due to HZ, PHN and other HZ complications.25 Indirect costs per HZ case were based on absenteeism and presenteeism-related work loss hours due to HZ based on a telephone survey of HZ patients.26 Utilities were calculated from baseline values for individuals without HZ and disutility associated with HZ. Additionally, individuals who are vaccinated have a risk per vaccine dose of experiencing AEs, which result in additional direct costs , indirect costs (working hours lost), and QALYs lost. The model includes four possible vaccine-related AEs: local/general reactions, outpatient visits, emergency room visits, and hospitalizations.In the model, vaccinated individuals incur costs for the vaccine price and administration cost. Vaccination reduces the probability of getting HZ and, given a case of HZ, reduces the QALY loss resulting from HZ. Individuals who experience HZ may experience complications. The HZ complications included in the model are PHN as well as ocular, neurological, cutaneous, and other non-pain complications. Recurrent HZ and complications may also occur. Probabilities of moving between health states are age-group–specific and are derived from the US-based peer-reviewed literature, as previously published.In the base-case analysis we evaluated the ICER, in terms of cost per QALY gained, vaccinating a cohort of 1 million individuals aged 60+ years old in the US population who had been previously vaccinated 5 years prior with ZVL, compared with no additional vaccination. The primary perspective was the societal perspective so as to include both direct medical costs and indirect costs. Costs and outcomes were presented over the remaining lifetimes of individuals. Life years, QALYs, and costs (in US dollars 2016) were discounted at 3% per year, consistent with a lifetime perspective.22 and are summarized in Table S2. Vaccine efficacy inputs for ZVL and RZV were obtained from clinical trials. Additional efficacy of ZVL against PHN cases was included; no additional efficacy of RZV against PHN was included. 12 In the absence of data for one dose of RZV, waning at the same rate as for ZVL was assumed. Vaccination coverage rates were assumed to be 100% for ZVL and RZV first-dose. Compliance with the second dose of RZV  was 69%, based on the vaccination series completion and compliance rates of hepatitis A and B among US adults.27 US-specific (year 2016) HZ-related and vaccine program costs, including management of AEs, were used as previously described.14 The base-case RZV price per dose was $140 compared with ZVL $196.91.28 Baseline utility values for the US population were taken from Janssen & Szende 2014,29 QALY loss per HZ case by vaccination status and PHN status were as reported in Pellissier.30Model inputs have been described in detail elsewhereA deterministic sensitivity analysis was conducted with ranges informed by published data where available. The results of the DSA were summarized in a tornado diagram. A probabilistic sensitivity analysis was conducted to consider the impact of the full uncertainty across model inputs on the ICERs for vaccination with RZV (a) versus no additional vaccination, and (b) versus ZVL revaccination. The results from 5,000 Monte Carlo simulations for each analysis, where inputs were simultaneously sampled from probability distributions (gamma for costs and beta for the other varied inputs), were presented on a cost-effectiveness plane, with lines illustrating hypothetical willingness to pay thresholds. The results from the PSA for RZV versus no additional vaccination were also presented as a cost-effectiveness acceptability curve.Threshold analyses were conducted to investigate, for a selected set of key inputs , the values that those inputs could hold and still maintain an ICER for RZV vs. no additional vaccination below various hypothetical willingness-to-pay thresholds ranging from $0 to $160,000 per QALY gained.et al. 201619 where the first-year HZ effectiveness of 69% dropped substantially to 4% in year 8, and Baxter et al. 201618 where the first-year PHN effectiveness of 83% was better sustained, falling to 70% in year 5; details are included in Table S1.Finally, we conducted scenario analyses. Firstly, in place of 5 years between the vaccination with ZVL and RZV, we studied 1 and 10 years. Secondly, we varied ZVL efficacy in line with data from two studies Tseng Zostavax is a trademark of Merck Sharp & Dohme Corp.Shingrix is a trademark of the GSK group of companies.


Human Immunodeficiency Virus (HIV) infection has become a global concern. Determining the factors leading to death among HIV patients helps controlling Acquired Immune Deficiency Syndrome (AIDS) epidemic. Up to now, little is known about mortality and its determinants among people living with HIV in the Middle East and North Africa (MENA) region, including Iran. The purpose of this study was to assess the risk factors of AIDS-Related Mortality (ARM) and Non-AIDS-Related Mortality (NARM) among people with HIV in Iran.This 20-year retrospective study was conducted on 1160 people with HIV whose data were collected from 1997 to 2017. The association of the study outcomes (ARM and NARM) with various study variables, including demographic status at the time of diagnosis and clinical indexes during the follow-up were examined to define the predictors of mortality among the patients. Regarding, Cox proportional hazard and competing risk models were fitted and Adjusted Hazard Ratios (AHR), Sub-distribution Hazard Ratio (SHR) and the 95% Confidence Intervals (CI) were reported.during the follow-up period, 391 individuals (33.7%) died with 86,375 person-years of follow-up. Of the total deaths, 251 (64.2%) and 140 (35.8%) were ARM and NARM, respectively. Rates of the mortality caused by AIDS and non-AIDS were 3.2 and 4.5 per 1000 person-months, respectively. Responding to combined Antiretroviral Treatment (cART) 6 months after initiation, receiving Pneumocystis Pneumonia (PCP) prophylaxis, and higher CD4 count at diagnosis, reduced the hazard of ARM and NARM. However, older age, late HIV diagnosis, and last HIV clinical stages increased the hazard of AIDS related to mortality. Additionally, male gender, older age, incarceration history, and last HIV clinical stages increased the non-AIDS mortality.Mortality caused by AIDS and non-AIDS remains high among people with HIV in Iran, particularly among males and those with late diagnosis. It seems that applying effective strategies to identify infected individuals at earlier stage of the infection, and targeting individuals with higher risk of mortality can decrease the mortality rate among HIV infected people. Human Immunodeficiency Virus (HIV) is still a major public health issue in all around the world. Since the beginning of the AIDS epidemic, approximately 78 million people have been infected by HIV, and 35 million people have died because of AIDS-related diseases . In the 2 (200–350 cell/mm2 before 2015). However, since 2019, the treatment regimen has been changed and HAART must currently be started as soon as the individual is diagnosed with HIV infection regardless of the level of CD4 [HIV is also a public health challenge in Iran. According to the latest estimates of the United Nations Program on HIV/AIDS (UNAIDS), a total of 61,000 people ) are living with HIV in Iran until 2018 . Out of l of CD4 .The importance of analysis of data on death among PLWH has been shown in several studies. The results of such studies could identify the strengths and weaknesses of health policies for treatment of HIV patients, to achieve a longer time of survival –19. The Recent studies have indicated that the main causes of death among PLWH are co-infections, such as oncogenic viruses, Hepatitis B Virus (HBV), Hepatitis C Virus (HCV), tuberculosis, medication-related toxicities, and consumption of illegal or recreational drugs. Moreover, several cohort studies have demonstrated that HIV/AIDS-related conditions are the most frequent causes of death , 3. HoweIn the present study, HIV patients’ survival was investigated among the patients who were registered in Shiraz HIV/AIDS Research Center during 1997–2017. With respect to the standard guidelines, the causes of death among HIV patients are classified to ARM and NARM , 11, 24.This follow-up study was conducted at Shiraz HIV/AIDS Research Center, a referral center for individuals living with HIV in Fars province, Iran. Nearly 5000 patients were diagnosed at this center and almost 1160 cases followed their ART line with complete medical files. Data were recorder from all patients at this referral center from August 1997 to May 2017 and were extracted to a checklist. The checklist was completed by trained staff in terms of the World Health Organization’s (WHO) recommendations  protocol . The CoDTwo sets of covariates were considered. The first set of covariates was evaluated at diagnosis, which included age (continuous), gender , education level , marital status , employment status , self-reported mode of HIV transmission , etc.), and CD4 count (continuous). The other set of covariates was measured after diagnosis , which included drug abuse , drug use), linkage to HIV care defined as having visited centers providing HIV care and having access to ART and medical care after being diagnosed with HIV , response to ART defined as indicating an improvement in clinical factors and/or increased CD4 counts by passing 6 months from ART initiation based on the medical records and physicians’ feedbacks , late HIV diagnosis defined as less than 1 year between HIV diagnosis through an advanced phase of AIDS , taking Pneumocystis Pneumonia (PCP) prophylaxis , having HCV co-infection , having HBV co-infection , having tuberculosis co-infection , and AIDS clinical stage defined based on WHO’s criteria (stages 1 and 2 vs. stages 3 and 4).p-values less than 0.20. All data analyses were carried out using Stata, version 14.Descriptive statistics were reported as mean, Standard Deviation (SD), median, and interquartile range  for continuous measures, and as absolute and relative frequencies for categorical ones. In order to accomplish survival analysis, Cox model and competing risks analysis method were used to apply Hazard Ratio (HR) and Sub-distribution Hazard Ratio (SHR) for both groups of death events as they were proposed by Fine and Gray (1999) . At Firsn = 857, 73.9%). The transmission route was mainly injecting drugs in males  and sexual contact in females . All descriptive statistics of the study variables have been presented in Table n = 241, 96%) were accounted for the main causes of ARM. Among these cases with NARM, the most frequent cause of death was external causes , including overdoses and driving accidents. Moreover, 14 deaths (7%) were related to cancer as shown in Table In this study, a total of 1160 AIDS patients were enrolled due to the inclusion criteria. The mean age of the patients at diagnosis was 35.22 years old (SD = 8.43). Additionally, 1109 patients (95.6%) had not finished the compulsory education. Besides, the majority of them were male . As most causes of death were AIDS related, ARMs had a similar trend to all-cause mortality. However, NARM remained fairly stable with a slight increase at the end of the study (from approximately one death/100 person-years to two deaths/100 person-years) Fig. .Fig. 2Mp-values more than 0.2. Therefore, they were excluded from all three multivariate models (Table p > 0.2) and consequently were removed.Based on the results of survival analysis on baseline characteristics, two variables; i.e., education level and employment status, had ls Table . Moreovep < 0.001). Actually, based on the “response to treatment”, ARM and NARM hazards were reduced by 91%  and 86% , respectively. Similarly, “PCP prophylaxis” reduced all-cause mortality by 57% , ARM by 67% , and NARM by 36% . Also, “Last clinical stage” was significantly associated with both ARM and NARD; however, it has no relationship with all-cause mortality. In addition, “stages 3 and 4” were associated with a higher risk of ARM  and a lower risk of NARM . Late HIV diagnosis, age, and CD4 count at diagnosis were identified as significant risk factors for ARM and all-cause mortality, but not for NARM. Each unit increase in baseline CD4 count declined all-cause mortality by 0.1%  and ARM by 0.2% . Additionally, 1 year increase in age of diagnosis, caused a 4% increase in all-cause mortality  and a 2% increase in ARM risk . Moreover, “late HIV diagnosis” was the most effective risk factor because it increased all-cause mortality and ARM risk by four and about three folds, respectively  . Furthermore, incarceration history increased the all-cause mortality by about 70% , and the risk of NARM by two folds . Finally, gender  was a significant risk factor only for NARM . Accordingly, overdose most frequently caused NARMs. However, most NARMs originated from non-AIDS malignancies, CVD, and liver diseases in developed countries. Indeed, ART has been reported to be more effective in developed countries where HIV patients receiving ART were shown to survive as long as the normal population do. As a result, the mean age in NARMs can be higher than that in ARMs [n = 56, 71% of external deaths). In fact, drug abuse has reduced the survival of HIV patients through raising NARMs even prior to the AIDS stage. In consistent with that, a recent study showed an association between serum albumin and NARM events and suggested that the association was stronger for smokers [The results of this study demonstrated that not only ARM is still a major cause of death, but also NARM poses a threat to PLWH. This is in agreement with other studies, as well , 27. It  in ARMs , 30. How in ARMs , the pre smokers .Previous studies indicated a rapidly increasing number of drug users in Iran from 1998 to 2003 –35. ThisIn the present study, almost 34% of the patients died during the study period, and about two out of every three deaths were related to opportunistic and AIDS-defining infections with a history of addiction. These findings indicated that addiction had a significant effect on the causes of death and survival of HIV patients in south of Iran regions. On the contrary, several studies from other countries have reported other factors as the major causes of death , 3, 28. 2 were the main reasons for NARM [Based on the current study findings, late HIV diagnosis in older ages, in stages 3 or 4, or with a lower CD4 count could significantly increase the risk of ARM. These results were in line with the findings of other studies , 37. Thefor NARM .The present study findings demonstrated a significant improvement in detection and treatment of HIV patients. However, the 90–90-90 targets have not been fulfilled in Iran . In 2010The strengths of the present study were using a relatively large sample and making use of competing risks model for performing statistical analysis of the data. However, limitation of this investigation were incomplete medical files, especially about viral load check before 2011. Another study limitation was the restricted access to the patients’ information, especially for the decreased ones. Another important limitation of this study was that, like several other diseases , 40, theThe results of the present study indicated that both ARM and NARM are substantially high among people with HIV in Iran. Applying effective strategies is needed to achieve on time diagnosis of individuals with HIV and provide them with HIV care and treatment services to enhance the survival of the patients. Moreover, males living with HIV require more attention when receiving HIV care and treatment as they experience a higher risk of NARM. In line with the available evidence, the findings of the current research revealed that early HIV care and treatment could substantially reduce ARM and NARM among HIV patients. Therefore, to reduce mortality rate among people with HIV, the current strategies should be revised to improve the timing of treatment initiation and also to optimize the adherence to the treatment.


All ischemic stroke cases were validated and categorized into subtypes. The lipophilic index was calculated based on fatty acid profiles in adipose tissue. Subjects were divided into quintiles and a weighted Cox proportional hazards regression model was used to calculate hazard ratios. After appropriate exclusions, a subcohort of 3194 subjects and 1752 cases of ischemic stroke were included. When comparing the fifth quintile of the lipophilic index with the first quintile, the hazard ratio for ischemic stroke was 0.92  and the trend across quintiles was not statistically significant (p = 0.1727). In conclusion, no association was found between the lipophilic index and ischemic stroke or its subtypes.Diet may influence the risk of ischemic stroke by several mechanisms. A potential and hitherto unknown mechanism may relate to an effect on the lipophilic index, which is a new and convenient indicator of membrane fluidity. This study investigated the association between the adipose tissue lipophilic index and ischemic stroke and its subtypes. A case-cohort study was conducted based on the Danish cohort study Diet, Cancer, and Health, which includes 57,053 subjects aged 50–64 years at enrolment. A subcohort ( Ischemic stroke is the second most common cause of death from vascular disease and the third leading cause of disability worldwide . StudiesThe lipophilic index is a weighted mean of the melting points of fatty acids and has been proposed as a simple method of determining overall membrane fluidity. The lipophilic index has previously been found to be associated with myocardial infarction , coronarThe aim of the study was to investigate the association between the adipose tissue lipophilic index and incidence of ischemic stroke. Furthermore, we examined whether associations differed between ischemic stroke subtypes.This study was a case-cohort study within the Danish cohort study Diet, Cancer, and Health. Between 1993 and 1997, 160,725 men and women aged 50–64 years were invited to participate in the study and the inclusion criteria were as follows: born in Denmark, living in the Copenhagen or Aarhus areas, and with no previous cancer diagnosis registered in the Danish Cancer Register. A total of 57,053 persons accepted the invitation. A subcohort of 3500 subjects was randomly drawn from within the Diet, Cancer, and Health cohort. At baseline, all cohort subjects completed both a lifestyle questionnaire and a food frequency questionnaire ,16. Basen = 569), stroke (n = 582), or missing questionnaire (n = 42) and, furthermore, cases and subjects from the subcohort were excluded if their fatty acid profile was incomplete (n = 335) or there were missing variables (n = 44). Previously, knowledge of more than 95% of total fatty acids has been suggested as an adequate proportion to represent a fatty acid composition , “MPi” is the melting point of the i-th fatty acid (°C), “k” is the number of fatty acids included in the calculation, and “i” represents the individual fatty acid [The lipophilic index was defined as the mean melting points of the fatty acids weighted according to the fatty acid composition (Equation (1)) ,9.(1) LItty acid ,9.Melting points were obtained from LipidBank, the official database of the Japanese Conference on the biochemistry of Lipids . Fatty aThe outcome of the study was ischemic stroke defined according to the World Health Organization’s definition as a rapidly developing disease with focal or global neurologic deficit of vascular origin which lasted more than 24 h . The cohSubjects were followed until the date of a diagnosis of ischemic stroke or censoring, which was the date of death not related to ischemic stroke, date of emigration, or end of follow-up (30 December 2009). This case-cohort study assessed the association between the lipophilic index as the exposure and incident ischemic stroke and additionally ischemic stroke subtypes as outcome. Since no generally accepted categorization of the lipophilic index existed, the association was explored by means of quintiles. To achieve quintiles representative of the background population, quintiles were defined based on the lipophilic index of the subcohort and cases added accordingly. Weighted Cox proportional hazards analysis was applied to calculate hazard ratios with 95% confidence intervals (CIs), with graphical evaluation of the association performed by exploring cubic splines of the lipophilic index. p-values were two-sided and we considered values <0.05 to be statistically significant. All analyses were conducted using Stata 15 .Age was used as the underlying time scale, and to account for the overweight of cases, the Cox regression analyses were weighted using the method of Kalbfleisch and Lawless . Hazard Adjusted models were constructed to evaluate the association between the lipophilic index and ischemic stroke. Potential confounders were included in three different adjusted models as follows: Model 1a included sex and age at baseline, where sex was included allowing for different baseline hazards while assuming equal associations for exposures . Model 1A subject was considered to have diabetes mellitus, hypertension, or hypercholesterolemia when the condition was self-reported on the questionnaire or treatment with insulin, antihypertensive, or lipid-lowering drugs was reported. Furthermore, diabetes mellitus was noted if the subject was registered in the National Diabetes Registry at baseline and hypercholesterolemia was noted if the baseline total s-cholesterol measurement was above 5 mmol/L. For the restricted cubic splines, continuous variables were modelled using three knots and for comparability purposes, the reference value was the average lipophilic index of the first quintile.After having applied the exclusion criteria, the study population consisted of 1752 cases of ischemic stroke and a subcohort of 3194 subjects, of whom 113 experienced an ischemic stroke during follow-up. Cases were further divided into ischemic stroke subtypes, including 300 cases of large artery atherosclerosis, 779 cases with small-artery occlusion, 99 with cardioembolism, 91 cases of stroke of other etiology, and 483 strokes of unknown etiology . CompareFurthermore, baseline characteristics when distributed across quintiles of the lipophilic index showed that subjects in the first quintile, who had the lowest lipophilic index, were more likely to have a higher body mass index, a lower level of physical activity, a lower intake of alcohol, hypertension, and atrial fibrillation/flutter than the subjects in the fifth quintile .n-6, 20:2 n-6, 20:3 n-6, and 20:4 n-3. A mean melting temperature of trans fatty acids 18:1 ∆6–8 was applied collectively, as differentiation between the trans fatty acids on the gas chromatography was not possible.The adipose tissue samples contained up to 34 fatty acids, but melting points were only available for 31 fatty acids, and therefore three fatty acids with unknown melting points were omitted. The sum of fatty acids still included more than 96% of total fatty acids. The included fatty acids were saturated fatty acids 12:0, 14:0, 15:0, 16:0, 17:0, 18:0, 19:0, 20:0, and 22:0, monounsaturated fatty acids 14:1 n-5, 16:1 n-7, 18:1 n-7, 18:1 n-9, 20:1 n-9, 20:1 n-11, 22:1 n-9, and 22:1 n-11, polyunsaturated fatty acids 18:2 n-6, 18:2 n-9, 18:3 n-3, 18:3 n-6, 20:4 n-6, 20:5 n-3, 22:5 n-3, and 22:6 n-3, and trans fatty acids 18:1 ∆6–8, 18:1 ∆-9, 18:1 ∆-10, 18:1 ∆-11, 18:2 ∆ct, and 18:2 ∆tc. The omitted fatty acids were the following polyunsaturated fatty acids 22:4 p = 0.1727).The hazard ratios for an association between the lipophilic index and total ischemic stroke did not indicate a clear trend across quintiles either . In the Regarding subtypes of ischemic stroke, there was no clear association between the lipophilic index and large artery atherosclerosis. The lipophilic index was associated with lower rates of small-artery occlusion and cardioembolism, although this association was not statistically significant, and when comparing the fifth quintile with the first quintile the hazard ratios were 0.95  and 0.69 , respectively.In this large case-cohort study, which included 1752 cases and a subcohort of 3194 subjects, no clear associations between the adipose tissue lipophilic index and the occurrence of total ischemic stroke as well as subtypes of ischemic strokes could be demonstrated.When considering the functionality of the cell membrane, it should be able to withstand mechanical stress but, at the same time, not be too rigid to respond to cell stimuli; thus, a u-shaped association between the lipophilic index and ischemic stroke would be reasonable. However, this study showed no association between the adipose tissue lipophilic index and total ischemic stroke, nor its subtypes.Strengths of our study include the study size of 1752 cases of ischemic stroke, with each case having been individually validated independently of the lipophilic index at baseline. Random error is thus not a likely explanation of our study results. Furthermore, each case was categorized according to the TOAST classification, making it possible to investigate stroke according to underlying etiologies. Also, the lipophilic index was calculated based on adipose tissue biopsies, which are objective measures of long-term fatty acid intake and endogenous exposure. Moreover, in the adjusted models, sex, age, and socioeconomic and lifestyle factors, as well as medical history, were added in steps to make it possible to differentiate between the impact of the potential confounders and to ease the interpretation of the results.The study also had some limitations. In general, the participation in the cohort was only 35% of those invited. However, it is unlikely that questionnaire answers and fatty acid analyses were affected by any systematic error related to the association between lipophilic index and ischemic stroke. Additionally, only middle-aged persons, primarily Caucasians, who had survived without a diagnosis of cancer or ischemic stroke until inclusion were included, hence the results may not be applicable for other age and ethnic groups. The adipose tissue biopsies were only obtained at baseline and changes during follow-up, which would influence the fatty acid composition, were not captured. Furthermore, medical history at baseline did not include pre-hypertensive or pre-diabetic subjects. The untreated subjects may be at higher risk of developing ischemic stroke. However, we did not adjust for changes in medical history in our analysis.Dietary patterns such as Mediterranean, and Dietary Approaches to Stop Hypertension (DASH) have been shown to be associated to the risk of ischemic stroke . A previOnly a few other studies regarding the lipophilic index and cardiovascular disease have been published previously. In a case-cohort study of a Dutch population including 2604 subjects and 479 cases with ischemic stroke, no association between the dietary lipophilic index and ischemic stroke or coronary heart disease was found , which iA Costa Rican matched case-control study by Toledo et al.  explorinIn the study by Toledo et al. , the meaSince both ischemic heart disease and ischemic stroke are primarily caused by atherosclerosis, the lipophilic index could, based on previous results, a priori be expected to be associated with ischemic stroke. This hypothesis could, however, not be substantiated by findings in this large study of validated cases of total ischemic stroke or subtypes of ischemic stroke, where we found no association between the lipophilic index and incident ischemic stroke. However, our study was an epidemiological study, which can generate hypotheses based on associations but not prove causality. It is still possible that changing a diet from saturated to more unsaturated fatty acids, which would be expected to raise the lipophilic index, as both monounsaturated and polyunsaturated fatty acids have lower melting points than saturated fatty acids , could cIn conclusion, this case-cohort study of 1752 ischemic stroke cases showed no association between the adipose tissue lipophilic index and ischemic stroke or any ischemic stroke subtype. While diet is important for risk of ischemic stroke, this does not seem to be mediated via cell membrane fluidity as measured by the adipose lipophilic index.


Growing interest in the biological activity of aminobisphosphonates has stimulated the development of methods for their synthesis. Although several general procedures were previously elaborated to reach this goal, aminobisphosphonate chemistry is still developing quite substantially. Thus, innovative modifications of the existing commonly used reactions, as well as development of new procedures, are presented in this review, concentrating on recent achievements. Additionally, selected examples of aminobisphosphonate derivatization illustrate their usefulness for obtaining new diagnostic and therapeutic agents. Bisphosphonates are a class of compounds that are currently receiving significant attention. Over 50 new papers are seen each week when searching the literature with the keyword “bisphosphonate” via Web of Science. More than 17,000 various bisphosphonate structures have been synthesized and described in the literature ,4,5,6,7 Thus, simple and effective procedures for their synthesis are becoming increasingly important. However, only a few general reactions leading to these compounds have been described to date and are only partially reviewed in the literature ,12. NoveIn this paper, we comprehensively review the recent studies (supplemented by older papers if necessary) on reactions applied to synthesize the most important class of bisphosphonates—aminobisphosphonates—and discuss their scope and limitations.There are contradictory reports of the origin of bisphosphonates . Most liThere are only a few general methods for the synthesis of aminobisphosphonates. However, there are many individual procedures described for their preparation ,12. In t1-Hydroxyethylidene-1,1-bisphosphonic acids are perhaps the oldest group of bisphosphonates. They are standardly prepared by a large-scale, one-step reaction of carboxylic acids with phosphorus trichloride and phosphorous or phosphoric acids, followed by hydrolysis with water; the procedure was optimized by Kieczykowski et al. . The rea1). In this case, free unblocked amino acids are used as substrates, and the final neutralization of reaction mixtures to a pH of approximately 4 causes precipitation of the desired products, which are formed in satisfactory yields and are of good purity . Th. Th66]. Simple three-component condensation of stoichiometric ratios of amines, diethyl phosphite and triethyl orthoformate, first reported in patent literature by Suzuki  and furtThe three-component reaction was applied to synthesize a large series of physiologically active compounds, in some cases of very complex chemical structures, including bone antiresorptive drug candidates ,80,81,8224 and 25) or formylation (compound 26) of amine moieties is observed . A. A48, wh50, which resulted in γ-phosphono-α–aminobisphosphonates 51  . S. S64 andA similar monomer, acryloylated pamidronate, has been used to obtain a hyaluronic acid derivative that is dually polymerized with cross-linkable hydrazide groups and bisphosphonate ligands. By mixing bisphosphonate polymer with calcium ions and aldehyde-derivatized hyaluronic acid, a hybrid hydrogel was obtained, which quickly mineralizes . Such a 66, N-(3-aminopropyl) methacrylamide 67, and tetra(ethylene glycol) diacrylate 68) (69 were obtained . A. A57 as Alendronate was also bound to conjugates of pullulan and paclitaxel, which were bound to the polysaccharide by a cathepsin K-sensitive tetrapeptide spacer. This should ensure release of paclitaxel in bones. Then, bisphosphonate was covalently conjugated to the sugar chain via a polyethyleneglycol chain, using a technique similar to that described above (identical to those shown in Aminobisphosphonic acids are gaining significant interest as a class of compounds with promising physiologic activity, with some representatives already commercialized as bone resorption inhibitors, and therefore useful drugs against osteoporosis and related bone disorders. This leads to both the modification of existing and the elaboration of novel procedures for their preparation. There are several commonly used reactions for this purpose; however, they have also been modified in the last decade to be tailored to specific biological needs. A few novel procedures have also been developed. Functionalization of simple aminobisphosphonic acids is a difficult task because of their strongly polar character. However, successful examples of functionalization of the amino moieties of these compounds have provided promising diagnostics and novel therapeutic agents.


Hence, it is time-consuming and expensive, in particular for the prototyping of microfluidic devices in low quantities. This article describes a laser-based process that enables the rapid manufacturing of enclosed micro-structures by laser micromachining and microwelding of two 1.1-mm-thick borosilicate glass plates. The fabrication process was carried out only with a picosecond laser (Trumpf TruMicro 5×50) that was used for: (a) the generation of microfluidic patterns on glass, (b) the drilling of inlet/outlet ports into the material, and (c) the bonding of two glass plates together in order to enclose the laser-generated microstructures. Using this manufacturing approach, a fully-functional microfluidic device can be fabricated in less than two hours. Initial fluid flow experiments proved that the laser-generated microstructures are completely sealed; thus, they show a potential use in many industrial and scientific areas. This includes geological and petroleum engineering research, where such microfluidic devices can be used to investigate single-phase and multi-phase flow of various fluids (such as brine, oil, and CO Microfluidic devices are used across a wide range of applications in many industrial and research areas, primarily in chemistry, biology, medicine, and pharmacology ,6,7,8,9.2 injection and trapping [Microfluidic devices are also used in geological and petroleum engineering research to investigate various processes governing the macroscopic behavior of subsurface systems at a pore level  ,11. Thestrapping ,14,15,16trapping ,18,19,20trapping , and tratrapping , at a’ sMicrofluidic devices can be manufactured from a range of materials, such as SU-8 photoresist, polydimethylsiloxane (PDMS), polymethyl methacrylate (PMMA), polylactic acid (PLA), cyclic olefin copolymer (COC), glass, or silicon. Glass, due to its unique combination of high transparency, hardness, thermal stability, electric insulation, surface stability, chemical inertness, and resistance to acids, is often a preferred substrate for the fabrication of microfluidic devices over silicon and polymers. Unfortunately, the conventional manufacturing of microfluidic devices from glass substrates is a complex, multi-step process that involves different fabrication techniques and tools ,24,25. TMicrofluidic patterns on glass materials are typically generated by either wet  etching or dry (reactive ion) etching. Wet etching of glass involves the use of strong chemicals, such as hydrofluoric acid (HF), to remove the material . This etThe reactive ion etching (RIE) of glass is an anisotropic process that enables the generation of microfluidic patterns with almost vertical sidewalls  . MicrochAlthough both etching techniques enable the precision generation of complex microfluidic patterns on glass substrates ,25, the 2 laser, an ultrafast laser, an excimer laser, or an ultraviolet Q-switched solid-state laser. Current laser systems, which often are integrated with a galvo-scanning system and computer-aided design (CAD) software, enable the generation of complex 2.5-dimensional (2.5D) patterns in which each channel may have a different depth and width. The surface elements of such patterns may be generated with different sets of laser parameters, and thus, they may have different widths as well as depths.Lasers enable the generation of microfluidic patterns on various materials, including glass materials ,30,31,32®33, or Pyrex™ [Ultrafast (femtosecond) lasers can also be used for the generation of truly three-dimensional (3D) microfluidic patterns inside glass materials, such as fused silica, Borofloatr Pyrex™ ,36,37,38In the past, we demonstrated that an ultrafast picosecond laser  can be an effective tool for the direct cutting, drilling, and micromachining of glass plates ,40, and ®33) glass plates with dimensions of 75 mm × 25 mm × 1.1 mm were used for the manufacturing of microfluidic devices. The flatness of the glass plates was ~λ/4. Borofloat®33 contains 81% SiO2, 13% B2O3, 4% Na2O/K2O, and ~2% Al2O3 [Borosilicate  was used in this work. The laser provides ~6 ps pulses ) with a maximum pulse repetition frequency (PRF) of 400 kHz. The model TruMicro 5×50 contains three outputs; each output emits a collimated laser beam of a different wavelength . The output laser beams are expanded and delivered to three separate galvo-scanners using appropriate high-reflection (HR)-coated mirrors. Each galvo-scanner is equipped with an approximately 160-mm-focal-length F-theta lens. The laser beams at the focus have different diameters and M2. The PRF value, in turn, was limited to 100 kHz because higher values could potentially lead to heat accumulation in the material, and consequently, the fusion of glass particles to the material surface [Microfluidic patterns and inlet/outlet ports were generated using the 515-nm wavelength. At this wavelength, the peak laser fluence and the machining resolution were the highest. The maximum fluence used for machining the glass plates was 31.1 J/cm surface .Prior to the laser treatment, the glass samples were cleaned with isopropanol and wiped off with lens tissues. Following the cleaning process, the glass plates were mounted in a holder that provided a clear aperture underneath the laser machining area, while the holder was fixed to XYZ linear stages , as shown in The same picosecond laser processing system was also used for the microwelding of glass plates; however, a different processing arrangement was used. In this case the 1030-nm wavelength was used at the maximum PRF of 400 kHz. The processing arrangement was the same as that described in Reference . The onlThe real challenge in laser microwelding is to bring two glass plates into sufficiently close contact prior to the process. This can be achieved by pressing one plate to the other. Once optical contact is provided, van der Waals forces are capable of holding the two materials together. This, however, requires the glass surfaces to be flat, smooth, and free of debris. The laser-machined glass plates were contaminated with small amounts of dust and glass particles that would prevent optical contact being achieved. Therefore, it was necessary to clean the plates prior to laser welding to ensure optical contact between the two materials. The cleaning was performed with the use of an ultrasonic bath. The glass plates were individually inserted into a beaker filled with methanol, and the beaker was placed into a water-filled tank of an ultrasonic bath, that was operated at room temperature for ~10 min. After the ultrasonic bath treatment, the samples were dried using a jet of ionized nitrogen. This last cleaning step was performed under an air hood that protected the samples from dust. The cover glass plates were cleaned in the same way.Recently, more effective cleaning of the laser-machined samples was achieved using hydrofluoric (HF) acid. In this method, the glass plates are placed into a beaker filled with a 5% HF solution, and are kept in this solution at room temperature for a couple of minutes. This removes any debris resulting from the laser machining process, even glass particles that fused to the glass surface.Following the cleaning process, the glass cover and the laser-machined glass plate are placed into contact, and a force is applied to bring them into optical contact. Such prepared samples are transferred to the holder, while the holder is fixed to XY nano-positioning stages , as shown in The laser-manufactured microfluidic devices were tested using the set-up shown in Initially, the microfluidic channels were filled with air. In the test, deionized (DI) water was injected into a microfluidic device through one of the inlet ports. After filling, the device was inspected for any water leakage. Following the visual inspection of the device, nitrogen was injected in order to remove the water. The maximum injection pressure was 1.5 bar. During these experiments, the flow of fluids was recorded using a digital camera .2 (ablation threshold) is required, together with a PRF < 400 kHz, and a pulse overlap in the range of 85–95%. If the overlap is too small, the result is partial machining and the generation of damage on the back surface of glass, whereas, if the overlap and PRF are too high, the resultant heat accumulated in the material can be sufficient to cause cracking.®, Alicona Ltd., Raaba, Austria). This instrument can measure many surface parameters, including depth, width, and surface roughness, with a vertical resolution down to 10 nm. 2. The PRF value and laser beam scan velocity (v) were 20 kHz and 40 mm/s, respectively. At these laser parameters, the distance between the centers of subsequent laser pulses (Δx) was 2 µm, corresponding to a 90.4% pulse overlap /2 ω0) × 100%, where 2 ω0 is the laser beam diameter).The laser-generated channels were measured and characterized using a 3D surface profilometer , and hatch = 1.8 µm. The depth of this area is 73 µm, and the surface roughness (Sa) is 2.2 µm .Larger areas (including channels wider than 14 µm) can also be generated by the picosecond laser, typically by raster scanning the beam. The depth can be controlled by selecting an appropriate combination of laser fluence (F), pulse overlap (O), and hatch distance . As can be seen in The laser microwelding system enables the generation of weld seams at the interface of two glass plates, as demonstrated in References ,42. Prev®33 glass plates. The glass plates are blank, i.e., they do not contain any laser-generated patterns. The glowing lines seen in The generation of weld seams occurs via multi-photon absorption and subsequent plasma generation inside the glass plates, as described in Reference . By focu2) since a single depth was required. In order to drill two inlet/outlet ports (through holes), a total of nine passes were used. To maintain the laser beam focus during the drilling process, the sample was moved by a 0.1-mm distance toward the galvo-scanner after each laser scan.The total time required for the generation of the microfluidic pattern and for the drilling of the inlet/outlet ports was approximately 15 min. Laser machining was carried out using three passes of the laser beam with PRF = 100 kHz and v = 150 mm/s. In this case, the laser fluence used had a constant value . These pressures did not cause physical damage to the devices. During the gas injection, nitrogen created preferential paths  to escape the device through the outlet port, as can be seen in Not all water could be removed from the microfluidic devices following the nitrogen injection. In some places, water was trapped in the microstructure, as shown in During the water injection at a pressure of ~1.5 bar, when the microfluidic device still contained air, it was observed that the pressure inside the device caused a little deformation of the glass plates, giving rise to a couple of optical fringes as a result of gap formation within the laser-machined area. Although the gap was very small (approximately 1 μm) and did not have any visible impact on the behavior of the flow of fluids, the authors consider this may be a problem in particular at high injection pressures, where injected fluids may start bypassing the microchannels by flowing through the gaps. One of the solutions to overcome this problem would be to use thicker  glass substrates for manufacturing the microfluidic devices. Another solution would be to place the microfluidic device inside a hermetic vessel and apply an external pressure onto the microfluidic device in order to compensate for the internal pressure. This solution requires a special vessel with a transparent window to provide optical access to the microfluidic device; however, it is feasible, as already shown in several publications ,17,18,19This paper describes a laser-based process suitable for the rapid manufacturing of glass microfluidic devices. Using this process, it is possible to generate almost arbitrary enclosed microfluidic structures. The lateral resolution of the patterns, however, is limited to the laser spot size used for micromachining. The fluid flow tests performed for the microfluidic devices proved that a good sealing of the laser-generated microstructures can be obtained using the picosecond laser microwelding process. Weld seams generated by the laser not only eliminate any existing gaps between two glass plates, bringing the materials into close contact, but they also confine the flow of fluids to the designated areas. Hence, the laser microwelding process seems to be an attractive alternative to processes such as anodic bonding and thermal bonding.Continuing work will focus on the investigation of the flow of different fluids through a range of pore network patterns generated by the laser. Currently, we are building a workstation dedicated to such experiments. The workstation will be equipped with a high-resolution camera with a high-zoom objective lens in order to observe the flow of fluids in microchannels, while fluids will be injected to the microfluidic devices at controlled flow rates using syringe pumps. In this way, it will be possible to investigate various fluid transport processes and to determine conditions at which injected fluids follow the microfluidic patterns, as well as identifying the limit beyond which fluids start flowing through small gaps between the glass plates, bypassing the microchannels.Finally, it should be highlighted that the laser microwelding process has some limitations. One of the limitations is the lateral dimension of weld seams that determines the minimum size of the areas suitable for welding. In our case, two parallel microchannels cannot be isolated from each other by producing weld seams between them if the clearance between the microchannels is less than 100 μm. Weld seams of smaller dimensions, however, should be possible to generate using a different combination of laser welding parameters, i.e., using a different scan speed, pulse energy, repetition rate, and pulse duration. Unfortunately, the last two parameters cannot be changed in our laser system. In addition, equipping the laser microwelding system with a visualization system would simplify the necessary precise positioning of the welding samples. Using such a system, weld seams could be readily generated in specific locations, even in small areas between individual microchannels. Another limitation of our laser microwelding system is a welding speed (currently 2 mm/s). Higher welding speeds, however, can be achieved using ultrafast lasers operating at higher PRFs. For instance, using a laser with a PRF of 2 MHz, it should be possible to increase the welding speed to 10 mm/s, reducing the total welding time of microfluidic devices by a factor of five.