To assess the impact of specialist contraceptive support after abortion on effective contraceptive use at six months and subsequent abortions within two years.Multicentre randomised controlled trial among women undergoing induced abortion in three London boroughs. Allocation was through electronically concealed stratified randomisation by centre, blinding clinicians and participants to arm allocation until interventions. Control group received standard care, comprising advice to follow up with their general practitioner or contraceptive clinic as needed. Intervention group additionally received specialist contraceptive support via telephone or face-to-face consultation at 2–4 weeks and 3 months post-abortion. Primary outcome was rate of effective contraceptive use at six months post-abortion. Secondary outcomes were subsequent abortions within two years.569 women were recruited between October 2011 and February 2013, randomised to intervention (282) and control (287) groups; 290  were available for primary outcome analysis. Intention-to-treat analysis showed no significant difference between the two groups in effective contraceptive use after abortion , long-acting contraceptive use , and subsequent abortion . Per-protocol analysis showed those who received the complete intervention package were significantly more likely to use effective contraception , in particular long-acting contraception  and showed a non-significant reduction in subsequent abortions within 2 years .Structured specialist support post-abortion did not result in significant use of effective contraception at six months or reduction in subsequent abortions within two years. Participants engaging with the intervention showed positive effect on effective contraception at six months post-abortion. The potential benefit of such intervention may become evident through further studies with increased patient participation. Abortion rates in England and Wales have declined in the last decade, however subsequent abortions are on the rise; in 2016, 38% of women undergoing abortions had one or more previous abortions as compared to 32% in 2006 ; p = 0·172 . AdditioData from 551 patients (270 (Intervention), 281 (Control)) were analysed for the intention-to-treat secondary outcome analysis. Subsequent abortions within one and two years post-index abortion were similar in the intervention and control groups; subsequent abortion within one year was10%, and between one and two years was 6%. Details of these results are illustrated in Further exploratory analyses on the intention-to-treat population showed that the use of LARC method at six months was higher in the intervention group, but not statistically significant . A non-significant effect was observed in patients’ satisfaction with their contraceptive method used at six months in the intervention group compared to the control group . Details of these results are illustrated in In the per-protocol population, 121 women received the complete intervention package but only 96 of these were available for the six-month follow-up. From the control arm, 148 women were available for the six-month follow-up. The baseline characteristics of the 244 patients included in the per-protocol analyses were also balanced .Women who received the complete intervention package were significantly more likely to use effective contraception  and use LARC  at six months post-abortion. However, satisfaction with chosen method of contraception remained non-significant between groups. There was a reduction in subsequent abortions within one and two years post-index abortion in the group that received the complete intervention package compared to control group, although this did not reach statistical significance . Details of these results are illustrated in In the per-protocol population, a higher proportion of women had changed from using no contraception or a non-LARC method prior to abortion to a LARC method at six months (50%) compared to those in the control group (31%) . The change from non-LARC or no method prior to abortion to LARC at six months post-abortion was two-fold higher in those who completed the intervention compared to the control group (odds ratio = 2·21 [95% CI 1·28 to 3.83]). These rates were also significant in the intention-to -treat population .This study showed that a structured specialist support package following abortion did not increase the use of effective contraception at six months post-abortion. There was no significant difference in the incidence of subsequent abortions within two years between the intervention and control groups. However, participants who engaged in the interventions showed a positive effect on effective contraceptive use, in particular long-acting contraceptive methods. A reduction was observed in subsequent abortions within one and two years in the per-protocol population, but did not reach statistical significance.A recent systematic review  includinTo our knowledge, our study is the first RCT in the United Kingdom assessing a specialist contraceptive intervention provided exclusively in the post-abortion period. One of the main challenges was recruitment, typical of studies recruiting from clinics dealing with sensitive matters such as abortion . The parOur randomisation process ensured that the study arms were balanced in their baseline characteristics. It was stratified by centre to eliminate bias arising from differences between abortion centres regarding client characteristics, pre-abortion contraceptive counselling and immediate post-abortion contraceptive provision.Despite the majority of our study participants reporting no plans for a pregnancy in the next 12–24 months , just ovInitiating contraception on the day of abortion, in particular LARC, has proven significant in the reduction of subsequent abortions ,26. A stIt has also been reported that women who have had a recent abortion have lower rates of contraceptive continuation and a higher risk of subsequent unintended pregnancy compared to women who never had an abortion . Recent Monitoring unintended pregnancies for extended periods of time is one of the most reliable indices of effectiveness of contraceptive counselling . PregnanA larger sample size with specialist intervention targeted for a select population, for example, those who remain undecided about their contraceptive method post-abortion, and women who undergo medical abortion, may have changed the outcome of the study. However, there are real practical challenges to recruitment in the abortion setting which could make studies with large sample sizes unachievable and very expensive. To eliminate the potential dilution of any intervention effect arising from lost-to-follow-ups, future studies could use subsequent abortions through data-linkages with routinely collected national abortion datasets as the primary outcome. However such studies need much bigger sample sizes. Furthermore, national population-level reporting of the incidence of abortions within one-to-two years after an abortion could help evaluate the impact of interventions to improve post-abortion contraception.The potential benefits of timely interventions tailored to support effective contraceptive use by women undergoing abortion may be realised through strategies which optimise patient engagement. Further research is required to identify contextual barriers to follow-up interventions to improve post-abortion contraception. Collaborative initiatives by abortion services and contraceptive specialists can help to engage patients in need of additional contraceptive support post-abortion, ensuring a seamless pathway of support.S1 Consort Checklist(DOC)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S1 Topic GuideTopic guide for data collection sections A, B, C, D and E.(DOCX)Click here for additional data file.S1 Trial Protocol(DOC)Click here for additional data file.


Previously, using fMRI, we demonstrated lower connectivity between right anterior superior temporal (ATL) and anterior subgenual cingulate (SCC) regions while patients with major depressive disorder (MDD) experience guilt. This neural signature was detected despite symptomatic remission which suggested a putative role in vulnerability. This randomised controlled double-blind parallel group clinical trial investigated whether patients with MDD are able to voluntarily modulate this neural signature. To this end, we developed a fMRI neurofeedback software (FRIEND), which measures ATL-SCC coupling and displays its levels in real time. Twenty-eight patients with remitted MDD were randomised to two groups, each receiving one session of fMRI neurofeedback whilst retrieving guilt and indignation/anger-related autobiographical memories. They were instructed to feel the emotion whilst trying to increase the level of a thermometer-like display on a screen. Active intervention group: The thermometer levels increased with increasing levels of ATL-SCC correlations in the guilt condition. Control intervention group: The thermometer levels decreased when correlation levels deviated from the previous baseline level in the guilt condition, thus reinforcing stable correlations. Both groups also received feedback during the indignation condition reinforcing stable correlations. We confirmed our predictions that patients in the active intervention group were indeed able to increase levels of ATL-SCC correlations for guilt vs. indignation and their self-esteem after training compared to before training and that this differed significantly from the control intervention group. These data provide proof-of-concept for a novel treatment target for MDD patients and are in keeping with the hypothesis that ATL-SCC connectivity plays a key role in self-worth.https://clinicaltrials.gov/ct2/show/results/NCT01920490 •Employs real-time fMRI of anterior temporal –subgenual cingulate connectivity•Previously decreased for guilt in major depressive disorder (MDD) beyond remission•This RCT shows MDD patients can increase connectivity in one neurofeedback session.•Active neurofeedback group increase self-esteem vs control neurofeedback group•Training-induced self-esteem increases correlate with connectivity increases This potentially mitigates a shortcoming of cognitive therapy  for majo showed iWe recently provided evidence that reduced positive memory biases, as successfully targeted in previous neurofeedback studies ; 2014, wClinicalTrials.gov: NCT01920490):1)Demonstrate that ATL-SCC coupling for guilt can be increased through one session of neurofeedback in the group seeing visual feedback based on increasing correlations during the guilt condition compared with the group seeing visual feedback based on keeping correlations at the same level during the guilt condition.2)Demonstrate that this increase in coupling is selective for guilt relative to indignation.3)Demonstrate that mood is not negatively affected by neurofeedback.4)Explore whether this short intervention decreases self-hate on the Interpersonal Guilt Questionnaire  and incrIn this double-blind RCT we sought to establish proof-of-concept for a neurofeedback approach aimed at self-blame-selective reductions in functional connectivity  between 2https://clinicaltrials.gov/ct2/show/results/NCT01920490).This randomised, controlled, double-blind, parallel group, clinical trial was pre-registered  for at least 2 months, currently not fulfilling criteria for an episode and remitted from symptoms for at least 2 months, and age ≥18.Exclusion criteria: current suicidal thoughts, other current DSM-IV axis-I disorders, a history of atypical major depressive episodes (DSM-IV), Global Assessment of Functioning scores below 80 as a sign of incomplete remission or co-morbidity, >2 points on the suicidality item of the Hamilton Depression Scale  without researchers being aware of the allocation. After seeking statistical advice, we amended our protocol to use minimised randomisation  affiliated with IDOR, and screening assessments were carried out by an independent clinical researcher as part of the IDOR neuropsychiatric and normal control studies recruitment procedures. During the screening visit, a closely supervised specialist trainee and/or neuropsychologist carried out an in-depth assessment using the MINI plus International Neuropsychiatric Interview  changing2.3[Time Frame for all measures: change from baseline after one session of fMRI neurofeedback training].Primary Outcome Measure:•Increase in correlation between ATL and SCC fMRI signal for guilt relative to indignation Correlations are computed by using average signal in the most highly activated voxels within a priori regions of interest (ROIs) in the right superior ATL and SCC region. The same a priori regions are also used to provide neurofeedback.Secondary Outcome Measures:•Beck Depression Inventory . This w•Interpersonal Guilt Questionnaire - Self-hate subscale ; 2002.•Rosenberg Self-Esteem Scale The following two exploratory outcome measures were used to determine whether there is a detectable effect on self-blaming emotions after one session of fMRI neurofeedback. This was not our primary aim in that this study was primarily designed to determine feasibility and safety rather than efficacy:2.4The first and fourth run  were identical and were used to determine pre- and post-neurofeedback effects. The first run was also necessary to define the 10% most activated voxels for guilt vs. subtraction or indignation vs. subtraction in the predefined SCC and ATL ROIs. These individualized ROIs were used to extract average signal for subsequent neurofeedback training. First and fourth runs consisted of four blocks of guilt and four blocks of indignation memories (15 volumes per block), interspersed with eight mental subtraction condition blocks (10 volumes each). The second and third run were identical, and participants were provided with visual feedback for training during these runs. The neurofeedback runs consisted of 360 volumes with 120 volumes for guilt and 120 for indignation distributed over 4 blocks (30 volumes per block) which were interspersed with 8 blocks (15 volumes) of mental subtraction (120 volumes) by serial subtraction of seven from a 3-digit number . Mental subtraction was used to allow patients to distract themselves emotionally from the scenarios and to decrease, so-called “resting-state” activity known to be present in the SCC .2.5Autobiographical events were selected by participants prior to the scanning session. They chose events that evoked guilt and entailed their own actions. They also had to choose events that evoked indignation and entailed other people acting. Participants had to define cue words pointing to these events to be displayed to them during the scanning session (2 scenarios/events for each condition). Neurofeedback instructions for both groups in both conditions were identical in that they were asked to bring up the level to which the bar is filled with colour on the thermometer-like visual feedback displayed to them whilst thinking about the event (see Supplementary Methods).•ACTIVE: GUILT-INCREASE-CORRELATION: Visual feedback reinforced increasing the correlation in fMRI signal between the right superior ATL and SCC regions during retrieval of guilt-related events.•CONTROL: GUILT-STABILISE-CORRELATION: Visual feedback reinforced stabilisation of the preceding degree of correlation in fMRI signal between the right superior ATL and SCC regions during the retrieval of guilt-related events. The level of the bar on the visual feedback thermometer-scale went up for correlations staying within the preceding range and down for correlations outside of the range.During the indignation condition, visual feedback reinforced stabilisation of the preceding degree of correlation between the ATL and SCC in both intervention groups. The two intervention groups were:The rationale for stabilisation as a control intervention was to provide feedback from the same brain regions as in the active group whilst being engaged in the same psychological task which avoids differences in the psychological aspects of the intervention in both groups. This design further avoids providing feedback from a brain region which is not relevant to the psychological aspects of the task and could thus create a mismatch between neurofeedback signal and psychological task.2.6http://fsl.fmrib.ox.ac.uk/fsl/fslwiki/OtherSoftware and http://idor.org/neuroinformatics/friend  was uses/friend . Signal-s/friend , see Sups/friend a&b was dIncreasing correlations were reinforced by increasing the level of the colour bar if correlation coefficients were higher than the weighted mean  plus one standard deviation of the correlation values so far. Stabilisation of correlations was reinforced by increasing the colour bar level if correlation coefficients kept close to the weighted mean  and lower colour bar levels if the correlation coefficient deviated positively or negatively from the previous average.Root mean squares (RMS) of movement parameters for translation and rotation were tracked on an ongoing basis and the neurofeedback screen displayed a warning to participants and investigators in real-time if movement exceeded allowable levels, excluding those volumes from calculations of feedback signal (see Supplementary Methods). Unfortunately, FRIEND does not record the number of these excluded volumes during real-time neurofeedback.2.7Images were acquired with a 3 T Achieva scanner  using a T2*-weighted echoplanar (BOLD contrast) sequence . Before each run, five dummy volumes were collected for T1 equilibration purposes. A SENSE factor of 1.5 and dynamic stabilisation were additionally used. These parameters were based on careful sequence optimization to maximize temporal signal-to-noise[20] in brain regions that normally suffer from magnetic susceptibility effects, including the SCC and ATL. High-resolution anatomical images were acquired with a 3D turbo field echo T1-weighted sequence . Head motion was restricted by using foam padding and straps over the forehead and under the chin.2.8http://www.ibm.com/analytics/us/en/technology/spss/) for each subject by modelling the interaction of z-transformed ATL signal time-course with two factors: condition  and time . The z-transformation was undertaken to obtain standardised regression coefficients. Cohen's d effect sizes were computed for each regression coefficient using the formula: 2 × t-value/square root of degrees of freedom  in the pre- and post-training acquisition as the outcome variables were derived from a general linear model in IBM SPSS 23 .t-test to compare the two intervention groups which as expected resulted in the same p-value as the interaction term produced by the repeated measures ANOVA. The alpha-level was set to p = .05, two-tailed test. Clinical trial statisticians usually recommend using an ANCOVA of the post-intervention outcome measures with baseline as a covariate for complete data. The reason why we have chosen a different approach here for our imaging data is, because we have two related imaging outcome measures (guilt and indignation) which we sought to contrast against each other, but also look at separately. To realise this in an ANCOVA, we would have had to use a Multivariate ANCOVA with 7 predictors to model all the interactions between baseline connectivity and intervention group. With an n of 28 this would have entailed a massive overfitting problem, rendering the model probably not reproducible  and one between-subject factor . To further examine and estimate the confidence intervals of the observed three-way interaction between condition, time, and group, we computed the difference between the regression coefficient effects for guilt vs. indignation for post- vs. pre-training for each individual. The resulting measure was examined using an independent oducible .33.1As predicted, we found an increase in ATL-SCC correlation for guilt vs. indignation after training compared to before training in the ACTIVE group as compared with the CONTROL group . ContrarFurther secondary analyses considering pre-training differences between groups in ATL-SCC connectivity (see Supplementary Results), confirmed our primary analysis that the ACTIVE group showed higher post-training ATL-SCC regression effects for guilt vs. indignation compared with the CONTROL group. Furthermore, this secondary analysis showed that this result was associated with the interaction of intervention group and pre-training ATL-SCC regression effects. This interaction effect was driven by a negative association of pre-training with post-training ATL-SCC regression effects for guilt vs. indignation in the ACTIVE group and a positive association in the CONTROL group. This indicated that patients in the ACTIVE group with low connectivity between the ATL and SCC for guilt vs. indignation prior to neurofeedback training were more likely to increase the levels of connectivity through training when compared with those who had high levels of connectivity prior to training already. The positive association of pre-training and post-training ATL-SCC connectivity in the CONTROL group reflected that the control intervention was aimed at stabilising pre-training levels of connectivity. Further, these secondary analyses showed that the weak trendwise neurofeedback-training effect difference between intervention groups on ATL-SCC regression coefficients for indignation in our primary data analysis disappeared when correcting for pre-training differences in ATL-SCC connectivity for indignation.3.2r = 0.47, p = .02, n = 23). Contrary to our expectations, we did not find training effects for self-hate on the Interpersonal Guilt Questionnaire after training compared to before training  that compared to the CONTROL intervention, the ACTIVE neurofeedback intervention would lead to an increase in ATL-SCC connectivity for guilt relative to indignation, whilst we were unable to demonstrate that the intervention increases connectivity for guilt on its own (aim 1). We also confirmed our prediction that there would be no safety concerns by showing stable Beck Depression Inventory scores (aim 3). Intriguingly, we were able to demonstrate an effect of the intervention in increasing global self-esteem (aim 4) despite this measure's focus on general feelings about oneself rather than a specific focus on the current state. Further strengthening the assumption that this training-induced increase in self-esteem was a result of the ATL-SCC connectivity increase for guilt vs. indignation, we found a positive correlation between self-esteem and connectivity changes.Two additional findings render a non-specific boost in positive emotions or perceived success as explanations for this effect unlikely. Firstly, the ACTIVE intervention appears to have been more difficult and thus patients had a much lower thermometer level display on average, thus getting much less positive feedback on their performance than the CONTROL intervention group. The high success rates of stabilisation neurofeedback show that this was an adequate control intervention which was unlikely to have interfered with self-efficacy perceptions shown to be important for neurofeedback outcomes . InteresOur finding of a direct relationship between ATL-SCC connectivity and global self-esteem in MDD provides further evidence for the important role of this neural system for self-worth in MDD . This diA more recent study , whose rOur finding that patients with lower baseline levels of ATL-SCC connectivity for guilt relative to indignation had stronger training effects in the ACTIVE group should be used to design future trials by stratifying patients on the basis of their functional connectivity patterns when assigning them to a blame rebalance neurofeedback intervention. This is likely to increase the treatment effects and thus the statistical power of the trial. It remains unclear why the ACTIVE group had lower baseline connectivity for guilt vs. indignation compared with the CONTROL group, but one clue is offered by one participant in the control group with the highest baseline connectivity levels in the study, who showed potential bipolar features. Future trials should employ DSM5 criteria with its improved sensitivity for bipolar spectrum conditions which we may have under-detected in the current DSM-IV-based study, leading to potentially uncontrolled group differences in the frequency of patients whose condition would have been better captured by a bipolar spectrum diagnosis.4.1Potential limitations of our study were that we cannot determine how representative our sample was of MDD patients seen in general outpatient specialist settings as we did not collect data on how many patients were pre-screened for eligibility by referring psychiatrists. Another limitation was that this study was designed as a proof-of-concept study with a limited sample size, so that our results need to be replicated in a larger sample, ideally with repeated neurofeedback sessions, and a longer follow-up period to demonstrate clinical benefits and further safety data.5Patients with MDD were shown to be able to modulate guilt-selective ATL-SCC functional connectivity after a single neurofeedback session and this improved their self-esteem. Furthermore, increases in self-esteem correlated with connectivity increases. Non-specific effects such as perceived success cannot explain these findings. Whilst preliminary, this is in keeping with the hypothesis of a relevant direct or indirect relationship between ATL-SCC connectivity and self-esteem, although confounding differences between the groups other than the received ATL-SCC connectivity feedback cannot be ruled out , and proRZ: Lundbeck-sponsored presentation on neurofeedback, co-investigator Livanova-funded study; Industry advice via Guidepoint Global; Industry collaborations with EMIS PLC and Alloc Modulo LTD. Private Clinical Practice at The London Depression Institute. AHY: Paid lectures and advisory boards for the following companies with drugs used in affective and related disorders: Astrazenaca, Eli Lilly, Lundbeck, Sunovion, Servier, Livanova, Janssen; Consultant to Johnson & Johnson; Lead Investigator for Embolden Study (AZ), BCI Neuroplasticity study and Aripiprazole Mania Study; Investigator initiated studies from AZ, Eli Lilly, Lundbeck, Wyeth, Janssen. JM: Shareholder of Rede D'Or hospitals (Brazil) and a managing partner of VHM LLC. (US). All these interests are unrelated to the study. The other authors have no conflicts of interest to declare.


A high proportion of notified tuberculosis cases in the Philippines are clinically diagnosed (63%) as opposed to bacteriologically confirmed. Better understanding of this phenomenon is required to improve tuberculosis control.To determine the percentage of smear negative presumptive tuberculosis patients that would be diagnosed by GeneXpert; compare clinical characteristics of patients diagnosed as tuberculosis cases; and review the impact that the current single government physician and a reconstituted Tuberculosis Diagnostic committee (expert panel) may have on tuberculosis over-diagnosis.This a cross-sectional study of 152 patients 15–85 years old with two negative Direct Sputum Smear Microscopy results, with abnormal chest X-ray who underwent GeneXpert testing and review by an expert panel.Thirty-two percent (48/152) of the sample were Xpert positive and 93% (97/104) of GeneXpert negatives were clinically diagnosed by a single physician. Typical symptoms and X-ray findings were higher in bacteriologically confirmed tuberculosis. When compared to the GeneXpert results the Expert panel’s sensitivity for active tuberculosis was high , specificity was low .Using the GeneXpert would increase the level of bacteriologically confirmed tuberculosis substantially among presumptive tuberculosis. An expert panel will greatly reduce over-diagnosis usually seen when a decision is made by a single physician. In 2016Local and international experts have noted the high level of TB cases that are Clinically Diagnosed (CD) (approximately 61% of notified cases in 2017) in the Philippines [GeneXpert MTB/RIF (Xpert), introduced in the Philippines in 2011, is a rapid diagnostic test with high sensitivity and specificity for TB. Despite the availability of Xpert, direct sputum smear microscopy (DSSM) remains the primary diagnostic tool for pulmonary tuberculosis (PTB), although the national program is gradually shifting to Xpert.Cavite, one of the largest provinces in the Philippines, has consistently low detection rates and high CD PTB ,4. BacteThe TB Diagnostic Committee (TBDC), composed of a local panel of experts from private and public sectors, was created in the 1990’s by the Department of Health (DOH) to improve the quality of diagnosis of sputum smear-negative cases using chest X-ray findings of patients from the TB Directly Observed Treatment (DOTS) Facility. The shortage of trained and qualified specialists especially in rural and remote areas in the Philippines however resulted in a longer turnaround time and delays in diagnosis and treatment. This led to a change in the Philippine Manual of Procedures which then gave an option of a single physician (SP) making a decision instead of a TBDC. This may have contributed to the steady rise of clinical diagnosis in recent years. Oftentimes an SP (sometimes acting as a ‘one-man TBDC’) can decide on diagnosis and treatment of clinical TB but some TBDCs continue to function .This study is part of a larger project called the Newton Agham Fund Impact assessment of diagnostic algorithms and tools for multi drug resistant (MDR-TB) and drug sensitive tuberculosis (TB) in the Philippines (TB-FIT). The primary aim of this study is to evaluate the role of GeneXpert in the diagnosis of BC TB among SNPT cases in a high CD prevalent area of Cavite.The study was conducted in two urban-based public DOTS centers in the province of Cavite [Patients seeking consult at Dasmariñas City and Trece Martires CHOs DOTS facilities with two negative sputum smears and with abnormal chest X-ray, were recruited into the study.Direct Sputum Smear Microscopy was performed using Ziehl-Neelsen microscopes. Patients without chest X-rays or without film available, were requested to obtain a chest X-ray at the CHO/De La Salle. Eligible patients were offered the Xpert test and instructed on the test evaluation and sputum collection procedure.Field coordinators secured informed consent and interviewed the eligible patients. A pre-tested tool was used to collect baseline data on demographics, clinical symptoms and chest X-ray. X-ray films were reproduced using a 1080 x 1920 resolution and 18 megapixels camera. X-ray results were classified based on standard radiologic reading of chest films utilized by certified local radiologic society (Philippine College of Radiology) –9. All XXpert testing was performed according to the manufacturer’s procedure. Those who were Xpert negative were referred to the corresponding SP of the participating sites for final disposition. Patients recommended for treatment by the SP, were considered as CD TB. In addition, a panel of experts consisting of 3 certified pulmonologists, a microbiologist and a radiologist reviewed the clinical profile and chest X-rays of all study participants. The panel was unaware of the Xpert results prior to their evaluations. If there was a disagreement with the diagnosis among the members, consensus of the majority (at least 3 of 5 panel members) was needed to carry out the diagnostic recommendation.Physicians were provided with the results of the Xpert prior to patient’s consult. Treatment statuses of the patients were obtained within a month after notification of Xpert result.A sample size of 152 SNPT patients were included in the study to be able to have 95% confidence level of estimating percentage of Xpert positivity at 21.3%  with relThe survey was paper-based and EPI Info for Windows v7.2 was used for data entry and analysis. A Chi-square test was used to determine significant differences in categorical variables such as demographic and clinical characteristics between those Xpert positive and CD patients, while a t-test was used to determine significant differences in continuous variables such as age and number of symptoms. Level of significance was set at p-value less than 0.05.This study was approved by the Independent Ethics Committee of the De La Salle Medical and Health Sciences Institute with the approval number of–DLSHSI-IEC (2017)– 02 -01- A. All participants were provided a written informed consent.From June to August 2017, we recruited 152 patients who fulfilled the study’s inclusion criteria. They were all smear-negative participants with an abnormal chest X-ray, and all consented to participate and be tested with Xpert. The mean age was 40.3 years (range 15–85 years); 62.7% were male.Programmatic Management of Drug-resistant TB Centre (PMDT) for treatment.Thirty-two percent  tested positive on Xpert and were enrolled in treatment at their corresponding DOTS facility. An average of 3 smear-negative presumptive PTB patients needed to be Xpert tested to detect 1 Bacteriologically Confirmed (BC) PTB. One rifampicin resistant case was detected and enrolled at the Ninety-seven of the 104 (93.3%) smear-negative, Xpert negative patients were CD by SP and were enrolled for treatment. Of the remaining seven, four were diagnosed with no PTB, three patients needed further diagnostic tests . Only 89 CD patients were enrolled to treatment . The remTables p-value = 0.004).In p-value <0.001). Prevalence of the following symptoms were higher among the BC patients than CD: cough , loss of appetite , weight loss , shortness of breath . Cough, weight loss, and hemoptysis were 4 times (3.7 to 4.4. OR) more likely in BC than in CD  Excluding those for further testing, the EP diagnosed active PTB in 39 of 40 Xpert positive patients yielding a 98% sensitivity (based on Xpert as the gold standard), and further diagnosed 35 out of 87 Xpert negatives as non-active TB yielding a 40% specificity.Of the 104 Xpert negative cases, the EP considered 52 cases, 28 of which were active PTB cases and 24 indeterminate cases. According to the panel these 24 indeterminate cases should be enrolled for treatment because of the presumption that it is better to treat them than leave them untreated since some may really have active TB. Yousang Ko et al. in 2018 evaluated the diagnostic performance of the radiographic CT scan activities in predicting definite and overall PTB in patients with presumed PTB and found that 10% of indeterminate PTB were culture confirmed .Across two DOTS centers in Cavite, testing SNPT cases with GeneXpert returned a bacteriological confirmation in 31% of cases (i.e. we only need to test 3 smear negative cases to detect 1 BC). This percentage is consistent with the range of figures from other studies (21% - 36.5%).In a fee paying hospital in Nepal 21% of smear negative cases were bacteriologically confirmed by Xpert and the number needed to test was five . A higheKnown clinical predictors of PTB such as cough, weight loss, hemoptysis and classic radiologic findings seen on x-ray, were observed among many Xpert positive cases as expected ,15. TherThree features may have contributed to the high SNPT. Firstly, clinical diagnosis of SNPT relied on chest X-ray findings as demonstrated among fifteen asymptomatic smear and Xpert negative cases who were diagnosed based on chest X-ray findings of apical infiltrates/densities. Secondly, the study inclusion criteria required an abnormal chest X-ray, which increased the likelihood of being CD, reaching 93% of all smear and Xpert negative subjects diagnosed as CD by the SP. Thirdly, there was a high percentage of CD cases among subjects whose consultation was for employment purposes.Based on the NTP Manual of Procedures, the TBDC or a Physician may diagnose patients as CD. This was introduced in 2014 due to slow turnaround times of the TBDC decisions which delayed treatment . While SUsing a published sensitivity of 67% and specificity of 98%  of XpertThe higher proportion of CD patients among those attending pre-employment ‘fit to work’ checks supports the hypothesis that there are non-medical factors leading to physicians enrolling patients onto TB treatment. The authors believe the high CD rate among those subjects is largely due to patient-pressure, with patients seeking to be placed on treatment as a way of being cleared as fit to work. The study sites are industrialized areas in Cavite where several industrial zones are located, and similarly in surrounding municipalities. Chest X-ray is a mandatory pre-employment procedure , and thoOver-diagnosis leads to additional health system and direct patient costs. Patients incur costs because of repeated visits to health centers, and exposure to potentially serious side effects from unnecessary drugs. Patients misdiagnosed to have TB may actually have another serious illness–e.g. cancer that requires immediate attention.In this study an SP made decisions on clinical diagnosis. A fully represented TBDC exemplified by the EP, may be able to diagnose correctly those with active TB (SNPT) missed by Xpert. This study suggests that a fully represented TBDC is still useful to address missed cases not identified by available diagnostic tests including Xpert and should be the preferred option in diagnosing SNPT.Recently, the WHO  releasedThe X-rays obtained in the two sites in both public and private were mostly non-digital, many were of poor quality. The use of Xpert as the gold standard instead of culture may have missed some cases because of its lower sensitivity. There may be truly CD cases that are both smear-negative and Xpert negative. It must also be pointed out however that the EP in this study may not truly represent TBDCs nationwide.The impact of Xpert on the diagnosis of confirmed TB is substantial, identifying potentially 31% of smear negative presumptive cases as BC PTB. Clinical and X-ray features among Xpert positive cases supported the accuracy of Xpert. The EP is likely to have high sensitivity but low specificity for diagnosing presumptive smear-negative tuberculosis when compared to the Xpert results. The number of patients enrolled to treatment was not influenced by Xpert results. There was still a high percentage of smear-negative, Xpert negative cases who were enrolled to treatment upon the decision of an SP. A combination of factors such as relying on chest X-ray for diagnosis of SNPT, inclusion criteria of abnormal chest X-ray, implication of opportunity cost among patients seeking pre-employment clearance contributed significantly to the high percentage of clinically diagnosed SNPT.


A recent study used CRISPR/Cas9 to reveal long-range looping between disease-related genes and their regulatory elements that is mediated by the CCCTC-binding factor (CTCF) in prostate cancer. Extensive studies are currently devoted to understanding the three-dimensional (3D) architecture of genomes, including the formation and function of chromatin loops, topologically associated domains (TADs) and transcriptional activity-based A and B compartments. These studies provide evidence that the spatial organization of the genome is a global regulator of gene transcription. DNA looping is the fundamental architectural unit of the 3D genome and builds long-range connections and communication between genes and their regulatory enhancer elements. In addition, genome-wide association studies (GWAS) have revealed that a large fraction of disease-associated mutations or genomic rearrangements are found in non-coding rather than coding regions of the genome, providing a clue that the long range communication between genes and regulatory elements might play an important role in human disease . A recenEnhancer-promoter looping facilitates the assembly of the transcriptional machinery at specific promoters and thus the effective initiation of transcription. Histone H3K27ac distinguishes active enhancers, which are the ones that frequently engage in looping to transcriptionally active genes, from inactive or poised enhancers. Chromosomal looping is also maintained by numerous architectural proteins and by long non-coding RNAs (lncRNAs) in addition to regulatory DNA elements. The major architectural protein CCCTC-binding factor (CTCF) is an 11 zinc-finger DNA-binding protein that associates with the cohesin complex and orchestrates long range interactions between remote enhancers and their target gene promoters to modulate gene transcription. Whether this regulation is direct or indirect and involving an insulator function is an open question.Genome Biology, Guo et al. [KCNN3 , was already known to be linked to PCa biology. The CRISPR editing results suggest that KCNN3 is normally held in a repressive loop by CTCF, revealing a novel mechanism underlying genetic susceptibility to PCa.In this issue of o et al.  describeGuo and colleagues set out to discover regulatory elements that are associated with 2181 PCa risk-associated SNPs [KCNN3 expression increased almost 100-fold while other nearby genes were unaffected. On chromosome 12, KRT78 was activated more than 100-fold, while the expression of some neighboring genes was modestly elevated. In contrast to these deletions, when the CTCF sites that paired with the PCa risk-associated CTCF anchor regions on the two chromosomes were deleted, there was only a modest or no influence on gene expression within the loop.To ask whether the enhancer- or insulator-related SNPs were involved in the regulation of chromosomal architecture, Guo et al.  employedKCNN3 and KRT78 are in genomic regions that are devoid of the H3K27ac mark. The authors investigated whether an enhancer element within the looped regions had become activated as assessed by H3K27ac acquisition but did not detect such a change. Instead, the result suggests that after the deletion, the genes came under the influence of a pre-existing enhancer outside the CTCF-mediated loop area that was compatible with only some of the genes that were newly available to it. As Guo et al. [KCNN3 or KRT78 transcription.What underlies the gene activation? Both KCNN3 and KRT78 after deletion of the two PCa-risk-associated CTCF anchor regions is especially notable in contrast to results obtained upon rapid removal of CTCF in embryonic stem cells [The 100-fold activation of HoxA cluster that functions to separate adjacent TADs. Abrogation of CTCF occupancy at this position resulted in the Hox genes’ becoming subject to transcriptional activation from outside their original domain [Hox cluster’s repressive genomic region from active chromatin and thus to maintain a low level of gene expression. In addition, a recent study showed that CRISPR-mediated disruption of CTCF-binding sites at an α-globin locus, subTAD, allowed the α-globin enhancers to activate genes on the other side of the lost boundary, which they normally cannot access [CRISPR/Cas9 technology is proving useful in creating precise deletions or mutations of CTCF motifs in order to determine CTCF function in cell-fate determination, gene regulation and genome topology. A nine-base pair (bp) homozygous deletion was generated by CRISPR/Cas9 editing in a core CTCF motif at a boundary in the l domain . Consistl domain , CTCF fut access . Anothert access .PDGFRA, leading to an enhanced gliomagenesis [CRISPR/Cas9 has been more broadly employed to determine how 3D genome architecture is related to disease states. For example, CRISPR/Cas9 deletion of CTCF sites at the boundaries of an insulated neighborhood containing proto-oncogenes was sufficient to induce their activation in non-malignant cells . Importaagenesis . In anotagenesis .Defining genome architectural mechanisms in disease-related gene regulation has great potential to illuminate the role of GWAS-identified noncoding variation in cis-regulatory elements and regulators of DNA topology. On the basis of the systematic approach described by Guo et al. , CRISPR/


Hereby, the supplemental data of the research article “Long-Term Prognostic Value of High-Sensitivity Troponin T added to N-Terminal Pro Brain Natriuretic Peptide Plasma Levels before Valve Replacement for Severe Aortic Stenosis” are presented [1]. It offers enhanced input on the predictive value of these biomarkers considering the influence of the presence of concomitant coronary artery disease (CAD) in various severities as well as an additional cox proportional hazard model on cardiovascular mortality. Furthermore, the receiver operating characteristic (ROC) curves are shown as figures. The material described increases therefore the understanding of the predictive value of these already routinely available biomarkers and reduces the risk of potential bias due to possible confounding factors. It also underlines the urge for a multi-factorial approach in diagnostics to detect the optimal point for referral to valve replacement other than just symptomatic status, an observed reduction in left ventricular ejection fraction or the presence of CAD with the necessity for coronary artery bypass grafting (CABG) [2]. The data of the 3595 patients were gathered retrospectively at a consortium of four university hospital centers in Austria and combined with prospectively collected data on cardiovascular and all-cause mortality. Out of those 622 (17.3%) had a one-vessel, 337 (9.4%) a two-vessel, 310 (8.6%) a three-vessel and 133 (3.7%) left-main disease. Coronary revascularisation either via concomitant coronary artery bypass grafting surgery  or via percutaneous coronary intervention  was performed in 853 (60.8%) patients.Occurrence of significant CAD separated by high-sensitivity Troponin T (hsTnT) groups is displayed in Three-vessel disease was found in 284 patients of whom 61 had normal hsTnT and 223 had elevated hsTnT. Kaplan-Meier curves for the prediction of cardiovascular and all-cause mortality by using the hsTnT groups in patients with three-vessel disease are presented in 2Data was acquired locally at each of the contributing hospitals in a retrospective way by using the electronic hospital information system or the electronic patient record. Patients were enrolled consecutively at all participating centers, inclusion and exclusion criteria were described in the main publication . Then, iSeveritiy of aortic valve stenosis was graded according to current guidelines . The corChi-square tests were used for the assessment of differences in the prevalence of CAD and hsTnT groups. Analysis of survival was performed by using either the Kaplan-Meier method (univariate) or a cox regression hazard model (multivariate). Calculations were conducted with IBM SPSS version 24 . Graphics were designed by using GraphPad PRISM, version 5 .


Metazoan animals are characterized by restricted phenotypic heterogeneity  of organisms within various species, a feature that contrasts sharply with intra-species morphological diversity observed in the plant kingdom. Robust emergence of morphogenic blueprint in metazoan animals reflects restricted autonomy of individual cells in adoption of fate outcomes such as differentiation. Fates of individual cells are linked to and influenced by fates of neighboring cells at the population level. Such coupling is a common property of all self-organising systems and propels emergence of order from simple interactions between individual cells without supervision by external directing forces. As a consequence of coupling, expected functional relationship between the constituent cells of an organ system is robustly established concurrent with multiple rounds of cell division during morphogenesis. Notably, the molecular regulation of multicellular coupling during morphogenic self-organisation remains largely unexplored. Here, we review the existing literature on multicellular self-organisation with particular emphasis on recent discovery that β-catenin is the key coupling factor that programs emergence of multi-cellular self-organisation by regulating synchronised cycling of individual cells. Developmental morphogenesis describes cellular and molecular events that instruct the final shape and size of metazoan tissues/organs together with functional specialization of the constituent cells . During Self-organisation describes emergence of order in a system without supervision by external directing forces . Self-orβ-Catenin is a metazoan cytoskeletal protein that can also function as a transcriptional factor upon translocation to the nucleus . In ordetrans-activates cyclin-D1 [A key signature of human neural progenitor cells  culturedyclin-D1  and C-myyclin-D1 , two majThese findings suggest that metazoan cell cycle, unlike that of unicellular organisms, has lost its autonomy and is coupled to other cycling cells. Recruitment of β-catenin to junctional complexes  signals It is noteworthy that the synchronicity of a cycling population in relation to the coupling strength (i.e. stability of adherens junctions) can be numerically predicted using the Kuramoto model. The Kuramoto model, named after the mathematician who proposed it, accurately describes the behavior of a large set of oscillators that are connected by a weak coupler . In thisProliferation/differentiation dichotomy, sub-lineage differentiation outcome and spatial navigation are three major events that shape the self-organisation signature of a cellular population. Emerging evidence suggests that proliferation/differentiation dichotomy is, in part, resolved by feedback from cell cycle . In factDecentralised regulation of morphogenesis by self-organisation improves robustness of the developmental landscape as explained above. As such, self-organisation could also improve the heritability of various traits by canalisation  of develIt is proposed that coupled cycling foreshadows a fundamental adaptive change that facilitated evolution and diversification of multicellular life forms. In coupled cycling the stringency of coupling is programmed by availability and subcellular localization of β-catenin that in turn invokes specific self-organisation signatures, anatomical patterns and functional portfolios that characterize tissues and organs. It is suggested that major morphogens the function via β-catenin, such as Wnt, may alternatively be reinterpreted as modulators of coupling strength.


Ovine pulmonary adenocarcinoma is a chronic respiratory disease of sheep caused by jaagsiekte sheep retrovirus (JSRV). OPA is a significant economic problem for sheep farmers in many countries and is a valuable animal model for some forms of human lung cancer. Here, we examined the changes in host gene expression that occur in the lung in response to JSRV infection. We identified a large number of genes with altered expression in infected lung, including factors with roles in cancer and immune system function. We also compared the data from OPA to previously published data from human lung adenocarcinoma and found a large degree of overlap in the genes that were dysregulated. The results of this study provide exciting new avenues for future studies of OPA and may have comparative relevance for understanding human lung cancer. Jaagsiekte sheep retrovirus (JSRV) is the etiologic agent of ovine pulmonary adenocarcinoma (OPA), a neoplastic lung disease of sheep. OPA is an important economic and welfare issue for sheep farmers and a valuable naturally occurring animal model for human lung adenocarcinoma. Here, we used RNA sequencing to study the transcriptional response of ovine lung tissue to infection by JSRV. We identified 1,971 ovine genes differentially expressed in JSRV-infected lung compared to noninfected lung, including many genes with roles in carcinogenesis and immunomodulation. The differential expression of selected genes was confirmed using immunohistochemistry and reverse transcription-quantitative PCR. A key finding was the activation of anterior gradient 2, yes-associated protein 1, and amphiregulin in OPA tumor cells, indicating a role for this oncogenic pathway in OPA. In addition, there was differential expression of genes related to innate immunity, including genes encoding cytokines, chemokines, and complement system proteins. In contrast, there was little evidence for the upregulation of genes involved in T-cell immunity. Many genes related to macrophage function were also differentially expressed, reflecting the increased abundance of these cells in OPA-affected lung tissue. Comparison of the genes differentially regulated in OPA with the transcriptional changes occurring in human lung cancer revealed important similarities and differences between OPA and human lung adenocarcinoma. This study provides valuable new information on the pathogenesis of OPA and strengthens the use of this naturally occurring animal model for human lung adenocarcinoma.IMPORTANCE Ovine pulmonary adenocarcinoma is a chronic respiratory disease of sheep caused by jaagsiekte sheep retrovirus (JSRV). OPA is a significant economic problem for sheep farmers in many countries and is a valuable animal model for some forms of human lung cancer. Here, we examined the changes in host gene expression that occur in the lung in response to JSRV infection. We identified a large number of genes with altered expression in infected lung, including factors with roles in cancer and immune system function. We also compared the data from OPA to previously published data from human lung adenocarcinoma and found a large degree of overlap in the genes that were dysregulated. The results of this study provide exciting new avenues for future studies of OPA and may have comparative relevance for understanding human lung cancer. Ovine pulmonary adenocarcinoma (OPA) is an infectious lung disease of sheep that is a significant economic problem and welfare concern for sheep producers in many countries , 2. The OPA is caused by jaagsiekte sheep retrovirus (JSRV), an exogenous betaretrovirus . Despitein vitro  –10. An u–in vitro , 12 and . We also compared the differential gene expression of OPA-affected animals with previously published data on the two most common types of non-small-cell lung carcinoma (NSCLC) in humans: lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC). Collectively, this study provides new information that greatly enhances our understanding of the host response to JSRV and provides a number of exciting new avenues for future work on OPA.n = 4) and mock-infected (n = 4) specific-pathogen-free (SPF) lambs. The samples used were obtained from a previous study (Whole-transcriptome profiling (RNA-Seq) was performed on lung tissue samples derived from JSRV-infected  showed an intermediate overall expression pattern  and C. Ohttps://david.ncifcrf.gov/  postinoculation), the lambs were culled. Each time a JSRV-infected lamb was euthanized, a healthy animal from the mock-infected control group was culled to provide age- and sex-matched control tissues. Tissues were collected from 24 locations in each lung and stored in liquid nitrogen for RNA extraction and in 10% buffered formalin for IHC. To study tissues from animals with natural disease, lung tissue samples were taken from four farm-raised adult sheep in the advanced stages of clinical OPA and four clinically healthy adult sheep. Cases which had no gross appearance of bacterial or parasitic infection were selected. All protocols involving animal handling and the use of postmortem material were approved by the Animal Welfare and Ethical Review Body of the Moredun Research Institute in accordance with the U.K. Animals (Scientific Procedures) Act 1986.Tissues were available from a previous study in which four SPF lambs were inoculated through intratracheal injection of JSRV at 6 days of age . Four adFor RNA-Seq, RNA was extracted from frozen tissue samples from seven distinct sites of the lungs of each animal and pooled in equal amounts. Total RNA from cryosectioned tissue was extracted using an RNeasy minikit (Qiagen) according to the manufacturer’s instructions. RNA concentration and purity were measured (ND-1000 NanoDrop spectrometer), and RNA integrity was confirmed . All samples had an RNA integrity number (RIN) greater than 8.0. Prior to sequencing, frozen lung tissue sections derived from the same lung sites were examined by histology, which confirmed the presence of OPA tumor lesions in the JSRV-infected animals and the absence of lesions in the mock-infected negative controls. Total RNA was processed (TruSeq RNA Library kit) to generate cDNA libraries according to the manufacturer’s instructions and subsequently sequenced with an Illumina HiSeq2000 instrument using 100-bp paired-end sequencing .AF105220.1) genomes using HISAT2-2.0.4 software  using the Cutadapt (v1.10) program . Nonredusoftware . The quasoftware . Transcrsoftware , and thesoftware  and fittsoftware . Differesoftware . Princip2 fold change value of each gene.Identification of enriched KEGG pathways in the upregulated and downregulated gene lists was performed with DAVID (v6.8) . Pathwayportal.gdc.cancer.gov). Data for 57 LUAD patients and 49 LUSC patients for which corresponding normal and tumor data were available were selected. Differentially expressed genes in human tumors were identified using the edgeR pipeline, as described above for the sheep data. The list of differentially expressed genes in human data was filtered for only those that have an orthologous gene in sheep (identified using the Ensembl BioMart data mining tool) and were differentially expressed in sheep. The comparison of the expression of these short-listed genes was made using the correlation between the expression profiles  in humans and in sheep. The gene lists obtained are presented in Data Set S5 in the supplemental material.To compare significantly expressed genes in JSRV-infected lung with data from human NSCLC, HTSeq counts from RNA-Seq data for LUAD and LUSC were obtained from the GDC portal (R2) of each primer set. In addition, the level of gene expression in experimental samples was ensured to lie within the limits of the standard curve. For comparison of RNA transcription levels between samples, results from the RT-qPCR experiments were normalized to those for two reference genes, the genes for succinate dehydrogenase (SDHA) and β-actin (ACTB) (predetermined to be stable reference genes using geNORM software [RT-qPCR was performed according to MIQE recommendations . Primerssoftware ). StatisTissue sections were routinely processed through graded alcohols and embedded in paraffin wax, and IHC was performed on sections (4 μm) mounted on charged glass microscope slides as previously described . The dilPRJEB27638.The raw RNA-Seq reads (fastq data) of each sample are present in the European Nucleotide Archive with the accession number


Moumne et al.), (2) mechanism of mitochondrial centric cardioprotection involving a large conductance calcium activated potassium channel (BKCa) , (3) involvement of the mitochondria and endoplasmic reticulum crosstalk in the development of heart failure , (4) a non-canonical role of telomerase in cardioprotection , and (5) the association of microRNA-1 with cardiac fibrosis . In addition to these excellent studies, two review articles on oxidant and antioxidant enzymes important in cardiovascular health and disease  and technologies used to develop personalized models of heart failure for drug discovery are included .Cardiovascular disease is associated with a myriad of molecular and genetic aberrations, but the details and impact of these changes on cellular and organ function are poorly understood . By utilAwad et al., the importance of oxidant and antioxidant enzymes on cardiovascular health is discussed. In particular, they summarize our current understanding of major oxidant and antioxidant pathways and their roles in both heath and disease. The authors detail currently available animal models with knockout of genes encoding oxidant and antioxidant enzymes and include known effects and phenotypes. Lastly, they cover important limitations of using these animal models for translational research. Altogether, the review gives an excellent summary of what we know today about the oxidant and antioxidant enzymes associated with cardiovascular disease.In the review by Greenberg et al. emphasizes the dire need for new and effective therapies for heart failure. They discuss the current challenges in heart failure research encompassing the heterogeneous nature of the disease, difficulty in obtaining tissue from heart failure patients, and the molecular changes that occur across time and space during disease progression. A promising solution to these problems is the culturing of human pluripotent stem cells. This method retains the genetic background and allows for the development of patient specific therapies. However, cells in culture are quite distinct and behave differently from those imbedded in native tissue matrices. To circumvent this problem, the authors go into exquisite detail describing the state-of-the-art advances using 3D human engineered heart tissue as screening and rapid development platforms for drug discovery. The development of these platforms could greatly advance our understanding of the causes and development of heart failure and provide impetus to carry research forward from bench to bedside.A review contribution by Moumne et al. provide compelling evidence that gestational hypoxia is strongly linked to cardiac malformations. By breeding wild-type females with heterozygous Nkx2-5 mutant knock-in mice (Nkx2-5+/R52G)  expressed in mitochondria. They employed an ex vivo Langendorff model of ischemia/reperfusion (I/R) injury using wild-type mice and mice overexpressing a gain-of-function subunit of BKCa (BKR207Q) (BKR207Q) . GeneticChen et al. revealed a novel cardioprotective pathway. They generated cardiac specific p53 KO mice by crossing floxed p53 mice with mice carrying α-myosin heavy chain Cre. After a 48-h thapsigargin treatment to induce ER stress, cardiomyocyte death in p53 KO mice decreased relative to wild-type mice. In addition, cardiac mitochondria from p53 KO mice retained complex I activity and pyruvate dehydrogenase content. Furthermore, the release of cytochrome c and apoptosis inducing factor was also inhibited in cardiac mitochondria from p53 KO mice after thapsigargin treatment. These findings provide an additional target to the arsenal of treatments for cardiac injury and heart failure.Using a cardiac specific knockout of the tumor suppressor gene p53, Ait-Aissa et al. on the importance of a telomere-independent function of telomerase in cardiac I/R injury. Using CRISPR/Cas9 technology, they generated transgenic rats with the catalytic-subunit of telomerase knockout (TERT−/−). In an ex vivo Langendorff experimental I/R model, they showed that TERT−/− paradoxically reduced myocardial infarction, but their mitochondrial function was impaired and they showed increased mitochondrial lesion when compared to wild-type. In addition, TERT−/− rats were more susceptible to I/R injury compared to the wild-type when both rat groups were treated with angiotensin II. In the presence of angiotensin-II infusion, infarct sizes increased after I/R and were even larger in the TERT−/− rats compared to the wild-type. This study opens up a new area of investigation regarding the mechanism behind the protective effect of TERT on cardiac tissue exposed to stressors such as I/R injury and in chronic heart disease.A fresh insight into cardioprotective mechanisms was provided by Valkov et al. identified a new role of this transcript in cardiac fibroblasts. Using chronic (4 weeks) subcutaneous injection of angiotensin II, they demonstrated a significant downregulation of miRNA-1 in cardiac fibroblasts and an increase in cardiac fibrosis in Sprague-Dawley rats. They validated this finding by overexpressing miRNA-1 level using adenovirus encoding miRNA-1 or by reducing miRNA-1 level using chemically modified sequence-specific miRNA-1 antagomir in primary fibroblasts in situ to inhibit or facilitate fibroblast proliferation, respectively. They further identified cyclin D2 and cyclin-dependent kinase 6 as the downstream targets of this transcript. From this study, there is evidence of an additional regulatory function of miRNA-1 in the heart that may furnish a new strategy to treat cardiac fibrosis in patients with heart failure.While miRNA-1 has long been established as a muscle specific miRNA, In summary, with the advent of gene editing tools and other molecular approaches, our understanding of cardiovascular physiology has immensely grown to the point where we are able to now design effective therapeutic strategies that hold promise in minimizing the effects of heart disease. Overall, the work in this Research Topic has contributed to our understanding of some of the underlying mechanisms that are the basis of cardiovascular disease by utilizing a range of novel genetic and molecular tools. We are now in an exciting time with new technological developments in drug design and therapeutics for enhanced cardiovascular health .JB, AC, DS, and JO-U contributed conception and design of this topic. All authors contributed to revision, read, and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


With the availability and capabilities of varied technologically enhanced learning activities, the blended learning approach has become increasingly popular in interprofessional education. The combined use of different technologically enhanced learning activities has not been fully examined, particularly to determine the effects of instructional sequences for effective learning outcomes.The objective of this study was to investigate whether the instructional sequences of a blended learning approach can improve students’ learning outcomes on interprofessional competencies.A randomized controlled study was conducted with 40 interprofessional health care teams. These teams undertook three technologically enhanced learning activities—Web-based instruction (WI), virtual reality (VR), and simulation exercise (SE)—after random assignment to three groups based on three different instructional sequences . Pretests and posttests were conducted to evaluate the students’ learning outcomes on interprofessional competencies.P<.05) and attitudes (P<.001) toward interprofessional team care about 1 month after the interprofessional learning activity. Although no significant difference was found (P=.06) between the WI-VR-SE and WI-SE-VR groups in the self-efficacy posttests, participants in the SE-WI-VR group reported significantly lower (P<.05) posttest scores than those in the WI-SE-VR group. The majority of the participants  selected the instructional sequence “WI-VR-SE” as their top preference.A total of 198 participants from the three groups completed the questionnaires. All three groups reported significant improvement in their levels of self-efficacy (This study shows that the instructional sequence of a blended learning approach can have a significant impact on students’ learning outcomes. The learning of concepts from WI followed by problem-solving activity in the SE was found to be a more effective learning sequence than the reverse sequence. We recommend that future studies focus on scaffolding students’ learning when planning instructional sequences for technologically enhanced learning activities within blended learning environments. Technologically enhanced learning in health professional education has evolved rapidly from basic text-based learning to inclusion of more multimedia features , also known as Web-based multimedia instruction, to provide self-directed learning opportunities for learners on didactic material. Yavner et al  describeWith the development of communication tools, there is a growing trend of electronic learning (e-learning) use in interprofessional learning to support interaction and promote information sharing among different health care professionals . E-learnAdvanced interactive technologies including game-based learning, virtual patient, and virtual reality have recently gained attention in health care education. A virtual reality environment generated by computers to create 3D realms allows every user to don the role of a virtual avatar and enables him/her to interact with the avatar in a real-time environment . The useBlended learning refers to combining computer-mediated learning with face-to-face interactions. This can involve a mix of Web-based technologies or various pedagogical approaches to support learning . A systeAlthough the combined use of different learning modalities within blended learning environments has become increasingly popular in the delivery of interprofessional education , it remaThis study aimed to investigate whether the instructional sequences of a blended learning approach can improve students’ learning outcomes on interprofessional competencies. The study also aimed to explore students’ evaluations of the different technologically enhanced learning modalities and their instructional sequences for the delivery of interprofessional education.After obtaining approval from institutional review boards of higher educational institutions, a prospective randomized controlled trial study with a pre-post test design was conducted on students undertaking health care courses  who were in their senior year at three tertiary educational institutions. Participants were recruited via convenience sampling using email and Facebook. The results of a previous study were used to estimate the sample size . For a mThe interprofessional training on multidisciplinary rounds for patient-centered team care was selected as the focus of the learning content. As shown in The participants were asked to complete questionnaires before and 1 month after the blended learning activity to evaluate their interprofessional competencies. A 5-item confidence scale with a 10-point scale, developed by Grundy  for measAn evaluation questionnaire was administered immediately after the blended learning activity to evaluate three learning modalities in relation to the levels of enthusiasm, help, and satisfaction on a 5-point Likert scale. A question on ranking the three instructional sequences in order of the learner’s preference was included in the evaluation questionnaire. All questionnaires were completed electronically to ensure that participants answered all the questions.t test was used to examine any significant changes between the baseline and posttest attitude scores measured by the ATIHCT and ISVS. Analysis of covariance was employed to evaluate the effect of the blended learning approach on attitudes and self-efficacy posttest scores by using pretest scores as a covariate.Descriptive statistics were used to represent the demographic characteristics of the study population and the evaluation of the learning strategies. Analysis of variance with a posthoc test was carried out to determine the differences between the groups on demographic characteristics. A paired A total of 207 health care students participated in the blended learning activity. However, only 198 completed the questionnaires , with a response rate of 95.7% for the one-month posttest questionnaires .P=.62), gender (P=.81), type of qualification (P=.81), and type of health care course (P=.21). This suggested homogeneity of the participants between the three groups.As shown in P<.05) of self-efficacy in performing interprofessional team care after interprofessional learning. However, the SE-WI-VR group had the lowest self-efficacy posttest mean scores. Between-group comparisons using analysis of covariance revealed a significant difference (P=.03) among the three groups in terms of self-efficacy posttest mean scores, with the SE-WI-VR group reporting significantly lower posttest scores than the WI-SE-VR groups after controlling the pretest scores (P=.04). There were no significant differences between the WI-VR-SE and WI-SE-VR groups in self-efficacy posttest scores after controlling the pretest scores (P=.06).As shown in P<.001) from the baseline scores for all three groups. However, no significant differences were found between the baseline and posttest scores on the ATIHCT for all three groups. Between-group comparisons also did not identify any significant differences in the attitude posttest scores for both the ISVS and ATIHCT after controlling the pretest scores.As shown in In terms of the participants’ top preferences for interprofessional learning through the three different instructional sequences of a blended learning approach, the majority of the participants (69.1%) selected the instructional sequence “WI-VR-SE” as their top preference; in addition, less than 10% of the participants chose the sequence “SE-WI-VR” as their top preferences and about 20% chose the sequence “WI-SE-VR.”The evaluation of the participants’ levels of self-efficacy in contributing to interprofessional patient-centered care, which was conducted 1 month following the interprofessional learning activity, indicated significant improvements from the baseline scores for all three groups. Drawing from the constructivist learning theory, case-based and problem-based learning approaches were incorporated into the blended learning strategies to help learners develop competencies in performing interprofessional patient-centered care. These approaches are known to enable participants to actively construct their knowledge based on their interpretations of experiences . FurtherAmong the three groups, participants who undertook the SE-WI-VR sequence were found to have the lowest improvement of self-efficacy levels, which was significantly lower than those who undertook WI-SE-VR. These findings suggest that the instructional sequences of a blended learning approach can significantly influence learners’ perceptions of learning outcomes. Our findings show that learning starting with instructional prompts on concepts followed by problem solving in SE was a more effective learning sequence than the reverse sequence. Although the problem-solving activity in SE may prepare learners to benefit from WI learning, the introduction of concepts seemed to be necessary, particularly for novice health care students who may not have sufficient domain knowledge to engage in problem-solving tasks in SE . This stAlthough no significant differences in self-efficacy scores were found between the instructional sequences WI-SE-VR and WI-VR-SE, the instructional sequence WI-VR-SE was chosen by the majority of the students as their top preferences. In this WI-VR-SE instructional sequence, scaffolding support was provided initially in WI through a video demonstration of desired performances using cognitive tools. This was followed by facilitator-led experiential learning in VR that fostered reflection and metacognition. The support was gradually decreased in SE, where students were given the opportunity to collaborate among themselves to apply their learning to problem solve a case scenario. According to Kim and Hannafin , in the Unlike self-efficacy in performing interprofessional team care, no significant differences were reported among the three groups in their attitudes toward interprofessional socialization and the health care team. Given the opportunities to engage in collaborative learning in VR and SE, all groups reported a significant improvement in attitudes toward interprofessional socialization. According to the social constructivist views of learning, the exchange and discussion of ideas through social interaction are critical for learners to construct meaningful knowledge . AlthougAlthough a robust study using a randomized controlled trial and a large sample size was employed in this study, an important limitation is the lack of performance measurement as the outcome measure. The use of self-reported surveys that measure changes in attitudes and self-perceived efficacy may not predict actual performances and may be subjected to social desirability. Future studies can measure the impact on team performance. Another limitation is that we did not incorporate a debriefing or feedback session in the SE after the students’ role-play experiences, which could have served as expert feedback and engage learners in reflection on actions.In conclusion, this study provides evidence of the effectiveness of a blended learning approach using a randomized controlled study. Technologically enhanced learning strategies based on the constructivist learning theory improves health care students’ interprofessional competencies. The study suggests that the instructional sequence of a blended learning approach can have a significant impact on students’ learning outcomes. The more effective learning sequence allows learners to grasp concepts from WI before presenting them with a problem-solving activity in SE. From the learners’ perspective, they preferred to start with concept building using WI, followed by experiential learning in VR and subsequent application through SE. This study provides recommendation for future practice to scaffold students’ learning when planning the instructional sequence of technologically enhanced learning activities within blended learning environments. Future studies can undertake a more robust outcome measurement by evaluating the effects of the instructional sequence on team performance.


Benign prostatic hyperplasia (BPH) rarely occurs in children or young males. In this case report, a 29-year-old male patient diagnosed with BPH coexisting with ketamine-associated uropathy was reported to investigate the possible relationship between BPH and ketamine-associated uropathy as well as therapeutic strategies.A 29-year-old male patient with a 3-year history of ketamine inhalation, complaining of dysuria with frequency and urgency, was admitted. Hydronephrosis, hydroureters, uneven bladder wall thickening and a tumour located in the outlet of the bladder were detected with computed tomography (CT). The patient agreed to cystoscopy under general anaesthesia. A spherical tumour with a diameter of approximately 2 cm was found to originate from the median lobe of the prostate and follicular lesions were diffusely distributed on the right bladder wall. The tumour and follicular lesions in the bladder were resected successfully, and pathology demonstrated BPH and chronic inflammation of the mucous membranes separately. The patient quit ketamine completely during the one-year follow-up. Dysuria was relieved completely and no tumour or follicular neoplasm recurrence was found.Inflammation in the urothelium, as a direct or indirect consequence of ketamine, may contribute to the development of BPH. Both surgical interventions to remove obstruction and ketamine cessation are necessary approaches. Benign prostatic hyperplasia (BPH) is a common disease that usually develops in middle-aged and older men. The occurrence of BPH in children or young men seems to be very rare. This report presents a case of BPH with ketamine-associated uropathy in a young patient which, to our knowledge, has never been reported before.A 29-year-old male patient was admitted on April 2017 because of dysuria with frequency and urgency. The patient began quitting ketamine for one year after 3 years of ketamine inhalation (once weekly at first and then once every 2–3 days up to a maximum of approximately 10 g/day). Duplex ultrasonography of the urinary system showed bilateral hydronephrosis and hydroureters, a thick-walled bladder and a post-void residual volume of approximately 100 ml. Computed tomography (CT) showed moderate bilateral hydronephrosis and hydroureters  seems to represent a prominent difference in BPH between elderly men and young males. According to the cases listed in Table To our knowledge, there has been no case of BPH with ketamine-associated uropathy ever reported in young patients. The exact pathophysiology of ketamine-associated cystitis remains greatly unknown. An important theory is that ketamine and/or its metabolites accumulates at a high concentration in urine, leading to a direct toxic effect on the urothelium and resulting in significant inflammation. The nuclear factor-kappa B (NF-κB) pathway was found to be activated during inflammatory signalling of ketamine-induced cystitis in the bladder in a rat model . FurtherKetamine-induced upper urinary tract lesions seem not to be paid enough attention due to a greater incidence of apparent lower urinary tract syndromes . HydroneThe general treatment of BPH in elderly men includes medical and surgical approaches. Medical approaches may not work due to the indefinite aetiology of BPH in young men. For ketamine-associated uropathy, the aims of management of ketamine-associated urinary symptoms are symptom control and preservation of renal function and are ultimately to prevent irreversible damage to the urinary tract . Due to The simultaneous occurrence of BPH and ketamine-associated uropathy in young males has never been reported before. Inflammation in the urinary system, as a consequence of ketamine, may contribute to the development of BPH. Both surgical interventions and ketamine cessation are important approaches for relieving symptoms. This case may deserve significant attention as ketamine addiction among young population increases globally.