Long-term follow-up of liver-directed, adeno-associated vector-mediated gene therapy in the canine model of hemophilia A.

Questions remain concerning the long-term efficacy, safety, and site(s) of transgene expression following adeno-associated vector (AAV) therapy. We report a long-term follow-up of 8 (male = 4, hemizygous, and female = 4, homozygous) dogs with severe hemophilia A treated with a single portal vein infusion of a B-domain-deleted (BDD)-canine FVIII (cFVIII) AAV vector (median dose = 1.25 × 1013 vg/kg, AAV2 = 4, AAV6 = 3, and AAV8 = 1). After a median follow-up of 10.8 years (8.2-12.0 years), persistent FVIII:C (median one-stage = 12.7%, chromogenic = 7.2%) was seen in all responding dogs (n = 6), with improvement in annualized bleed rates (pre = 3.9 vs post = 0.3 event per year; P = .003). Anti-AAV capsid neutralizing antibodies (nAbs) toward the dosed capsid were detected throughout the study, with limited cross-reactivity to other capsids. nAb titers for all capsid serotypes declined with time, although they remained at levels precluding redosing with the same capsid. AAV-BDD-cFVIII DNA was detected in the liver of all dogs (median = 0.15 vg per diploid genome), with lower levels in the spleen in 4 dogs (median = 0.005 vg per diploid genome). Consistent with the liver-specific promoter, BDD-cFVIII mRNA was only detected in the liver. Postmortem examination demonstrated no evidence of chronic liver disease or liver malignancy. Persistent FVIII expression and an improved bleeding phenotype was seen for more than a decade after vector delivery. This is the longest follow-up reported in a preclinical model supporting long-term efficacy and safety of AAV-mediated gene therapy.

Systematic development of a mobile preconception lifestyle programme for couples undergoing IVF: the PreLiFe-programme.

STUDY QUESTION: Can we develop a preconception lifestyle programme for couples undergoing IVF that is in line with their needs. SUMMARY ANSWER: A mobile preconception lifestyle programme was systematically developed based on expert opinion, literature and needs of IVF-patients. WHAT IS KNOWN ALREADY: A healthy lifestyle prior to conception is not only beneficial for the general health of couples, but evidence on its importance for their reproductive health and the health of their children is also emerging. So far, the vast majority of fertility clinics do not offer a lifestyle programme for couples undergoing IVF. Therefore, the present study aimed to develop a lifestyle programme for IVF-couples. STUDY DESIGN, SIZE, DURATION: The development of the PreLiFe-programme was guided by the steps of the Medical Research Council (MRC) framework for developing complex interventions, a systematic approach for developing theory- and evidence-based health promotion interventions. PARTICIPANTS/MATERIALS, SETTINGS, METHODS: First, the evidence base on lifestyle programmes for IVF-couples was reviewed. Second, several iterations between an expert panel, the literature, and quantitative and qualitative data from IVF-patients identified the content, the format, behaviour change techniques and theory of the programme. Third, the PreLiFe-programme was produced and the expected process and outcomes of a randomized controlled trial assessing it were modelled. Finally, user tests among experts and patients and pilot tests among patients were conducted. MAIN RESULTS AND ROLE OF CHANCE: The finally developed PreLiFe-programme is a mobile application to be used autonomously by both partners of IVF-couples during the first year of IVF, in combination with motivational interviewing over the telephone every three months (i.e. blended care). The PreLiFe-programme provides advice and skills training on physical activity, diet and mindfulness based stress reduction and is in part tailored based on monitoring and tracking the lifestyle of patients. Based on the literature the expert panel considers it plausible that all three components contribute to IVF-success rates. The PreLiFe-programme is likely to be acceptable to patients as it meets the need of patients for lifestyle advice and treatment information. LIMITATIONS, REASON FOR CAUTION: The pilot in IVF-couples had a 3-month duration. The feasibility of the PreLiFe-programme in other infertile populations and/or upon longer use is yet to be examined. Whether the PreLiFe-programme effectively improves lifestyle and IVF-success rates is currently being examined in a trial randomizing heterosexual couples starting IVF to the PreLiFe-programme or an attention-control group for 12 months. WIDER IMPLICATIONS OF THE FINDINGS: If the PreLiFe-programme improves lifestyle and the chance of IVF-success, it will be a powerful tool and provide guidance for implementing lifestyle programmes in fertility clinics. STUDY FUNDING/COMPETING INTEREST(S): Funded by the Research Foundation Flanders (FWO-TBM (Applied Biomedical Research with a Primary Social finality); reference: T005417N). The authors have no conflict of interest to report. TRIAL REGISTRATION NUMBER: NCT03790449.

Impact of ARTs on oncological outcomes in young breast cancer survivors.

STUDY QUESTION: What is the risk of recurrence in young breast cancer survivors who undergo ARTs following completion of anticancer treatment? SUMMARY ANSWER: ART in breast cancer survivors does not appear to have a negative impact on disease-free survival. WHAT IS KNOWN ALREADY: In healthy women, fertility treatment does not increase the risk of developing breast cancer. At the time of breast cancer diagnosis and before starting anticancer treatments, several studies have shown the safety of performing ART. However, the safety of ART in breast cancer survivors following completion of anticancer treatment remains under-investigated. In general, breast cancer survivors are counselled to avoid any hormonal treatment but there are limited data available on the effect of short exposure to high oestradiol levels during ART. The largest study in this regard included 25 breast cancer survivors exposed to ART and did not show a detrimental effect of ART on patient survival. Hence, taking into account that pregnancy after breast cancer does not affect cancer prognosis, defining the safety of ART in breast cancer survivors remains a priority. STUDY DESIGN, SIZE, DURATION: We conducted a retrospective multicentric matched cohort study including a cohort of breast cancer survivors who underwent ART (exposed patients) between January 2006 and December 2016. Exposed patients who were eligible for the study were matched according to known breast cancer prognostic factors. Matched breast cancer survivors did not undergo ART (non-exposed patients) and were disease-free for a minimum time that was not less than the time elapsed between breast cancer diagnosis and first ART for the matched ART-exposed patients. PARTICIPANTS/MATERIALS, SETTING, METHODS: Data were retrieved from all survivors who had been diagnosed with breast cancer in eight participating centres at an age of ≤40 years, without metastasis, ongoing pregnancy, pre-existing neoplasia or ovarian failure. ART included ovarian stimulation for IVF/ICSI, clomiphene citrate treatment and hormone replacement therapy for embryo transfer. Data were collected from an oncological database for the selection of breast cancer patients in the non-exposed group. Exposed patients were matched (1:2) for germline BRCA status, tumour stage, anticancer treatment and age, whenever feasible. Matched groups were compared at baseline according to characteristics using conditional logistic regression. Kaplan-Meier curves were constructed to compare time to recurrence between groups, with the time of ART as starting point that has been adjusted in the non-exposed group. The analyses were performed using Stata IC/15.1. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 39 breast cancer patients in the ART group were eligible for the analysis and were matched with 73 controls. There was no statistical difference between the two groups for the presence of BRCA mutation, tumour characteristics, use of (neo)adjuvant chemotherapy and of adjuvant endocrine therapy. Exposed patients were younger than non-exposed patients (mean age 31.8 vs 34.3 years, respectively; P < 0.001). In the ART group, 89.7% were nulliparous at diagnosis compared to 46.6% of controls (P < 0.001). ART was performed at a mean age of 37.1 years old, after a median time of 4.1 years following breast cancer diagnosis (range: 1.5-12.5). Median anti-Müllerian hormone at the time of ART was 0.28 ng/ml (range: 0-4.4) and median serum oestradiol peak level was 696.5 pg/ml (range: 139.7-4130). Median follow-up time from first attempt of ART was 4.6 years (range: 2.4-12.5) in the ART group. Adjusted follow-up time for the non-exposed group was 6.9 years (range: 1.1-16.5 years) (P = 0.004). In the ART group, 59% of patients had a pregnancy after breast cancer compared to 26% in the non-exposed patients (P = 0.001). Breast cancer relapsed in 7.7% versus 20.5% women in the ART and non-exposed groups, respectively (hazard ratio 0.46, 95% CI 0.13-1.62, P = 0.23). Median time to relapse was 1.3 (range: 0.3-2.7) years versus 4.5 (range: 0.4-11.1) years after ART and adjusted time in the ART and non-exposed groups, respectively (P = 0.14). LIMITATIONS, REASONS FOR CAUTION: Although this is the first and largest multicentric study addressing the impact of ART on breast cancer recurrence to provide data on oestrogen exposure, only a small number of patients could be included. This reflects the reluctance of breast cancer survivors and/or oncologists to perform ART, and highlights the need for a prospective data registry to confirm the safety of this approach. This would offer the possibility for these patients, who are at a high risk of infertility, to fully benefit from ART. WIDER IMPLICATIONS OF THE FINDINGS: Although recent studies have proven that pregnancy after breast cancer has no detrimental impact on prognosis, counselling patients about the safety of ART remains challenging. Our study provides reassuring data on the use of ART in breast cancer survivors with favourable prognostic factors, for when natural conception fails. STUDY FUNDING/COMPETING INTEREST(S): M.C. and I.D. are funded by FNRS, Télévie-FNRS and Fonds Erasme. M.D.V. is a CooperSurgical scientific advisory board member and receives lecture fees for MSD, Gedeon-Richter and Ferring, outside the submitted work. M.L. has acted as a consultant for Roche and Novartis and has received honoraria from Theramex, Roche, Lilly, Pfizer, Novartis and Takeda, outside the submitted work. I.D. has acted as a consultant for ROCHE and has received speaker's fees from Novartis, outside the submitted work. E.d.A. has received honoraria and is a Roche/GNE, Novartis, SeaGen and Zodiac scientific advisory board member, has received travel grants from Roche/GNE and GSK/Novartis, and has received research grants from Roche/GNE, Astra-Zeneca, GSK/Novartis and Servier, outside the submitted work. A.D. is a recipient of a research grant from Ferring Pharmaceuticals and receives lecture and/or consultancy fees from Merck, Gedeon-Richter and Ferring Pharmaceuticals, outside the submitted work. The remaining authors have no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: N/A.

Diversity of activity participation determines bone mineral content in the lower limbs of pre-pubertal children with developmental coordination disorder.

This study examined the relationships between activity participation and bone mineralization in children with developmental coordination disorder. Limited participation in physical, recreational, social, and skill-based and self-improvement activities contributed to lower bone mineral content. For improved bone health, these children should participate in a variety of activities, not only physical activities. INTRODUCTION: Limited activity participation in children with developmental coordination disorder (DCD) may have a negative impact on bone mineral accrual. The objectives of this study were to compare bone mineralization and activity participation patterns of pre-pubertal children with DCD and those with typical development, and to determine the association between activity participation patterns and bone mineralization in children with DCD. METHODS: Fifty-two children with DCD (mean age = 7.51 years) and 61 children with typical development (mean age = 7.22 years) participated in the study. Appendicular and total body (less head) bone mineral content (BMC) and bone mineral density (BMD) were evaluated by a whole-body dual-energy X-ray absorptiometry scan. Activity participation patterns were assessed using the Children's Assessment of Participation and Enjoyment (CAPE) questionnaire. RESULTS: Children with DCD had lower appendicular and total body BMCs and BMDs than children with typical development overall (p < 0.05). They also had lower CAPE total activity and physical activity diversity scores (p < 0.05). After accounting for the effects of age, sex, height, lean mass, and fat mass, the total activity diversity score remained independently associated with leg BMC in children with DCD, explaining 5.1% of the variance (p = 0.030). However, the physical activity diversity score was no longer associated with leg BMC (p = 0.090). CONCLUSIONS: Diversity of activity participation and bone mineralization were lower in pre-pubertal children with DCD. Decreased total activity participation diversity was a contributing factor to lower BMC in the legs of children with DCD.

Stable pneumorectum using an inline glove - a cost-effective technique to facilitate transanal total mesorectal excision.

AIM: Transanal total mesorectal excision (taTME) is a novel approach for resection of the rectum. Use of a standard insufflator to create pneumorectum, however, results in bellowing-large heaving motions from insufflation of air that can frustrate surgery. We report the successful application of our technique, stable pneumorectum using an inline glove (SPRING), for the performance of transanal rectal excision in a series of 17 patients using a standard laparoscopic insufflator. METHOD: A retrospective review of 17 patients using the SPRING technique was performed between October 2015 and October 2016. Characteristics of these patients were evaluated, and technique-related short-term outcome was reviewed. RESULTS: The SPRING technique was successfully used in patients who underwent both minimally invasive (n = 14) and open (n = 3) approaches in the abdominal stage of the surgery. In the 12 patients who had rectal cancer for whom SPRING was used to facilitate taTME there were no conversions to an alternative access for rectal resection, the median duration of the TME part of the operation was 95 min (62-147) and there was one R1 resection (8%). Billowing was not a significant problem in any of the 17 patients during the surgery. CONCLUSION: In this case series we have successfully shown the feasibility of the SPRING technique as a practical and cost-effective solution to the problem of billowing during taTME.

Strategy to detect pre-existing immunity to AAV gene therapy.

Gene therapy may offer a new treatment option, particularly for patients with severe hemophilia, based on recent research. However, individuals with pre-existing immunity to adeno-associated viruses (AAVs) may be less likely to benefit from AAV vector-based therapies. To study pre-existing AAV5 immunity in humans, we validated two complementary, sensitive, and scalable in vitro assays to detect AAV5 total antibodies and transduction inhibition (TI). Using these two assays, we found that 53% of samples from 100 healthy male individuals were negative in both assays, 18% were positive in both assays, 5% were positive for total antibodies but negative for TI and, of interest, 24% were negative for total antibodies but positive for TI activity, suggesting the presence of non-antibody-based neutralizing factors in human plasma. Similar findings were obtained with 24 samples from individuals with hemophilia A. On the basis of these results, we describe the development of a dual-assay strategy to identify individuals without total AAV5 antibodies or neutralizing factors who may be more likely to respond to AAV5-directed gene therapy. These assays offer a universal, transferrable platform across laboratories to assess the global prevalence of AAV5 antibodies and neutralizing factors in large patient populations to help inform clinical development strategies.

Hyperglycemia impedes definitive endoderm differentiation of human embryonic stem cells by modulating histone methylation patterns.

Exposure to maternal diabetes during fetal growth is a risk factor for the development of type II diabetes (T2D) in later life. Discovery of the mechanisms involved in this association should provide valuable background for therapeutic treatments. Early embryogenesis involves epigenetic changes including histone modifications. The bivalent histone methylation marks H3K4me3 and H3K27me3 are important for regulating key developmental genes during early fetal pancreas specification. We hypothesized that maternal hyperglycemia disrupted early pancreas development through changes in histone bivalency. A human embryonic stem cell line (VAL3) was used as the cell model for studying the effects of hyperglycemia upon differentiation into definitive endoderm (DE), an early stage of the pancreatic lineage. Hyperglycemic conditions significantly down-regulated the expression levels of DE markers SOX17, FOXA2, CXCR4 and EOMES during differentiation. This was associated with retention of the repressive histone methylation mark H3K27me3 on their promoters under hyperglycemic conditions. The disruption of histone methylation patterns was observed as early as the mesendoderm stage, with Wnt/ß-catenin signaling being suppressed during hyperglycemia. Treatment with Wnt/ß-catenin signaling activator CHIR-99021 restored the expression levels and chromatin methylation status of DE markers, even in a hyperglycemic environment. The disruption of DE development was also found in mouse embryos at day 7.5 post coitum from diabetic mothers. Furthermore, disruption of DE differentiation in VAL3 cells led to subsequent impairment in pancreatic progenitor formation. Thus, early exposure to hyperglycemic conditions hinders DE development with a possible relationship to the later impairment of pancreas specification.

Polycystic ovarian morphology and the diagnosis of polycystic ovary syndrome: redefining threshold levels for follicle count and serum anti-Müllerian hormone using cluster analysis.

STUDY QUESTION: Can cluster analysis be used to differentiate between normo-ovulatory women with normal ovaries and normo-ovulatory women with polycystic ovarian morphology (PCOM) in a non-subjective manner? SUMMARY ANSWER: Cluster analysis can be used to accurately and non-subjectively differentiate between normo-ovulatory women with normal ovaries and normo-ovulatory women with PCOM. WHAT IS KNOWN ALREADY: Currently, PCOM is diagnosed using a fixed threshold level, i.e. 12 or more follicles per ovary, and is one of the diagnostic criteria of polycystic ovary syndrome (PCOS). However, PCOM is also encountered in normo-ovulatory women, suggesting that it could just represent a normal variant. On the other hand, recent studies have shown subtle endocrine abnormalities in women with isolated PCOM that resemble those found in women with PCOS. Because of the strong correlation between anti-Müllerian hormone (AMH) and follicle number, a high serum AMH level has been proposed as a surrogate marker for PCOM and could, therefore, be integrated in the diagnostic classifications for PCOS. STUDY DESIGN, SIZE, DURATION: This was a retrospective observational cohort study. Original cohorts had been recruited for previous studies between 1998 and 2010. Two hundred ninety-seven regularly cycling women and 700 women with PCOS were eligible for inclusion. PARTICIPANTS/MATERIALS, SETTING, METHODS: Cluster analysis was performed in 297 regularly cycling women. After exclusion of 'PCOM' clusters, each 'non-PCOM' cluster (young, n = 118 and old, n = 100) was included in the construction of a receiver operating characteristics curve to test the diagnostic performance of follicle number per ovary (FNPO) and serum AMH in discriminating similarly aged full-blown PCOS patients (n = 411 and 237, respectively) from normal regularly cycling non-PCOM women. MAIN RESULTS AND ROLE OF CHANCE: The optimal number of clusters was four; age was the most important classifying variable, followed by the FNPO and serum AMH. Two distinct clusters of normo-ovulatory women with PCOM were isolated and differed solely by age, i.e. 'young' and 'old'. Both 'PCOM' clusters had their similarly aged counterpart of 'non-PCOM' clusters. Likewise, two clusters comprised women younger than 30 years, with (n = 28, 'PCOM regularly cycling women') or without (n = 118, 'normal regularly cycling women') features of PCOM (increased FNPO and/or serum AMH). The two other clusters in older women could be labelled 'normal regularly cycling women' or 'PCOM regularly cycling women' (n = 100 and 51, respectively). The prevalence of PCOM was significantly greater in old than in young regularly cycling women controls. In the young population, after exclusion of the 'PCOM regularly cycling women', the diagnostic performance of AMH, expressed by area under the curve (AUC) (AUC = 0.903; CI (0.876-0.930)) to differentiate PCOS women from normal regularly cycling women was similar to that using the FNPO (AUC = 0.915, CI (0.891-0.940)) (P = 0.25), confirming results from earlier studies. In the old population, the diagnostic performance of AMH was greater than that of FNPO (AUCs = 0.948 (0.927-0.970) vs 0.874 (0.836-0.912), respectively, P = 0.00035). Cut-off levels of AMH and antral follicle count distinguishing regularly cycling non-PCOM women from PCOS women were higher in young women than in older women. LIMITATIONS, REASONS FOR CAUTION: Data of normal women were obtained from earlier studies, aiming to measure normal endocrine values. Apparently, the strong effect of age in cluster analysis revealed a dichotomy in the age distribution among the cohort of regularly cycling women included. This was involuntary since in none of the original studies, eligibility was limited by age and there was considerable overlap in age ranges of the cohorts. Transvaginal ultrasound was performed using a 6.5-8 mHz probe and our data confirm that this threshold level for FNPO is still valid if using such probe frequencies, although the use of devices with a maximum frequency lower than 8 mHz has become obsolete. Obviously, newer ultrasound scanner using higher transducer frequency will facilitate the detection of more follicles. WIDER IMPLICATIONS OF THE FINDINGS: Our data support the use of AMH as a surrogate for ultrasound to define PCOM, which is one of the three items of the Rotterdam classification. They also show that age should be taken into account to define the optimal threshold. The fact that the prevalence of PCOM was increased in the older regularly cycling women, may be due to 'attenuated' PCOS, a phenomenon that has been described in ageing women with PCOS. These women might have had anovulatory cycles in the past and have become ovulatory with increasing age, and were, therefore, eligible for this study. However, since most women included at older age have had spontaneous pregnancies in the past, PCOM at older age may be associated with a subclinical form of PCOS, which may also be present in young regularly cycling women. STUDY FUNDING/COMPETING INTEREST(S): No funding was received for this study. J.S.E.L. has received grants and support from Ferring, MSD, Organon, Merck-Serono, Schering Plough and Serono during recruitment and analysis of data for this study. S.L.F., A.D. and D.D. do not have any conflict of interest.

A randomized controlled trial of self-regulated modified constraint-induced movement therapy in sub-acute stroke patients.

BACKGROUND AND PURPOSE: Emerging research suggests the use of self-regulation (SR) for improving functional regain in patients post stroke. SR is proposed to produce an added effect to effective modified constraint-induced movement therapy (mCIMT). This study aimed to examine the effect of a self-regulated mCIMT programme (SR-mCIMT) for functional regain in patients with sub-acute stroke. METHODS: Eighty-six patients completed the trial: SR-mCIMT, n = 29; mCIMT, n = 31; or conventional functional rehabilitation, n = 26. All interventions were 2-week therapist-guided training. Outcome measurements, taken by a blinded assessor, examined arm function [Action Research Arm Test (ARAT), Fugl-Meyer Assessment (FMA)], daily task performance [Lawton Instrumental Activities of Daily Living Scale (Lawton IADL)] and self-perceived arm use in functional tasks [Motor Activity Log (MAL)]. RESULTS: Significant differences were found with the SR-mCIMT outperforming the other groups after the intervention (ARAT, P = 0.006; FMA, Lawton IADL and MAL, all Ps < 0.001). In terms of the carry-over effect, the SR-mCIMT group outperformed in the hand and coordination subscales of ARAT and FMA (P = 0.012-0.013) and the self-perceived quality of arm use (P = 0.002). CONCLUSION: A combination of SR and mCIMT could produce an added effect in functional regain in patients post stroke.

Apical lymph node dissection of the inferior mesenteric artery.

AIM: It is controversial whether a high or low ligation of the inferior mesenteric artery (IMA) is superior. The former allows an extended lymph node clearance whereas the latter preserves the distal vascular supply via the left colic artery (LCA). Apical lymph node dissection of the IMA (ALMA) harvests nodal tissue along the IMA proximal to the LCA whilst performing a low ligation. This anatomically replicates the oncological benefit of high ligation and the vascular preservation of low ligation. Our study evaluates the nodal yield of ALMA and the short-term outcome of this technique. METHOD: We retrospectively studied 19 patients with sigmoid or rectal cancer who underwent curative surgical resection with ALMA. All ALMAs were performed with a standard technique previously described (Kobayashi et al., Surg Endosc 2005, 20:563-9; Sekimoto et al. Surg Endosc 2010, 25:861-6) . The lymph node yield from the dissection (the ALMA specimen) was compared with the total lymph node yield. Data on the LCA anatomy, time required to perform ALMA, complications and postoperative recovery were evaluated. RESULTS: ALMA was successful in 18 patients. Median postoperative hospitalization was 5 (2-26) days without ALMA-related morbidity or mortality. The median lymph node yield was 20 (9-41) and a median of 14.3 (0-80)% were harvested with ALMA. Two patients not having neoadjuvant chemoradiotherapy had fewer than 12 lymph nodes, excluding nodes harvested from ALMA. The average time required for ALMA was 18 min. CONCLUSION: ALMA is a safe and feasible technique, allowing extended lymphadenectomy without sacrificing the LCA. In this small group of patients none were upstaged due to cancerous involvement of the proximal nodes.

Laparoscopic excision of retrorectal tumours.

AIM: The aim of this study was to demonstrate a laparoscopic technique for the excision of retrorectal tumours. METHOD: A retrospective review of cases identified from a prospectively maintained database was undertaken. RESULTS: Ten patients (seven female; median age 45 years (range 23-79) underwent successful laparoscopic excision with no significant morbidity or intra-operative mishaps. CONCLUSION: The procedure was deemed to be safe and we include a video to show the operative technique.

Surgical strategies in the management of recurrent retrorectal tumours.

BACKGROUND: The aim of this study was to review a consecutive series of patients who had undergone excision of recurrent retrorectal tumours and propose surgical strategies to tackle such recurrences. METHODS: Patients were identified from a prospectively maintained database. Demographic details, preoperative imaging and pathology, intra- and post-operative problems and follow-up details were noted. RESULTS: Fifteen patients (11 females) with a median age of 38 years (range 19-75 years) underwent excision of recurrent retrorectal tumours (13 benign) between 2002 and 2012. The median interval between the first and second surgical procedure was 3.5 years (range 1-19 years). Three patients had surgery performed via the transperineal approach, while 12 patients had resection via the abdominal approach. En bloc resection of adjacent organs was needed in three patients. Major pelvic bleeding occurred in two patients. R0 resection was achieved in all 15 patients, and there have been no subsequent recurrences [median follow-up 73 months (range 12-148 months)]. CONCLUSIONS: Benign recurrent retrorectal tumours can be safely excised usually without sacrifice of adjacent organs, while en bloc resection is needed for malignant tumours.

[Identification of endogenous control genes for gene expression studies in peripheral blood of patients with coronary artery disease].

The selection of stable endogenous control genes is critical for normalization of quantitative real-time PCR (qPCR) data. In this study, we aimed to identify a suitable set of control genes to be used as endogenous references for gene expression evaluation in human peripheral blood samples among coronary artery disease patients. The expression levels of 12 endogenous control genes procured from TATAA Biocenter (Goteborg, Sweden) were measured in five acute coronary syndrome patients and five chronic stable angina patients. Gene expression stability was analyzed using two different software applications i.e geNorm and NormFinder. Results suggested that beta-glucuronidase is the most stable endogenous control, followed by hypoxanthine-guanine phosphoribosyltransferase. The NormFinder analysis further confirmed that beta-glucuronidase and hypoxanthine-guanine phosphoribosyltransferase were on the first rank order with the most stable expression among endogenous control genes analyzed and 60S acidic ribosomal protein P0. Besides this, the expression levels of 18S rRNA were revealed to be highly variable between coronary heart disease patients. We thus recommend the use of beta-glucuronidase and hypoxanthine-guanine phosphoribosyltransferase as reference genes for accurate normalization of relative quantities of gene expression levels in coronary artery disease patients using qPCR. Also the use of 18S rRNA as a control gene should be avoided.

APSIC guide for prevention of catheter associated urinary tract infections (CAUTIs).

BACKGROUND: The Asia Pacific Society of Infection Control launched the APSIC guide for prevention of catheter associated urinary tract infections in July 2022. It aims to highlight practical recommendations in a concise format designed to assist healthcare facilities in the Asia Pacific region to achieve high standards in infection prevention and control practices during the management and care of patients with a urinary catheter. METHODS: The guidelines were developed by an appointed workgroup comprising experts in the Asia Pacific region, following reviews of previously published guidelines and recommendations relevant to each section. RESULTS: It recommends that healthcare institutions have a catheter associated urinary tract infection prevention program that includes surveillance and the use of the insertion and maintenance bundles. Implementation of the bundles is best done using a quality improvement approach with a multidisciplinary team. CONCLUSIONS: Healthcare facilities should aim for excellence in care of patients with urinary catheters. It is recommended that healthcare facilities have a catheter associated urinary tract infection prevention program as part of their Infection Prevention and Control program.

Lipid and glucose abnormalities and associated factors among children living with HIV in Asia.

BACKGROUND: Children living with HIV (CLHIV) on prolonged antiretroviral therapy (ART) are at risk for lipid and glucose abnormalities. Prevalence and associated factors were assessed in a multicentre, Asian longitudinal paediatric cohort. METHODS: CLHIV were considered to have lipid or glucose abnormalities if they had total cholesterol ≥200 mg/dL, high-density lipoprotein (HDL) ≤35 mg/dL, low-density lipoprotein (LDL) ≥100 mg/dL, triglycerides (TG) ≥110 mg/dL, or fasting glucose >110 mg/dL. Factors associated with lipid and glucose abnormalities were assessed by logistic regression. RESULTS: Of 951 CLHIV, 52% were male with a median age of 8.0 (interquartile range [IQR] 5.0-12.0) years at ART start and 15.0 (IQR 12.0-18.0) years at their last clinic visit. 89% acquired HIV perinatally, and 30% had ever used protease inhibitors (PIs). Overall, 225 (24%) had hypercholesterolemia, 105 (27%) low HDL, 213 (58%) high LDL, 369 (54%) hypertriglyceridemia, and 130 (17%) hyperglycemia. Hypercholesterolemia was more likely among females (versus males, aOR 1.93, 95% CI 1.40-2.67). Current PIs use was associated with hypercholesterolemia (current use: aOR 1.54, 95% CI 1.09-2.20); low HDL (current use: aOR 3.16, 95% CI 1.94-5.15; prior use: aOR 10.55, 95% CI 2.53-43.95); hypertriglyceridemia (current use: aOR 3.90, 95% CI 2.65-5.74; prior use: aOR 2.89, 95% CI 1.31-6.39); high LDL (current use: aOR 1.74, 95% CI 1.09-2.76); and hyperglycemia (prior use: aOR 2.43, 95% CI 1.42-4.18). CONCLUSION: More than half and one-fifth of CLHIV have dyslipidemia and hyperglycemia, respectively. Routine paediatric HIV care should include metabolic monitoring. The association between PIs use and dyslipidemia emphasizes the importance of rapidly transitioning to integrase inhibitor-containing regimens.

Study of transforming growth factor alpha for the maintenance of human embryonic stem cells.

Human embryonic stem cells (hESCs) have great potential for regenerative medicine as they have self-regenerative and pluripotent properties. Feeder cells or their conditioned medium are required for the maintenance of hESC in the undifferentiated state. Feeder cells have been postulated to produce growth factors and extracellular molecules for maintaining hESC in culture. The present study has aimed at identifying these molecules. The gene expression of supportive feeder cells, namely human foreskin fibroblast (hFF-1) and non-supportive human lung fibroblast (WI-38) was analyzed by microarray and 445 genes were found to be differentially expressed. Gene ontology analysis showed that 20.9% and 15.5% of the products of these genes belonged to the extracellular region and regulation of transcription activity, respectively. After validation of selected differentially expressed genes in both human and mouse feeder cells, transforming growth factor α (TGFα) was chosen for functional study. The results demonstrated that knockdown or protein neutralization of TGFα in hFF-1 led to increased expression of early differentiation markers and lower attachment rates of hESC. More importantly, TGFα maintained pluripotent gene expression levels, attachment rates and pluripotency by the in vitro differentiation of H9 under non-supportive conditions. TGFα treatment activated the p44/42 MAPK pathway but not the PI3K/Akt pathway. In addition, TGFα treatment increased the expression of pluripotent markers, NANOG and SSEA-3 but had no effects on the proliferation of hESCs. This study of the functional role of TGFα provides insights for the development of clinical grade hESCs for therapeutic applications.

[Ultraviolet Mie lidar observations of aerosol extinction in a dust storm case over Macao].

Atmospheric aerosol over Macao was monitored by using a 355 nm Mie scattering lidar during the dust event on March 22nd, 2010. Vertical profiles of aerosol extinction coefficients were obtained and correlated with local PM10 concentration. The near-surface aerosol extinction coefficients have good agreement with PM10 concentration values. The aerosol extinction vertical profiles showed that there were distinct layers of dust aerosol concentration. The source and tracks of dust aerosol were analyzed by back-trajectory simulation. Observations showed that this lidar could run well even in dust storm episode, and it would help to further the study on aerosol properties over Macao.

Attitudes and practices of healthcare providers regarding gestational diabetes: results of a survey conducted at the 2010 meeting of the International Association of Diabetes in Pregnancy Study Group (IADPSG).

OBJECTIVE: To investigate the attitudes and practices of healthcare providers regarding gestational diabetes mellitus and to identify differences between provider types and geographical locations. RESEARCH DESIGN AND METHODS: A survey was distributed to participants at the 2010 GDM Pasadena II: International Conference regarding management of gestational diabetes. RESULTS: Of 250 distributed surveys, there was a 60.8% (n = 152) response rate. Most responders believed it was cost beneficial to treat and screen for gestational diabetes. Approximately 2/3 accepted that the diagnosis of Type 2 diabetes could be made in pregnancy. For the diagnosis of gestational diabetes, c. 60% opted for a one-step screen while 30% opted for a two-step screen. Most responders were comfortable treating a non-pregnant woman with Type 2 diabetes. Ninety per cent of responders initiated antenatal testing at 32-34 weeks for medication-treated gestational diabetes, with a wider variation for diet-treated gestational diabetes. Obstetricians were less comfortable than other providers in treating a non-pregnant woman with Type 2 diabetes, but were more likely to prescribe glyburide. Providers in the USA, compared with those from other countries, were more likely to perform a Caesarean section based on fetal weight, use glibenclamide (glyburide), initiate exercise post-delivery and less likely to check 2-h postprandial glucose levels. At least 80% of responders routinely screened for Type 2 diabetes at 6-12 weeks post-delivery. CONCLUSION: There are differences internationally and by provider types in attitudes and management regarding gestational diabetes. These findings may be useful in developing strategies for implementing the recent guidelines by the International Association of the Diabetes and Pregnancy Study Groups.

The effect of dichloroacetate on health- and lifespan in C. elegans.

Aging is associated with increased vulnerability to chronic, degenerative diseases and death. Strategies for promoting healthspan without necessarily affecting lifespan or aging rate have gained much interest. The mitochondrial free radical theory of aging suggests that mitochondria and, in particular, age-dependent mitochondrial decline play a central role in aging, making compounds that affect mitochondrial function a possible strategy for the modulation of healthspan and possibly the aging rate. Here we tested such a "metabolic tuning" approach in nematodes using the mitochondrial modulator dichloroacetate (DCA). We explored DCA as a proof-of-principle compound to alter mitochondrial parameters in wild-type animals and tested whether this approach is suitable for reducing reactive oxygen species (ROS) production and for improving organismal health- and lifespan. In parallel, we addressed the potential problem of operator bias by running both unblinded and blinded lifespan studies. We found that DCA treatment (1) increased ATP levels without elevating oxidative protein damage and (2) reduced ROS production in adult C. elegans. DCA treatment also significantly prolonged nematode health- and lifespan, but did not strongly impact mortality doubling time. Operator blinding resulted in considerably smaller lifespan-extending effects of DCA. Our data illustrate the promise of a "metabolic tuning" intervention strategy, emphasize the importance of mitochondria in nematode aging and highlight operator bias as a potential confounder in lifespan studies.