Increased Cardiac Arrhythmogenesis Associated With Gap Junction Remodeling With Upregulation of RNA-Binding Protein FXR1.

BACKGROUND: Gap junction remodeling is well established as a consistent feature of human heart disease involving spontaneous ventricular arrhythmia. The mechanisms responsible for gap junction remodeling that include alterations in the distribution of, and protein expression within, gap junctions are still debated. Studies reveal that multiple transcriptional and posttranscriptional regulatory pathways are triggered in response to cardiac disease, such as those involving RNA-binding proteins. The expression levels of FXR1 (fragile X mental retardation autosomal homolog 1), an RNA-binding protein, are critical to maintain proper cardiac muscle function; however, the connection between FXR1 and disease is not clear. METHODS: To identify the mechanisms regulating gap junction remodeling in cardiac disease, we sought to identify the functional properties of FXR1 expression, direct targets of FXR1 in human left ventricle dilated cardiomyopathy (DCM) biopsy samples and mouse models of DCM through BioID proximity assay and RNA immunoprecipitation, how FXR1 regulates its targets through RNA stability and luciferase assays, and functional consequences of altering the levels of this important RNA-binding protein through the analysis of cardiac-specific FXR1 knockout mice and mice injected with 3xMyc-FXR1 adeno-associated virus. RESULTS: FXR1 expression is significantly increased in tissue samples from human and mouse models of DCM via Western blot analysis. FXR1 associates with intercalated discs, and integral gap junction proteins Cx43 (connexin 43), Cx45 (connexin 45), and ZO-1 (zonula occludens-1) were identified as novel mRNA targets of FXR1 by using a BioID proximity assay and RNA immunoprecipitation. Our findings show that FXR1 is a multifunctional protein involved in translational regulation and stabilization of its mRNA targets in heart muscle. In addition, introduction of 3xMyc-FXR1 via adeno-associated virus into mice leads to the redistribution of gap junctions and promotes ventricular tachycardia, showing the functional significance of FXR1 upregulation observed in DCM. CONCLUSIONS: In DCM, increased FXR1 expression appears to play an important role in disease progression by regulating gap junction remodeling. Together this study provides a novel function of FXR1, namely, that it directly regulates major gap junction components, contributing to proper cell-cell communication in the heart.

Epithelial stem cells and implications for wound repair.

Activation of epithelial stem cells and efficient recruitment of their proliferating progeny plays a critical role in cutaneous wound healing. The reepithelialized wound epidermis has a mosaic composition consisting of progeny that can be traced back both to epidermal and several types of hair follicle stem cells. The contribution of hair follicle stem cells to wound epidermis is particularly intriguing as it involves lineage identity change from follicular to epidermal. Studies from our laboratory show that hair follicle-fated bulge stem cells commit only transient amplifying epidermal progeny that participate in the initial wound re-epithelialization, but eventually are outcompeted by other epidermal clones and largely disappear after a few months. Conversely, recently described stem cell populations residing in the isthmus portion of hair follicle contribute long-lasting progeny toward wound epidermis and, arguably, give rise to new interfollicular epidermal stem cells. The role of epithelial stem cells during wound healing is not limited to regenerating stratified epidermis. By studying regenerative response in large cutaneous wounds, our laboratory uncovered that epithelial cells in the center of the wound can acquire greater morphogenetic plasticity and, together with the underlying wound dermis, can engage in an embryonic-like process of hair follicle neogenesis. Future studies should uncover the cellular and signaling basis of this remarkable adult wound regeneration phenomenon.

Deficiency in metabolic regulators PPARγ and PTEN cooperates to drive keratinizing squamous metaplasia in novel models of human tissue regeneration.

Hindgut-derived endoderm can differentiate into rectal, prostatic, and bladder phenotypes. Stromal-epithelial interactions are crucial for this development; however, the precise mechanisms by which epithelium responds to stromal cues remain unknown. We have previously reported ectopic expression of peroxisome proliferator-activated receptor-γ2 (PPARγ2) increased androgen receptor expression and promoted differentiation of mouse prostate epithelium. PPARγ is also implicated in urothelial differentiation. Herein we demonstrate that knockdown of PPARγ2 in benign human prostate epithelial cells (BHPrEs) promotes urothelial transdifferentiation. Furthermore, in vitro and in vivo heterotypic tissue regeneration models with embryonic bladder mesenchyme promoted urothelial differentiation of PPARγ2-deficient BHPrE cells, and deficiency of both PPARγ isoforms 1 and 2 arrested differentiation. Because PTEN deficiency is cooperative in urothelial pathogenesis, we engineered BHPrE cells with combined knockdown of PPARγ and PTEN and performed heterotypic recombination experiments using embryonic bladder mesenchyme. Whereas PTEN deficiency alone induced latent squamous differentiation in BHPrE cells, combined PPARγ and PTEN deficiency accelerated the development of keratinizing squamous metaplasia (KSM). We further confirmed via immunohistochemistry that gene expression changes in metaplastic recombinants reflected human urothelium undergoing KSM. In summary, these data suggest that PPARγ isoform expression provides a molecular basis for observations that adult human epithelium can be transdifferentiated on the basis of heterotypic mesenchymal induction. These data also implicate PPARγ and PTEN inactivation in the development of KSM.

Sex hormone-binding globulin and heart failure hospitalizations in patients with dysglycemia: Experiences from the outcome reduction with an Initial Glargine Intervention trial.

INTRODUCTION: Sex hormone-binding globulin (SHBG), which binds most of circulating testosterone in blood, has been linked to dysglycemia and cardiovascular disease but the relationship with heart failure remains unclear. AIM: To study the relation between SHBG and heart failure hospitalizations. METHODS: SHBG levels were analysed in dysglycemic participants at high cardiovascular risk (n = 8401) followed for a median of 6.2 years in the Outcome Reduction with an Initial Glargine Intervention trial. Cox regression was used to estimate hazard ratios (HRs) per one standard deviation increase for heart failure hospitalizations adjusted for age, comorbidities, biochemical data (including testosterone) and pharmacological treatment. RESULTS: 5553 men and 2848 women were included. Heart failure hospitalizations occurred in 349 (6.3 %) men and 123 (4.3 %) women. One standard deviation increase in SHBG was independently associated with an increased risk of heart failure hospitalizations in men (HR 1.15, 95 % CI 1.03-1.28; p = 0.011) but not in women (HR 1.15; 95 % CI 0.96-1.39; p = 0.14). CONCLUSIONS: In patients with dysglycemia and high cardiovascular risk, increasing SHBG was associated with greater risk of HF hospitalizations independent of testosterone concentrations in men but not in women, suggesting the effects could be mediated through androgen-independent pathways.

Efficacy and Tolerability of Ramucirumab Plus Erlotinib in Taiwanese Patients with Untreated, Epidermal Growth Factor Receptor-Mutated, Stage IV Non-small Cell Lung Cancer in the RELAY Study.

BACKGROUND: In RELAY, a randomized, double-blind, phase III trial investigating the efficacy and safety of ramucirumab+erlotinib (RAM+ERL) or ERL+placebo (PBO) in patients with untreated, stage IV, epidermal growth factor receptor (EGFR)-mutated non-small cell lung cancer (NSCLC), RAM+ERL demonstrated superior progression-free survival (PFS) versus PBO+ERL, with no new safety signals. OBJECTIVE: The aim of this paper was to report efficacy and tolerability findings for the Taiwanese participants of RELAY. PATIENTS AND METHODS: Patients were randomized 1:1 to RAM+ERL or ERL+PBO. Primary endpoint was investigator-assessed PFS. Secondary endpoints included objective response rate (ORR), duration of response (DoR) and tolerability. Data for the current analysis are reported descriptively. RESULTS: In RELAY, 56 Taiwanese patients were enrolled; 26 received RAM+ERL, 30 received ERL+PBO. The demographic profile of the Taiwanese subgroup was consistent with that of the overall RELAY population. Median PFS for RAM+ERL/ERL+PBO, respectively, was 22.05 months/13.40 months (unstratified hazard ratio 0.4; 95% confidence interval 0.2-0.9); ORR was 92%/60%; median DoR was 18.2 months/12.7 months. All patients experienced one or more treatment-emergent adverse events (TEAEs); those most commonly reported were diarrhea and dermatitis acneiform (58% each) for RAM+ERL and diarrhea (70%) and paronychia (63%) for PBO+ERL. Grade ≥  3 TEAEs were experienced by 62%/30% of RAM+ERL/PBO+ERL patients, respectively, and included dermatitis acneiform (19%/7%), hypertension (12%/7%), and pneumonia (12%/0%). CONCLUSIONS: PFS for the Taiwanese participants of RELAY receiving RAM+ERL versus ERL+PBO was consistent with that in the overall RELAY population. These results, together with no new safety signals and a manageable safety profile, may support first-line use of RAM+ERL in Taiwanese patients with untreated EGFR-mutant stage IV NSCLC. TRIAL REGISTRATION: www. CLINICALTRIALS: gov , NCT02411448.

SOdium-glucose CO-transporter inhibition in patients with newly detected Glucose Abnormalities and a recent Myocardial Infarction (SOCOGAMI).

AIMS/HYPOTHESIS: Established dysglycaemia (impaired glucose tolerance [IGT] or type 2 diabetes [T2DM]) is a risk factor for further cardiovascular events in patients with coronary artery disease. Sodium-glucose cotransporter 2 inhibitors reduce this risk. The aim of the present investigation was to test the hypothesis that empagliflozin exerts beneficial effects on myocardial function in patients with a recent acute coronary syndrome and newly detected dysglycaemia. METHODS: Forty-two patients (mean age 67.5 years, 81 % male) with recent myocardial infarction (n = 36) or unstable angina (n = 6) and newly detected IGT (n = 27) or T2DM (n = 15) were randomised to 25 mg of empagliflozin daily (n = 20) or placebo (n = 22) on top of ongoing therapy. They were investigated with oral glucose tolerance tests, stress-perfusion cardiac magnetic resonance imaging (CMR) and echocardiography at three occasions: before randomisation, after seven months on study drug and three months following cessation of such drug. Primary outcome was a change in left ventricular (LV) end-diastolic volume (LVEDV) and secondary outcomes were a change in a) systolic and diastolic LV function; b) coronary flow reserve; c) myocardial extracellular volume (ECV) in non-infarcted myocardium; d) aortic pulse wave velocity. RESULTS: Empagliflozin induced a significant decrease in fasting and post load glucose (p < 0.05) and body weight (p < 0.01). Empagliflozin did not influence LVEDV, LV systolic or mass indexes, coronary flow reserve, ECV or aortic pulse wave velocity. Echocardiographic indices of LV diastolic function (E/e' and mitral E/A ratio) were not influenced. No safety concerns were identified. CONCLUSIONS/INTERPRETATION: Empagliflozin had predicted effects on the dysglycaemia but did not influence variables expressing LV function, coronary flow reserve and ECV. An explanation may be that the LV function of the patients was within the normal range.

Hysteroscopic transcervical resection. A straightforward method corrects bleeding related to cesarean section scar defects.

Cesarean section defect had been found to be one of the causes of prolonged bleeding in women with previous cesarean delivery. Lin's hysteroscopy (TCR)-metroplasty method had focused on 4 predisposed anatomical defects, which ensured correction of the cesarean section defect. With this simple procedure, the patients had greatly improved their quality of life, as well as discomfort.

Androgen receptor polymorphism, testosterone levels, and prognosis in patients with acute myocardial infarction.

Aims: Low testosterone has been associated with cardiovascular disease in men but with contradictory findings. Testosterone bind to the androgen receptor and polymorphisms of the receptor gene such as CAG repeat length may affect transcriptional activity, possibly mitigating testosterone effects. The aims were to study the CAG repeat length and testosterone levels at four time points following a myocardial infarction (MI) and to analyse possible relationships between CAG repeat length and cardiovascular prognosis. Methods and results: Male patients admitted for acute MI (n = 122) from the Glucose in Acute Myocardial Infarction study were included. Blood samples were drawn at four time points (day after admission, at discharge, and at 3 and 12 months post-infarction) for assessment of testosterone levels. Patients were followed for a median of 11.6 years. Cox regression analyses were performed for CAG repeat length by one unit increment and by > vs. ≤median for cardiovascular events and all-cause mortality. Median CAG repeat length was 20. There was no difference in testosterone levels at each time point when dividing the cohort into ≤ vs. >CAG repeat median (=20). There was no association between CAG repeat length either as a continuous or categorical variable in unadjusted and age-adjusted Cox analyses for cardiovascular events. While CAG >20 was associated with all-cause mortality in unadjusted analyses (hazard ratio 2.19, 95% confidence interval 1.13-4.22; P = 0.02), it did not remain significant following adjustment for age. Conclusion: CAG repeat length was not associated with testosterone levels or prognosis in men with acute MI.

Testosterone and sex hormone-binding globulin in dysglycemic women at high cardiovascular risk: A report from the Outcome Reduction with an Initial Glargine Intervention trial.

AIMS: Total and free testosterone and sex hormone-binding globulin may affect cardiovascular prognosis in women. The objective was to study the association between sex hormones and prognosis in women with dysglycemia and high cardiovascular risk. METHODS: This epidemiological report included dysglycemic women from the Outcome Reduction with an Initial Glargine Intervention trial (n = 2848) with baseline total testosterone and sex hormone-binding globulin. Free testosterone was calculated with the Vermeulen formula. Cox regression analyses adjusted for variables including age, previous diseases and pharmacological treatments were used to estimate the association between these levels and the composite cardiovascular outcome (death from cardiovascular causes, nonfatal myocardial infarction or nonfatal stroke) and all-cause mortality per one standard deviation. RESULTS: Patients (73% post-menopausal) were followed for a median of 6.1 years during which 377 cardiovascular events and 389 deaths occurred. In Cox analyses, total and free testosterone were not associated with any outcomes, but sex hormone-binding globulin was related to all-cause mortality in age adjusted (HR 1.15; 95% CI 1.06-1.24; p < 0.01) and fully adjusted analyses (HR 1.14; 95% CI 1.05-1.24; p < 0.01). CONCLUSIONS: Increasing levels of baseline sex hormone-binding globulin were associated with an increased risk of all-cause mortality in dysglycemic women at high cardiovascular risk. TRIAL REGISTRATION: ClinicalTrials.gov no. NCT00069784.

Testosterone, sex hormone-binding globulin and risk of cardiovascular events: A report from the Outcome Reduction with an Initial Glargine Intervention trial.

AIMS: Testosterone and its binding protein sex hormone-binding globulin have been associated with cardiovascular disease and dysglycaemia. However, information on the prognostic implication in patients at high cardiovascular risk with dysglycaemia is inconsistent. The study objective was to determine whether testosterone and/or sex hormone-binding globulin predict cardiovascular events or death in dysglycaemic patients. METHODS: Dysglycaemic males at high cardiovascular risk ( n = 5553) who participated in the Outcome Reduction with an Initial Glargine Intervention (ORIGIN) trial and provided baseline blood samples were studied. Testosterone and sex hormone-binding globulin were measured at baseline and used to estimate free testosterone. Low levels of total and free testosterone were defined as ≤300 ng/dl and ≤7 ng/dl, respectively. Patients were followed for six years for cardiovascular events (defined as the composite of cardiovascular death, non-fatal myocardial infarction or stroke) and all-cause mortality. RESULTS: The mean total and free testosterone levels were 416.6 ng/dl and 8.4 ng/dl, and low levels were present in 13% and 37% of the patients. The median sex hormone-binding globulin level was 35 nmol/l. In Cox regression models adjusted for age, previous diseases and pharmacological treatment, neither total nor free testosterone predicted cardiovascular events. However, a one-standard-deviation increase in sex hormone-binding globulin predicted both cardiovascular events (hazard ratio 1.07; 95% confidence interval 1.00-1.14; p = 0.03) and all-cause mortality (hazard ratio 1.13; 95% confidence interval 1.06-1.21; p < 0.01). CONCLUSION: Sex hormone-binding globulin, but not total testosterone, predicts cardiovascular disease and all-cause mortality in dysglycaemic males at high cardiovascular risk.

Urinary Bone Turnover Markers as Target Indicators for Monitoring Bisphosphonate Drug Treatment in the Management of Osteoporosis.

BACKGROUND: Osteoporosis is a debilitating disease characterized by bone micro-architecture degradation contributing to fragility fractures. Currently, determining bone mineral density (BMD) via dual-energy X-ray absorptiometry (DXA) is the most reliable form of diagnosing osteoporosis and managing pharmacological treatment regimens. However, changes in BMD occur slowly (i.e., several months) and DXA does not reflect the metabolic rate of bone turnover. Alternatively, biochemical bone turnover markers are metabolic indicators released into serum and/or urine, and their quantity reflects the metabolic activity of bone. bone turnover markers show a rapid response following antiresorptive drug administration, and enzyme-linked immunosorbent assays (ELISA) have been established and used in clinical trials to help assess and predict fracture risk independent of BMD. OBJECTIVE: This review highlights various established bone turnover markers that have found utility in the clinic as reliable and standardized indicators of bone turnover, with attention to those used to assess efficacy of bisphosphonate drug therapy - particularly in monitoring medication adherence in patients with postmenopausal osteoporosis. RESULTS: We posit that the use of urinary bone turnover markers values determined by immunoassay or ELISA at routine clinic visits might serve as valuable feedback to healthcare professionals and patients to help monitor the efficacy and adherence of bisphosphonate therapy and disease progression. CONCLUSION: Our belief is that when assessed in combination with an algorithm of independent risk factors, measuring urinary bone turnover markers using a point of care kit may find utility in the osteoporosis clinic as an accessible, non-invasive and cost-effective alternative for the routine assessment of efficacy of bisphosphonate therapies.

Dynamics of testosterone levels in patients with newly detected glucose abnormalities and acute myocardial infarction.

OBJECTIVE: Low testosterone has been associated with increased cardiovascular risk and glucose abnormalities. This study explored the prevalence of low testosterone, dynamics over time and prognostic implications in acute myocardial infarction patients with or without glucose abnormalities. METHODS: Male acute myocardial infarction patients (n = 123) and healthy controls (n = 124) were categorised as having normal or abnormal glucose tolerance (impaired glucose tolerance or diabetes) by oral glucose tolerance testing. Testosterone was measured at hospital admission, discharge, 3 and 12 months thereafter in patients. Patients and controls were followed for 11 years for major cardiovascular events (cardiovascular death/acute myocardial infarction/stroke/severe heart failure). RESULTS: At hospital admission, more patients had low testosterone (⩽300 ng/dl) and lower median levels than controls (64 vs 28%; p < 0.001 and 243 vs 380 ng/dl; p < 0.01). At the subsequent time points, testosterone had increased to 311, 345 and 357 ng/dl. Patients with abnormal glucose tolerance had the highest prevalence (75%) of low levels. In adjusted Cox regression models, neither total nor free testosterone predicted major cardiovascular events. CONCLUSION: Low testosterone levels were common in male acute myocardial infarction patients in the acute phase, especially in the presence of abnormal glucose tolerance, but increased over time indicating that testosterone measured in close proximity to acute myocardial infarction should be interpreted with caution.

Regeneration of fat cells from myofibroblasts during wound healing.

Although regeneration through the reprogramming of one cell lineage to another occurs in fish and amphibians, it has not been observed in mammals. We discovered in the mouse that during wound healing, adipocytes regenerate from myofibroblasts, a cell type thought to be differentiated and nonadipogenic. Myofibroblast reprogramming required neogenic hair follicles, which triggered bone morphogenetic protein (BMP) signaling and then activation of adipocyte transcription factors expressed during development. Overexpression of the BMP antagonist Noggin in hair follicles or deletion of the BMP receptor in myofibroblasts prevented adipocyte formation. Adipocytes formed from human keloid fibroblasts either when treated with BMP or when placed with human hair follicles in vitro. Thus, we identify the myofibroblast as a plastic cell type that may be manipulated to treat scars in humans.

Electrographic seizures during therapeutic hypothermia for neonatal hypoxic-ischemic encephalopathy.

Electrographic seizures are common in neonates with hypoxic-ischemic encephalopathy, but detailed data are not available regarding seizure incidence during therapeutic hypothermia. The objective of this prospective study was to determine the incidence and timing of electrographic seizures in term neonates undergoing whole-body therapeutic hypothermia for hypoxic-ischemic encephalopathy as detected by conventional full-array electroencephalography for 72 hours of therapeutic hypothermia and 24 hours of normothermia. Clinical and electroencephalography data were collected from 26 consecutive neonates. Electroencephalograms were reviewed by 2 pediatric neurophysiologists. Electrographic seizures occurred in 17 of 26 (65%) patients. Seizures were entirely nonconvulsive in 8 of 17 (47%), status epilepticus occurred in 4 of 17 (23%), and seizure onset was in the first 48 hours in 13 of 17 (76%) patients. Electrographic seizures were common, were often nonconvulsive, and had onset over a broad range of times in the first days of life.

Gold nanoparticle-assisted protein enrichment and electroelution for biological samples containing low protein concentration--a prelude of gel electrophoresis.

Protein enrichment is essential for biological samples that contain low protein concentrations, especially for proteomic studies that require sufficient quantities for subsequent MS analysis. Traditional precipitation methods, however, are limited in the sample volume and protein concentration required to cause efficient precipitations. We showed that gold nanoparticles (Au-NPs) can be easily applied to concentrate proteins from more than 15 mL of human urine, in which the total protein concentration is less than 1.4 ppm. Moreover, Au-NP-aggregated proteins can be directly applied to gel electrophoresis for Au-NP-protein dissociation followed by free protein separation as well as for the subsequent in-gel digestion and protein identification by mass spectrometry. We compared this method with trichloroacetic acid (TCA) precipitation method, one of the most common precipitation methods, and TCA method showed no enrichment effect for protein samples with large volumes (>2 mL) or with low protein concentrations (4 ppm). Therefore, Au-NP aggregation is not only a simple and efficient method for enriching a broad range of proteins, it is also particularly useful for concentrating proteins from a relatively large volume of dilute biological fluids, under which TCA method is ineffective.