Circulating androgen regulation by androgen-catabolizing gut bacteria in male mouse gut.

Abnormally high circulating androgen levels have been considered a causative factor for benign prostatic hypertrophy and prostate cancer in men. Recent animal studies on gut microbiome suggested that gut bacteria are involved in sex steroid metabolism; however, the underlying mechanisms and bacterial taxa remain elusive. Denitrifying betaproteobacteria Thauera spp. are metabolically versatile and often distributed in the animal gut. Thauera sp. strain GDN1 is an unusual betaproteobacterium capable of catabolizing androgen under both aerobic and anaerobic conditions. We administered C57BL/6 mice (aged 7 weeks) with strain GDN1 through oral gavage. The strain GDN1 administration caused a minor increase in the relative abundance of Thauera (≤0.1%); however, it has profound effects on the host physiology and gut bacterial community. The results of our ELISA assay and metabolite profile analysis indicated an approximately 50% reduction in serum androgen levels in the strain GDN1-administered male mice. Moreover, androgenic ring-cleaved metabolites were detected in the fecal extracts of the strain GDN1-administered mice. Furthermore, our RT - qPCR results revealed the expression of the androgen catabolism genes in the gut of the strain GDN1-administered mice. We found that the administered strain GDN1 regulated mouse serum androgen levels, possibly because it blocked androgen recycling through enterohepatic circulation. This study discovered that sex steroids serve as a carbon source of gut bacteria; moreover, host circulating androgen levels may be regulated by androgen-catabolizing gut bacteria. Our data thus indicate the possible applicability of androgen-catabolic gut bacteria as potent probiotics in alternative therapy of hyperandrogenism.

Omics and mechanistic insights into di-(2-ethylhexyl) phthalate degradation in the O2-fluctuating estuarine sediments.

Di-(2-ethylhexyl) phthalate (DEHP) represents the most used phthalate plasticizer with an annual production above the millions of tons worldwide. Due to its inadequate disposal, outstanding chemical stability, and extremely low solubility (3 mg/L), endocrine-disrupting DEHP often accumulates in urban estuarine sediments at concentrations above the predicted no-effect concentration (20-100 mg/kg). Our previous study suggested that microbial DEHP degradation in estuarine sediments proceeds synergistically where DEHP side-chain hydrolysis to form phthalic acid represents a bottleneck. Here, we resolved this bottleneck and deconstructed the microbial synergy in O2-fluctuating estuarine sediments. Metagenomic analysis and RNA sequencing suggested that orthologous genes encoding extracellular DEHP hydrolase NCU65476 in Acidovorax sp. strain 210-6 are often flanked by the co-expressed composite transposon and are widespread in aquatic environments worldwide. Therefore, we developed a turbidity-based microplate assay to characterize NCU65476. The optimized assay conditions (with 1 mM Ca2+ and pH 6.0) increased the DEHP hydrolysis rate by a factor of 10. Next, we isolated phthalic acid-degrading Hydrogenophaga spp. and Thauera chlorobenzoica from Guandu estuarine sediment to study the effect of O2(aq) on their metabolic synergy with strain 210-6. The results of co-culture experiments suggested that after DEHP side-chain hydrolysis by strain 210-6, phthalic acid can be degraded by Hydrogenophaga sp. when O2(aq) is above 1 mg/L or degraded by Thauera chlorobenzoica anaerobically. Altogether, our data demonstrates that DEHP could be degraded synergistically in estuarine sediments via divergent pathways responding to O2 availability. The optimized conditions for NCU65476 could facilitate the practice of DEHP bioremediation in estuarine sediments.

Integrated Multi-omics Investigations Reveal the Key Role of Synergistic Microbial Networks in Removing Plasticizer Di-(2-Ethylhexyl) Phthalate from Estuarine Sediments.

Di-(2-ethylhexyl) phthalate (DEHP) is the most widely used plasticizer worldwide, with an annual global production of more than 8 million tons. Because of its improper disposal, endocrine-disrupting DEHP often accumulates in estuarine sediments in industrialized countries at submillimolar levels, resulting in adverse effects on both ecosystems and human beings. The microbial degraders and biodegradation pathways of DEHP in O2-limited estuarine sediments remain elusive. Here, we employed an integrated meta-omics approach to identify the DEHP degradation pathway and major degraders in this ecosystem. Estuarine sediments were treated with DEHP or its derived metabolites, o-phthalic acid and benzoic acid. The rate of DEHP degradation in denitrifying mesocosms was two times slower than that of o-phthalic acid, suggesting that side chain hydrolysis of DEHP is the rate-limiting step of anaerobic DEHP degradation. On the basis of microbial community structures, functional gene expression, and metabolite profile analysis, we proposed that DEHP biodegradation in estuarine sediments is mainly achieved through synergistic networks between denitrifying proteobacteria. Acidovorax and Sedimenticola are the major degraders of DEHP side chains; the resulting o-phthalic acid is mainly degraded by Aestuariibacter through the UbiD-dependent benzoyl coenzyme A (benzoyl-CoA) pathway. We isolated and characterized Acidovorax sp. strain 210-6 and its extracellular hydrolase, which hydrolyzes both alkyl side chains of DEHP. Interestingly, genes encoding DEHP/mono-(2-ethylhexyl) phthalate (MEHP) hydrolase and phthaloyl-CoA decarboxylase-key enzymes for side chain hydrolysis and o-phthalic acid degradation, respectively-are flanked by transposases in these proteobacterial genomes, indicating that DEHP degradation capacity is likely transferred horizontally in microbial communities. IMPORTANCE Xenobiotic phthalate esters (PAEs) have been produced on a considerably large scale for only 70 years. The occurrence of endocrine-disrupting di-(2-ethylhexyl) phthalate (DEHP) in environments has raised public concern, and estuarine sediments are major DEHP reservoirs. Our multi-omics analyses indicated that complete DEHP degradation in O2-limited estuarine sediments depends on synergistic microbial networks between diverse denitrifying proteobacteria and uncultured candidates. Our data also suggested that the side chain hydrolysis of DEHP, rather than o-phthalic acid activation, is the rate-limiting step in DEHP biodegradation within O2-limited estuarine sediments. Therefore, deciphering the bacterial ecophysiology and related biochemical mechanisms can help facilitate the practice of bioremediation in O2-limited environments. Furthermore, the DEHP hydrolase genes of active DEHP degraders can be used as molecular markers to monitor environmental DEHP degradation. Finally, future studies on the directed evolution of identified DEHP/mono-(2-ethylhexyl) phthalate (MEHP) hydrolase would bring a more catalytically efficient DEHP/MEHP hydrolase into practice.

Efficient hexabromocyclododecane-biodegrading microorganisms isolated in Taiwan.

The potential toxicity of hexabromocyclododecane (HBCD), its persistence in the environment, and its high bioaccumulation characteristics pose a need to remediate HBCD in the environment. Bacillus cereus and B. subtilis species complexes we isolated from Taiwan soil are capable of degrading HBCD. B. cereus can degrade HBCD with a half-life only 0.911 days. The highest efficiency of HBCD degradation by B. cereus was achieved at pH 7.0, 35 °C, and 0.10 ppm HBCD. The removal mechanism of HBCD by B. cereus is debromination and its pathway was proposed. The addition of surfactant Tween 60 improved HBCD removal but the addition of CaO2, slow-releasing oxygen, did not. These findings can facilitate the bioremediation of HBCD in the environment.

Comparison of Percutaneous Transluminal Angioplasty with Stenting for Treatment of Central Venous Stenosis or Occlusion in Hemodialysis Patients: A Systematic Review and Meta-analysis.

OBJECTIVES: A common cause of hemodialysis vascular access dysfunction, a primary cause of morbidity and mortality in patients undergoing hemodialysis, is central venous stenosis or occlusion. The present study compared percutaneous transluminal angioplasty (PTA) with percutaneous transluminal angioplasty and stent placement (PTS) for treatment of central venous stenosis or central vein occlusion in hemodialysis patients. METHODS: A systematic literature review was conducted using database searches of PubMed, Cochrane, and Embase. Articles were selected using the Population/Intervention/Comparator/Outcomes (PICO) process. Outcomes included the rate of procedural success, primary patency, assisted primary patency, re-intervention subjects, re-intervention rate, and adverse events. RESULTS: A total of eight studies were included in the meta-analysis with subjects in the PTA group (n = 281) or PTS group (n = 192). Primary patency rate between PTA and PTS groups at 3-, 6-, 12-, or 24-month follow-up was not different (all p > 0.05). Patients treated with PTA had greater assisted primary patency rates than the PTS group (OR = 1.03, 1.73, 1.03, and 2.00 at 3, 6, 12, and 24 months, respectively). However, the statistical assessment only showed significantly at 24-month follow-up (p = 0.01). CONCLUSIONS: The present meta-analysis revealed that, compared to PTS, PTA may provide increased assisted primary patency for endovascular treatment of central vein stenosis or occlusion in patients undergoing hemodialysis. LEVEL OF EVIDENCE: Level 3a.

Retroconversion of estrogens into androgens by bacteria via a cobalamin-mediated methylation.

Steroid estrogens modulate physiology and development of vertebrates. Conversion of C19 androgens into C18 estrogens is thought to be an irreversible reaction. Here, we report a denitrifying Denitratisoma sp. strain DHT3 capable of catabolizing estrogens or androgens anaerobically. Strain DHT3 genome contains a polycistronic gene cluster, emtABCD, differentially transcribed under estrogen-fed conditions and predicted to encode a cobalamin-dependent methyltransferase system conserved among estrogen-utilizing anaerobes; an emtA-disrupted DHT3 derivative could catabolize androgens but not estrogens. These data, along with the observed androgen production in estrogen-fed strain DHT3 cultures, suggested the occurrence of a cobalamin-dependent estrogen methylation to form androgens. Consistently, the estrogen conversion into androgens in strain DHT3 cell extracts requires methylcobalamin and is inhibited by propyl iodide, a specific inhibitor of cobalamin-dependent enzymes. The identification of the cobalamin-dependent estrogen methylation thus represents an unprecedented metabolic link between cobalamin and steroid metabolism and suggests that retroconversion of estrogens into androgens occurs in the biosphere.

Temporal and spatial surveys of polybromodiphenyl ethers (PBDEs) contamination of soil near a factory using PBDEs in northern Taiwan.

Polybrominated diphenyl ethers (PBDEs), previously commonly used as flame retardants, should be monitored in the environment since some are listed as persistent organic pollutants. A contaminated site near a northern Taiwan factory using decabrominated diphenyl ether (deca-BDE) was identified based on a vegetable PBDEs monitoring project in 2013. The subsequent spatial and temporal survey of that contaminated site shows the contamination ingredients in soils were close to ones used by the factory, indicating that contamination was from the factory, possibly through an exhaust vent. The average concentration of deca-BDE in the main contaminated soil was 615 µg/kg d. w. (dry weight) soil in 2015, slightly decreasing to 604 µg/kg d. w. soil in 2016, increasing to 844 µg/kg d. w. soil in 2017, and then slightly decreasing to 670 µg/kg d. w. soil in 2018. The slight change of deca-BDE and the minor change in low brominated congener level indicate a low degradation rate. The contamination of peripheral sites was around 5000 µg/kg d. w. soil for one PBDEs sampling site that was higher than those around or within the main contaminated farm, indicating serious pollution. Concentrations of PBDEs in different soil depths show that depth 2-15 cm accounted for the greatest PBDEs accumulation, indicating that deca-BDE pollution had been present over time and transported into deeper soil. There can be PBDEs uptake by crops consumed by humans, as shown in our previous studies, so continuous monitoring of PBDEs in this site is important and treatments should be established urgently.

Outcomes of Endovascular Therapy With the Controlled Antegrade Retrograde Subintimal Tracking (CART) or Reverse CART Technique for Long Infrainguinal Occlusions.

PURPOSE: To compare the safety, efficacy, and clinical outcomes associated with the controlled antegrade retrograde subintimal tracking (CART) or reverse CART (r-CART) technique to the conventional retrograde approach in the treatment of patients with long infrainguinal occlusions. METHODS: From May 2008 to April 2014, 121 patients failed antegrade recanalization and underwent a retrograde approach to recanalize long infrainguinal occlusions. Patients who underwent successful endovascular therapy (EVT) by the conventional retrograde approach (CRA group) were compared to patients who had successful EVT using the CART/r-CART technique (CART group) after failure of a bidirectional approach. The efficacy, safety, vessel patency, and other clinical outcomes were compared between the groups. RESULTS: Fifty-eight patients (mean age 71.6 ± 12.2 years; 32 men) underwent successful EVT (47.9%, 58/121) using the conventional retrograde approach (CRA group), while 44 patients (mean age 70.8 ± 11.1 years; 31 men) among the 50 patients who underwent the CART/r-CART technique were successfully treated (88.0%, 44/50). Both groups had similar average occlusion lengths and gained 100% immediate hemodynamic success after EVT. There was no significant difference between the groups regarding procedure-related complications. During follow-up, 28 patients died (p=0.380), but there were no differences in the rates of major (p=0.279) or minor amputation (p=0.417) between the groups. There was no difference in the 2-year primary patency (31% vs 24%, p=0.686), assisted primary patency (66% vs 76%, p=0.251), target vessel revascularization (65% vs 54%, p=0.845), or sustained clinical success (52% vs 46%, p=0.995) rates between the CRA and CART groups, respectively. CONCLUSION: Based on acceptable safety, efficacy, and follow-up results in this study, the CART/r-CART technique can salvage patients with long peripheral occlusions after failure of the conventional antegrade or retrograde approach.

QT interval Independently Predicts Mortality and Heart Failure in Patients with ST-Elevation Myocardial Infarction.

OBJECTIVES: Heart-rate corrected QT (QTc) interval predicts cardiovascular mortality or all-cause mortality in the general population. Little is known about the best cut-off value of QTc interval for predicting clinical events in patients with ST-elevation myocardial infarction (STEMI). METHODS: We enrolled 264 patients with STEMI who received measurement of QTc intervals at ER (QTc-ER), on day 2 (QTc-D2), and on day 3 (QTc-D3) of hospitalization. Clinical events, including all-cause death and readmission for heart failure, were followed for 2 years. RESULTS: Prolonged QTc-ER, but not QTc-D2 or QTc-D3, well predicted clinical events with the best cut-off value of 445 ms. Patient with QTc-ER > 445 ms had lower left ventricular ejection fraction at baseline and at 6 months. Kaplan-Meier survival curves showed that the combination of QTc-ER > 445 ms and N-terminal pro-brain natriuretic peptide (NT-pro BNP) > 936 pg/mL was a strong predictor of clinical events (p<0.001). In multivariable Cox regression analysis, the independent predictors of death and heart failure were QTc-ER (p<0.001), log NT-proBNP (p<0.001), diabetes mellitus (p<0.001), history of stroke (p=0.001), and left ventricular end diastolic volume index (p<0.001). CONCLUSION: QTc-ER > 445 ms independently predicts clinical events in STEMI, providing incremental prognostic value to established clinical predictors and NT-proBNP.

Feasibility and Clinical Outcomes of Peripheral Drug-Coated Balloon in High-Risk Patients with Femoropopliteal Disease.

BACKGROUND: Clinical outcomes of the drug-coated balloon (DCB) procedure in high-risk patients with femoropopliteal (FP) disease have not been investigated sufficiently. METHODS: This retrospective, single-center study analyzed 87 patients (39% dialysis) and 97 affected legs (64% critical limb ischemia [CLI]) that underwent DCB for symptomatic FP disease from March 2013 to September 2014. Risk stratification was based on FeDCLIP (female, diabetes, dialysis, CLI, lesion length >150 mm and poor runoff) score. The DCB outcomes among the different risk groups were compared and factors predicting restenosis were analyzed during follow-up. RESULTS: Most of study participants (84%) were moderate to high-risk patients. The procedural success rate was 100% and the 30-day major adverse vascular event rate was 2.1%. The mean lesion length was 178 ± 106 mm and the mean follow-up time was 428 ± 145 (range 50-782) days. The binary restenosis-free and clinically driven target lesion revascularization (CD-TLR)-free rates at 12 months were 77.5% and 84.3%, respectively, for all participants. No significant differences were observed in 1-year binary restenosis and CD-TLR rates in the low-, moderate-, and high-risk groups (60%, 84%, and 73%: p = 0.396; 78%, 89%, and 80%: p = 0.635, respectively). In multivariate analysis, lesion length >150 mm (Hazard ratio [HR]: 8.00, 95% confidence interval (CI) 1.12 to 55.6, p = 0.038) and Rutherford class 6 (HR: 7.09, 95% CI, 1.15 to 43.5, p = 0.034) were identified as independent predictors of binary restenosis. CONCLUSIONS: Despite general comorbidities and advanced limb ischemia, 1-year outcomes of DCB in high-risk patients with FP disease were effective. The DCB procedure holds promise to improve vessel patency; however, lesion length >150 mm and major tissue loss were independent predictors for binary restenosis after the treatment.

Endovascular intervention in Taiwanese patients with critical limb ischemia: patient outcomes in 333 consecutive limb procedures with a 3-year follow-up.

BACKGROUND/PURPOSE: Midterm outcomes of endovascular intervention (EVI) for critical limb ischemia (CLI) have not been previously reported in Taiwan. This study assessed the safety, feasibility, and patient-oriented outcomes for CLI patients after EVI. METHODS: From June 2005 to December 2011, 270 patients underwent EVI for CLI of 333 limbs. Primary patency (PP), assisted primary patency (AP), limb salvage, sustained clinical success (SCS), secondary SCS (SSCS), and survival were assessed using Kaplan-Meier analysis. RESULTS: The procedural success rate was 89%, and the periprocedural mortality and major complication rates within 30 days were 0.6% and 6.9%, respectively. During the mean follow-up time of 27 ± 20 months (1-77), 64 patients died and 25 legs required major amputation. Eighty-one percent of the patients with tissue loss had wound healing at 6 months and 75% of the patients were ambulatory, with or without assisting devices, at 1 year. The overall survival and limb salvage rates at 3 years were 70% and 90%, respectively. The PP and AP at 1 and 3 years were 58% and 37% and 79% and 61%, respectively. The SCS and SSCS were 65% and 46% and 80% and 64% at 1 and 3 years, respectively. CONCLUSION: In Taiwan, EVI was a safe and feasible procedure for CLI patients, with a high procedural success rate and lower complication rate. Sustained limb salvage and clinical success can be afforded with an active surveillance program and prompt intervention during midterm follow-up.

Growth-differentiation factor-15 and major cardiac events.

BACKGROUND: Growth-differentiation factor (GDF)-15 is a strong predictor of cardiovascular events in patients with ST-elevation myocardial infarction (STEMI). However, the effects of GDF-15 on left ventricular (LV) remodeling have not been clearly elucidated. The aim of this study is to investigate whether GDF-15 will be of benefit in predicting LV remodeling, heart failure and death in patients with STEMI. METHODS: The authors enrolled 216 patients with STEMI who received measurement of GDF-15 level on day 2 of hospitalization. Echocardiographic studies were performed at baseline and were repeated 6 months later. Clinical events, including all-cause death and readmission for heart failure, were followed up for a maximum of 3 years. RESULTS: Patients with GDF-15 levels above the median had lower LV ejection fraction at baseline (43.9% versus 48.0%, P = 0.041) and at 6 months (51.5% versus 56.9%, P = 0.025). In univariable regression model, log-transformed GDF-15 level was not a predictor of increase in LV end-diastolic volume index at 6 months (P = 0.767). Kaplan-Meier survival curves showed that the combination of high GDF-15 and high N-terminal pro-B-type natriuretic peptide was a strong predictor of death and heart failure (P < 0.001). In multivariable Cox regression model, the independent predictors of death and heart failure were age, GDF-15 level and diabetes mellitus. CONCLUSIONS: High GDF-15 level is a strong predictor of death and heart failure in patients with STEMI. Although patients with higher GDF-15 levels tend to have lower LV ejection fraction, they have similar degree of the increase in LV end-diastolic volume index at 6 months.

Clinical predictors of long-term outcomes in patients with critical limb ischemia who have undergone endovascular therapy.

Clinical predictors of long-term outcomes in patients with critical limb ischemia (CLI) treated with endovascular therapy (EVT) remain unclear. In this study, clinical predictors of long-term outcomes in EVT-treated patients with CLI were investigated. In this prospective, observational study, we analyzed a total of 253 Taiwanese patients with CLI with 314 limbs who underwent EVT between 2005 and 2012. Cox models were used to estimate hazard ratios of death, limb loss, and sustained clinical success (SCS). Multivariate analysis showed that age, atrial fibrillation (AF), end-stage renal disease (ESRD), and albumin were significant predictors of mortality. Patients with coronary artery disease and low albumin levels had a significant risk of major limb amputation, while AF, ESRD, and albumin were significant, independent predictors of SCS. In addition to previously reported predictors, we showed that AF and malnutrition can be used to predict long-term outcome in EVT-treated patients with CLI.

Involvement of OpcR, a GbsR-type transcriptional regulator, in negative regulation of two evolutionarily closely related choline uptake genes in Bacillus subtilis.

The osmoprotectant glycine betaine can be generated intracellularly from conversion of the exogenous precursor choline by enzymes encoded by the gbsAB operon in Bacillus subtilis. Uptake of choline from outside B. subtilis cells is mediated through two evolutionarily closely related ATP-binding cassette transporters, OpuB and OpuC. Expression of the opuB operon and of the opuC operon is known to be osmoinducible. Here, we show that choline exerts a suppressive effect on opuC expression during normal growth and under osmotic stress. In the absence of the choline-responsive repressor GbsR, opuB expression is also suppressed by choline. We also report that a gene (formerly yvbF, now designated opcR) located immediately upstream of the opuC operon negatively regulates transcription of the opuC operon and, in the absence of GbsR, also that of the opuB operon. An inverted repeat (TTGTAAA-N8-TTTACAA) that overlaps with the -35 hexamer of the promoters of both operons has been identified as the OpcR operator. OpcR belongs to the GbsR-type transcriptional regulators. Its orthologues with unknown function are present in some other Bacillus species. Moreover, deletion analyses revealed that a region located further upstream of the promoters of the opuB operon and the opuC operon is critical for expression of both operons during normal growth and under osmotic stress. Osmotic induction of these two operons appears not to be OpcR mediated. OpcR is not a choline-responsive repressor. The possible biological role of OpcR is discussed.

Comparison of immediate and 2-year outcomes between excimer laser-assisted angioplasty with spot stent and primary stenting in intermediate to long femoropopliteal disease.

BACKGROUND: To compare the clinical outcomes between excimer laser-assisted angioplasty (ELA) with spot stent (group A) and primary stenting (group B) in intermediate to long femoropopliteal disease. METHODS: Outcomes of 105 patients totaling 119 legs treated with two different strategies were analyzed retrospectively in a prospectively maintained database. RESULTS: Baseline characteristics were similar in both groups. Better angiographic results and lesser increase of serum C-reactive protein levels (0.60 ± 0.72 versus 2.98 ± 0.97 mg/dL, P < 0.001) after the intervention were obtained in Group B. Group A had inferior 1-year outcomes due to higher rate of binary restenosis (67% versus 32%, P = 0.001) and lower rate of primary patency (40% versus 58%, P = 0.039). Rates of amputation-free survival, target vessel revascularization, assisted primary patency, and stent fracture at 24 months were similar in both groups (80% versus 82%, P = 0.979, 65% versus 45%, P = 0.11, 78% versus 80%, P = 0.75 and 6.3% versus 6.8%, P = 0.71, resp.). CONCLUSION: Greater vascular inflammation after ELA with spot stent resulted in earlier restenosis and inferior 1-year clinical outcomes than primary stenting. This benefit was lost in the primary stenting group at 2 years due to late catch-up restenosis. Active surveillance with prompt intervention was required to maintain the vessel patency.

Endovascular sharp recanalization for calcified femoropopliteal artery occlusion.

Endovascular intervention of peripheral chronic total occlusion (CTO) is technically challenging and time consuming. Various techniques and devices are used to facilitate lesion crossing and improve the success rate of the procedure. However, these new devices are quite expensive and not readily available. We report 2 cases of peripheral CTO wherein the occlusions were successfully crossed by using stiff end of Terumo glidewire. This sharp recanalization may be a useful technique for the recanalization of calcified peripheral CTOs when conventional techniques fail and new devices are not readily available, but it is accompanied by the risk of distal atheroembolism.