Evaluation of glucose-lowering medications in older people: a comprehensive systematic review and network meta-analysis of randomized controlled trials.

BACKGROUND: Type 2 diabetes mellitus (T2DM) is increasingly being diagnosed in older adults. Our objective is to assess the advantages and potential drawbacks of different glucose-lowering medications in this specific population. METHODS: A network meta-analysis was conducted to identify randomized controlled trials that examined patient-centered outcomes in adults aged ≥65 years with T2DM. We searched PubMed, Cochrane CENTRAL, and Embase up to September 23, 2023. Quality of eligible studies were assessed using the Cochrane RoB 2.0 tool. RESULTS: A total of 22 trials that involved 41 654 participants were included, incorporating sodium-glucose cotransporter-2 (SGLT2) inhibitors, glucagon-like peptide-1 receptor agonists (GLP-1RAs), dipeptidyl peptidase-4 (DPP-4) inhibitors, metformin, sulfonylureas (SU) and acarbose. Our findings reveal that GLP-1RAs reduce the risk of major adverse cardiovascular events (risk ratio [RR], 0.83; 95% confidence interval [CI], 0.71 to 0.97) and body weight (mean difference [MD], -3.87 kg; 95% CI, -5.54 to -2.21). SGLT2 inhibitors prevent hospitalization for heart failure (RR, 0.66; 95% CI, 0.57 to 0.77), renal composite outcome (RR, 0.69; 95% CI, 0.53 to 0.89), and reduce body weights (MD, -1.85 kg; 95% CI, -2.42 to -1.27). SU treatment increases the risk of any hypoglycaemia (RR, 4.19; 95% CI, 3.52 to 4.99) and severe hypoglycaemia (RR, 7.06; 95% CI, 3.03 to 16.43). GLP-1RAs, SGLT2 inhibitors, metformin, SU and DPP-4 inhibitors are effective in reducing glycaemic parameters. Notably, the number of treatments needed decreases in most cases as age increases. CONCLUSIONS: Novel glucose-lowering medications with benefits that outweigh risks should be prioritized for older patients with diabetes.

Association between spirometry pattern, left ventricular diastolic function, and mortality.

BACKGROUND: Spirometric abnormalities have been related to incident heart failure in general population, who generally have preserved left ventricular ejection fraction (LVEF). We aimed to investigate the association between spirometric indices, cardiac functions and clinical outcomes. METHODS: Subjects presenting with exertional dyspnoea and received spirometry and echocardiography were eligible for this study. Forced vital capacity (FVC) and forced expiratory volume in the first second (FEV1)/FVC ratio were measured to define the spirometry patterns: normal (FEV1/FVC ≥ 70%, FVC ≥ 80%), obstructive (FEV1/FVC < 70%, FVC ≥ 80%), restrictive pattern (FEV1/FVC ≥ 70%, FVC < 80%) and mixed (FEV1/FVC < 70%, FVC < 80%). The diastolic dysfunction index (DDi) was the counts of the indicators, including septal e' velocity <7 cm/s, septal E/e' > 15, pulmonary artery systolic pressure > 35 mmHg and left atrial dimension >40 mm. RESULTS: Among a total of 8669 participants (65.8 ± 16.3 years, 56% men), 3739 (43.1%), 829 (9.6%), 3050 (35.2%) and 1051 (12.1%) had normal, obstructive, restrictive and mixed spirometry pattern, respectively. Subjects with restrictive or mixed spirometry pattern had higher DDi and worse long-term survival than those with obstructive or normal ventilation. FVC but not FEV1/FVC was predictive of 5-year mortality, independent of age, sex, renal function, LVEF, DDi, body mass index, and comorbidities (hazard ratio, 95% confidence intervals: .981, .977-.985). Furthermore, there was an inverse nonlinear relationship between FVC and DDi, suggesting the declined FVC may mediate 43% of the prognostic hazard of left ventricular diastolic dysfunction. CONCLUSIONS: The restrictive spirometry pattern or the declined FVC was associated with left ventricular diastolic dysfunction, which aggravated the long-term mortality in the ambulatory dyspnoeic subjects.

Promotion of the Differentiation of Dental Pulp Stem Cells into Oligodendrocytes by Knockdown of Heat Shock Protein 27.

Stem cell-based therapy has been evaluated in many different clinical trials for various diseases. This capability was applied in various neurodegenerative diseases, such as multiple sclerosis, which is characterized by demyelination, axonal injury, and neuronal loss. Dental pulp stem cells (DPSCs) are mesenchymal stem cells from the oral cavity that have been studied with potential application for the regeneration of different tissues. Heat shock protein 27 (HSP27) regulates neurogenesis in the process of neural differentiation of placenta multipotent stem cells. Here, we hypothesize that HSP27 expression is also critical for the neural differentiation of DPSCs. An evaluation of the possible role of HSP27 in the differentiation of DPSCs was performed using gene knockdown and neural immunofluorescent staining. We found that HSP27 played a role in the differentiation of DPSCs and that knockdown of HSP27 in DPSCs rendered cells to oligodendrocyte progenitors; i.e., small hairpin specific for HSP27 DPSCs exhibited NG2-positive immunoreactivity and gave rise to oligodendrocytes or type-2 astrocytes. This neural differentiation of DPSCs may have clinical significance in the treatment of patients with neurodegenerative diseases. In conclusion, our data provide an example of the oligodendrocyte differentiation of a DPSC model, which may be applied in human regenerative medicine.

Colorectal cancer concurrent gene signature based on coherent patterns between genomic and transcriptional alterations.

BACKGROUND: The aim of the study was to enhance colorectal cancer prognostication by integrating single nucleotide polymorphism (SNP) and gene expression (GE) microarrays for genomic and transcriptional alteration detection; genes with concurrent gains and losses were used to develop a prognostic signature. METHODS: The discovery dataset comprised 32 Taiwanese colorectal cancer patients, of which 31 were assayed for GE and copy number variations (CNVs) with Illumina Human HT-12 BeadChip v4.0 and Omni 25 BeadChip v1.1. Concurrent gains and losses were declared if coherent manners were observed between GE and SNP arrays. Concurrent genes were also identified in The Cancer Genome Atlas Project (TCGA) as the secondary discovery dataset (n = 345). RESULTS: The "universal" concurrent genes, which were the combination of z-transformed correlation coefficients, contained 4022 genes. Candidate genes were evaluated within each of the 10 public domain microarray datasets, and 1655 (2000 probe sets) were prognostic in at least one study. Consensus across all datasets was used to build a risk predictive model, while distinct relapse-free/overall survival patterns between defined risk groups were observed among four out of five training datasets. The predictive accuracy of recurrence, metastasis, or death was between 61 and 86% (cross-validation area under the receiver operating characteristic (ROC) curve: 0.548-0.833) from five independent validation studies. CONCLUSION: The colorectal cancer concurrent gene signature is prognostic in terms of recurrence, metastasis, or mortality among 1746 patients. Genes with coherent patterns between genomic and transcriptional contexts are more likely to provide prognostication for colorectal cancer.

Revisiting 'intensive' blood glucose control: A causal directed acyclic graph-guided systematic review of randomized controlled trials.

AIM: To clarify the importance of HbA1c reduction and antidiabetic drug use in preventing major adverse cardiovascular events (MACE) for patients with type 2 diabetes (T2D). MATERIALS AND METHODS: We conducted an updated systematic review of contemporary large randomized controlled trials assessing the relative efficacy and safety of antidiabetic drugs with less hypoglycaemia risk in adult T2D patients. Mixed-effects meta-regression was performed to examine the associations of HbA1c reduction with subsequent risk of macrovascular and microvascular events. We evaluated the potential mediating role of HbA1c reduction in the relationship between antidiabetic drugs and MACE. RESULTS: Eighteen placebo-controlled trials comprising 155 610 participants were included. The effects of treatment differed among antidiabetic drug classes for most adverse outcomes with high heterogeneity (I2 : 63.7%-95.8%). Mean HbA1c reduction was lowest with dipeptidyl peptidase-4 inhibitors (0.30%), followed by sodium-glucose co-transporter-2 inhibitors (0.46%), and was highest with glucagon-like peptide-1 receptor agonists (0.58%) and thiazolidinediones (0.60%). Lower relative risks of MACE were significantly associated with larger reductions in achieved HbA1c (ß -0.3182; 95% CI: -0.5366 to -0.0998; P = .0043), even after adjusting for drug classes. When considering HbA1c lowering as a mediator to be controlled, beneficial effects owing to specific antidiabetic treatment for MACE were not observed (χ2  = 1.4494; P = .6940). The proportion mediated by HbA1c reduction was 50.0%-63.5% for these antidiabetic agents. CONCLUSIONS: The main benefits of antidiabetic agents might result from the reduction in blood sugar levels and are generally independent of drugs used. Risk reduction in MACE was proportional to the magnitude of HbA1c decrease conferred by antidiabetic agents with less hypoglycaemic hazard.

The extended concurrent genes signature for disease-free survival in breast cancer.

BACKGROUND/PURPOSE: Previously we had identified concurrent genes, which highlighted the interplay between copy number variation (CNV) and differential gene expression (GE) for Han Chinese breast cancers. The merit of the approach is to discovery biomarkers not identifiable by conventional GE only data, for which phenotype-correlation or gene variability is the criteria of gene selection. MATERIALS AND METHODS: Thirty-one comparative genomic hybridization (CGH) and 83 GE microarrays were performed, with 29 breast cancers assayed from both platforms. Potential targets were revealed by Genomic Identification of Significant Targets in Cancer (GISTIC) from CGH arrays. Concurrent genes and genes with significant GISTIC scores were used to derive the extended concurrent genes signature, which was consensus from leading edge analysis across all studies and a supervised partial least square (PLS) regression predictive model of disease-free survival was constructed. RESULTS: There were 1584 concurrent genes from 29 samples with both CGH and GE microarrays. Enriched concurrent genes sets for disease-free survival were identified independently from 83 GE arrays and another one with Han Chinese origin as well as three studies of Western origin. For five studies with disease-free survival follow up, prognostic discrepancy was observed between predicted high-risk and low-risk group patients. CONCLUSION: We concluded that through parallel analyses of CGH and GE microarrays, the proposed extended concurrent gene expression signature can identify biomarkers with prognostic values.

Functional Plasmon-Activated Water Increases Akkermansia muciniphila Abundance in Gut Microbiota to Ameliorate Inflammatory Bowel Disease.

Inflammatory bowel disease (IBD) is associated with dysbiosis and intestinal barrier dysfunction, as indicated by epithelial hyperpermeability and high levels of mucosal-associated bacteria. Changes in gut microbiota may be correlated with IBD pathogenesis. Additionally, microbe-based treatments could mitigate clinical IBD symptoms. Plasmon-activated water (PAW) is known to have an anti-inflammatory potential. In this work, we studied the association between the anti-inflammatory ability of PAW and intestinal microbes, thereby improving IBD treatment. We examined the PAW-induced changes in the colonic immune activity and microbiota of mice by immunohistochemistry and next generation sequencing, determined whether drinking PAW can mitigate IBD induced by 2,4,6-trinitrobenzene sulfonic acid (TNBS) and dysbiosis through mice animal models. The effects of specific probiotic species on mice with TNBS-induced IBD were also investigated. Experimental results indicated that PAW could change the local inflammation in the intestinal microenvironment. Moreover, the abundance of Akkermansia spp. was degraded in the TNBS-treated mice but elevated in the PAW-drinking mice. Daily rectal injection of Akkermansia muciniphila, a potential probiotic species in Akkermansia spp., also improved the health of the mice. Correspondingly, both PAW consumption and increasing the intestinal abundance of Akkermansia muciniphila can mitigate IBD in mice. These findings indicate that increasing the abundance of Akkermansia muciniphila in the gut through PAW consumption or other methods may mitigate IBD in mice with clinically significant IBD.

Deciphering Genetic Alterations of Taiwanese Patients with Pancreatic Adenocarcinoma through Targeted Sequencing.

Pancreatic adenocarcinoma (PAC) is the 8th leading cause of cancer-related deaths in Taiwan, and its incidence is increasing. The development of PAC involves successive accumulation of multiple genetic alterations. Understanding the molecular pathogenesis and heterogeneity of PAC may facilitate personalized treatment for PAC and identify therapeutic agents. We performed tumor-only next-generation sequencing (NGS) with targeted panels to explore the molecular changes underlying PAC patients in Taiwan. The Ion Torrent Oncomine Comprehensive Panel (OCP) was used for PAC metastatic lesions, and more PAC samples were sequenced with the Ion AmpliSeq Cancer Hot Spot (CHP) v2 panel. Five formalin-fixed paraffin-embedded (FFPE) metastatic PAC specimens were successfully assayed with OCP, and KRAS was the most prevalent alteration, which might contraindicate the use of anti-EGFR therapy. One PAC patient harbored a FGFR2 p. C382R mutation, which might benefit from FGFR tyrosine kinase inhibitors. An additional 38 samples assayed with CHP v2 showed 100 hotspot variants, collapsing to 54 COSMID IDs. The most frequently mutated genes were TP53, KRAS, and PDGFRA (29, 23, 10 hotspot variants), impacting 11, 23, and 10 PAC patients. Highly pathogenic variants, including COSM22413 (PDGFRA, FATHMM predicted score: 0.88), COSM520, COSM521, and COSM518 (KRAS, FATHMM predicted score: 0.98), were reported. By using NGS with targeted panels, somatic mutations with therapeutic potential were identified. The combination of clinical and genetic information is useful for decision making and precise selection of targeted medicine.

MitoTox: a comprehensive mitochondrial toxicity database.

BACKGROUND: Mitochondria play essential roles in regulating cellular functions. Some drug treatments and molecular interventions have been reported to have off-target effects damaging mitochondria and causing severe side effects. The development of a database for the management of mitochondrial toxicity-related molecules and their targets is important for further analyses. RESULTS: To correlate chemical, biological and mechanistic information on clinically relevant mitochondria-related toxicity, a comprehensive mitochondrial toxicity database (MitoTox) was developed. MitoTox is an electronic repository that integrates comprehensive information about mitochondria-related toxins and their targets. Information and data related to mitochondrial toxicity originate from various sources, including scientific journals and other electronic databases. These resources were manually verified and extracted into MitoTox. The database currently contains over 1400 small-molecule compounds, 870 mitochondrial targets, and more than 4100  mitochondrial toxin-target associations. Each MitoTox data record contains over 30 fields, including biochemical properties, therapeutic classification, target proteins, toxicological data, mechanistic information, clinical side effects, and references. CONCLUSIONS: MitoTox provides a fully searchable database with links to references and other databases. Potential applications of MitoTox include toxicity classification, prediction, reference and education. MitoTox is available online at http://www.mitotox.org .

Residual risk stratification of Taiwanese breast cancers following curative therapies with the extended concurrent genes signature.

INTRODUCTION: The aim of the study was to perform digital RNA counting to validate a gene expression signature for operable breast cancers initially treated with curative intention, and the risk of recurrence, distant metastasis, and mortality was predicted. METHODS: Candidate genes were initially discovered from the coherent genomic and transcriptional alternations from microarrays, and the extended concurrent genes were used to build a risk stratification model from archived formalin-fixed paraffin-embedded (FFPE) tissues with the NanoString nCounter. RESULTS: The extended concurrent genes signature was prognostic in 144 Taiwanese breast cancers (5-year relapse-free survival: 89.8 and 69.4% for low- and high-risk group, log-rank test: P = 0.004). Cross-platform comparability was evidenced from significant and positive correlations for most genes as well as equal covariance matrix across 64 patients assayed for both microarray and digital RNA counting. DISCUSSION: Archived FFPE samples could be successfully assayed by the NanoString nCounter. The purposed signature was prognostic stratifying breast cancer patients into groups with distinct survival patterns, and clinical applicability of the residual risk model was proved.

Effect of Acoustic Cardiography-guided Management on 1-year Outcomes in Patients With Acute Heart Failure.

BACKGROUND: The electromechanical activation time (EMAT) normalized by cardiac cycle length (%EMAT) and the third heart sound (S3) strength, as measured by automated acoustic cardiography, are predictive of postdischarge adverse events in patients with acute heart failure (AHF). The aim of this study was to evaluate whether the acoustic cardiography-guided management improves outcomes in patients with AHF when it is compared with the conventional therapy. METHODS AND RESULTS: This prospective single-blind study randomized 225 patients with AHF (74.1 ± 14.5 years of age, 26.2% women, and left ventricular ejection fraction 38.4 ± 14.4%) before discharge to the EMAT-guided group (n = 114) with the postdischarge treatment goals to reduce %EMAT to < 15% and S3 < 5, and the symptom-guided group (n = 111) to adjust medications without knowledge of the results of acoustic cardiography. The primary endpoints were rehospitalization for heart failure and total mortality during 1-year follow-up. The 2 groups were well matched in age and predischarge %EMAT and S3 strength. After a mean follow-up period of 238.1 ± 140.8 days, a significant reduction in the primary endpoints was seen in the EMAT-guided group compared with the symptom-guided group (43 events vs 61 events, P = 0.0095). Kaplan-Meier curves demonstrated significant differences in the time to first event, favoring the EMAT-guided group in the total study population (n = 225, hazard ratio and 95% confidence interval: 0.61, 0.42-0.91, log-rank P = 0.0129), as well as in the prespecified subgroup of patients with predischarge %EMAT > 15% (n = 85; 0.32, 0.16-0.65, P = 0.0008). CONCLUSIONS: In patients hospitalized due to AHF, EMAT-guided postdischarge management was superior to the conventional symptoms-driven therapy in terms of 1-year outcomes (ClinicalTrials.gov number NCT01298232).

Feasibility and rationale of direct current cardioversion immediately after transcatheter percutaneous edge-to-edge mitral valve repair.

AIMS: Atrial fibrillation (AF) is a frequent comorbidity among patients with severe mitral regurgitation (MR). Direct current (DC) cardioversion is one of the strategies for rhythm control. However, the safety and feasibility of immediate DC cardioversion after MitraClip are not elucidated. METHODS AND RESULTS: In this study, patients with symptomatic severe MR who underwent MitraClip were included. After fixing the MR, synchronized DC cardioversion was attempted for those with AF. A total of consecutive 60 patients, 36 subjects (60%), comorbid with AF. DC cardioversion was performed in 30 patients (mean age of 76.0 ± 9.3 years), and the successful conversion was achieved in 15 patients (50%). There was no any adverse event related to the cardioversion. Subjects with sustained conversion to SR experienced significant improvement in 6MWT (failed: 285 ± 110-308 ± 135 m, P = .278; successful: 269 ± 109 m-328 ± 78, P = .047) and reduction in NT-proBNP level (failed: 4411 ± 7401-3296 ± 4299 ng/mL, P = .217; successful: 4094 ± 2735-2353 ± 2856 ng/mL, P = .026) at 1 month. CONCLUSIONS: Direct current cardioversion seemed to be safe and feasible immediately after the transcatheter edge-to-edge mitral valve repairs. Subjects who maintain SR experienced better functional improvement.

Rare Concurrence of Two Congenital Disorders: Miller-Dieker Syndrome and T-Cell Lymphopenia.

Miller-Dieker syndrome (MDS; OMIM 247200) is a rare contiguous gene deletion syndrome associated with lissencephaly and characteristic facial dysmorphism. T-cell lymphopenia is an immunodeficiency disorder which can be early detected by newborn blood screening, and all live vaccines should be avoided. We report a 2.32-Mb microdeletion at chromosome 17p13.3p13.2 and T-cell lymphopenia in a 6-month-old male infant with MDS. This is, to our knowledge, the first description of these 2 conditions co-occurring in the same patient.

Ganoderma microsporum immunomodulatory protein induces apoptosis and potentiates mitomycin C-induced apoptosis in urinary bladder urothelial carcinoma cells.

Current chemotherapy and immunotherapy treatments followed by transurethral resection for urinary bladder urothelial carcinoma (UC) usually suffer from poor prognosis and high recurrence rate. Design and modification of current formulation with the novel adjuvants are needed. A recombinant protein derived from Ganoderma microsporum named as Ganoderma microsporum immunomodulatory protein (GMIP) was used to treat UC cells. We found GMIP elicits a dose-dependent and time-dependent anti-UC cell proliferation effect, with a half-maximal inhibition concentration (IC50 ) comparable to mitomycin C (MMC), a commonly used chemotherapy agent. After GMIP treatment, UC cells showed apoptotic phenomenon including cell cycle arrest in the G1 phase, elevated sub-G1 population, mitochondrial membrane potential loss, up-regulated p21 expression, p21 nuclear translocation, caspase activation, and PARP cleavage in a p53-independent but p21-mediated pathways. Unlike lung cancer cells, GMIP treated UC cells showed no autophagic scheme including Beclin-1, an autophagy to apoptosis switch marker, was not cleaved by caspase 3 and slight LC3B-II accumulation. Also, the classic autophagic inhibitor, chloroquine had no effect in GMIP-mediated cell death made us conclude that GMIP induced apoptosis through caspase activation but not autophagy in UC cells. Additionally, GMIP showed synergistic effects with MMC in killing UC cells and thus decreased the concentration of MMC usage to reach the comparable apoptotic effects. Our results delineate novel strategies for treatment of UC by GMIP alone or in combination with MMC application and provide a promising therapeutic cocktail for better treatment of urinary bladder urothelial carcinoma.

Blood glucose reduction by diabetic drugs with minimal hypoglycaemia risk for cardiovascular outcomes: Evidence from meta-regression analysis of randomized controlled trials.

AIMS: To investigate the effects of blood glucose control with antihyperglycaemic agents with minimal hypoglycaemia risk on cardiovascular outcomes in patients with type 2 diabetes (T2D). MATERIALS AND METHODS: Randomized controlled trials (RCTs) comparing the relative efficacy and safety of antidiabetic drugs with less hypoglycaemia risk were comprehensively researched in MEDLINE, Embase and the Cochrane Library up to January 27, 2018. Mixed-effects meta-regression analysis was conducted to explore the relationship between haemoglobin A1c (HbA1c) reduction and the risk of major adverse cardiovascular events (MACE), myocardial infarction, stroke, cardiovascular death, all-cause death, and hospitalization for heart failure. RESULTS: Ten RCTs comprising 92 400 participants with T2D were included and provided information on 9773 MACE during a median follow-up of 2.6 years. The mean HbA1c concentration was 0.42% lower (range, 0.27%-0.86%) for participants given antihyperglycaemic agents than those given placebo. The meta-regression analysis demonstrated that HbA1c reduction was significantly associated with a decreased risk of MACE (ß value, -0.39 to -0.55; P < 0.02) even after adjusting for possible confounding factors including age, sex, baseline HbA1c, duration of follow-up, difference in achieved systolic blood pressure, difference in achieved body weight, and risk difference in hypoglycaemia. Lowering HbA1c by 1% conferred a significant risk reduction of 30% (95% confidence interval, 17%-40%) for MACE. By contrast, the meta-regression analysis for trials using conventional agents failed to demonstrate a significant relationship between achieved HbA1c difference and MACE risk (P > 0.74). CONCLUSIONS: Compared with placebo, newer T2D agents with less hypoglycaemic hazard significantly reduced the risk of MACE. The MACE reduction appears to be associated with HbA1c reduction in a linear relationship.

Upregulation of the growth arrest-specific-2 in recurrent colorectal cancers, and its susceptibility to chemotherapy in a model cell system.

Colorectal cancer (CRC) is one of the most common life-threatening malignances worldwide. CRC relapse markedly decreases the 5-year survival of patients following surgery. Aberrant expression of genes involved in pathways regulating the cell cycle, cell proliferation, or cell death are frequently reported in CRC tumorigenesis. We hypothesized that genes involved in CRC relapse might serve as prognostic indicators. We first evaluated the significance of gene sequences in the feces of patients with CRC relapse by consulting a public database. Tumorigenesis of target tissues was tested through tumor cell growth, cell cycle regulation, and chemotherapeutic efficacy. We found a highly significant correlation between CRC relapse and growth arrest-specific 2 (GAS2) gene expression. Based on cell models, the overexpressed GAS2 was associated with cellular growth rate, cell cycle regulation, and with chemotherapeutic sensitivity. Cell division was impaired by treating cells with 2-[4-(7-chloro-2-quinoxalinyloxy)phenoxy]-propionic acid (XK469), even when the cells were overexpressing GAS2. Thus, downregulation of GAS2 expression might control CRC relapse after curative resection. GAS2 could serve as a noninvasive marker from the feces of patients with prediagnosed CRC. Our findings suggest that GAS2 could have potential clinical applications for predicting early CRC relapse after radical resection, and that XK469 might impair tumor cell division by reducing GAS2 expression or blocking its cellular translocation. This will help in selecting the best therapeutic option, 5-fluorouracil in combination with XK469, for patients overexpressing GAS2 in CRC cells. Thus, GAS2 might act as a prognostic biomolecule and potential therapeutic target in patients with CRC relapse.

Genome-wide identification of blood DNA methylation patterns associated with early-onset hepatocellular carcinoma development in hepatitis B carriers.

The etiology of early-onset hepatocellular carcinoma (HCC) among hepatitis B virus (HBV) carriers remains unclear. DNA methylation levels in peripheral leukocytes have been associated with different environmental exposures and immune or inflammatory response. We aimed to identify methylation signatures of peripheral leukocytes that could track hepatitis B progression to HCC, especially for early-onset HCC. We first performed an epigenome-wide association analysis on 48 matched case-control pairs in a nested case-control study within a 22-yr follow-up cohort of HBV carriers. Through this analysis we found that progression to early-onset HCC involved methylation variable positions across the genome, in which a substantial proportion displayed significant variation due to HBV viral load, chronic hepatitis status, and/or leukocyte subtype composition, and these associations were significantly enriched among genes in immune pathways. Methylation at probes cg00300879, cg06872964, and cg07080864, that are located within the proximal promoter of CNKSR1, IFI44L, and PENK, respectively, was validated by bisulfite pyrosequencing and findings were replicated in a case-sibling study of early-onset HCC (134 cases vs. 174 sibling controls). Furthermore, a high methylation score, constructed using the three probes, was predictive for the risk of early-onset HCC in two datasets (adjusted-odds ratios = 0.21-0.32, P ≤ 0.0206). This association was also observed for late-onset HCC (adjusted-odds ratio = 0.42-0.47, P ≤ 0.0194) in a nested case-control study (120 cases vs. 178 controls). In prospective analysis, change in the score was detected 5-9 yr before HCC onset. Blood-based methylation profiling provides new insights into the complexity of virus-host interaction underlying HBV-related HCC, holding promise for the disease risk management. © 2016 Wiley Periodicals, Inc.

Progressive accumulation of mutations in the hepatitis B virus genome and its impact on time to diagnosis of hepatocellular carcinoma.

UNLABELLED: To evaluate how hepatitis B virus (HBV) genetic variation affected progression from chronic carrier state to hepatocellular carcinoma (HCC), we analyzed HBV full-length sequences in blood obtained <1-20 years before diagnosis from 117 HCC cases and 118 controls nested in a cohort of 4,841 HBV carriers, for whom HBV genotypes B and C are predominant. The relationship between each viral single-nucleotide polymorphism (SNP) and HCC development was assessed using ordinal logistic models according to five periods of time to diagnosis (TTD). Thirty-one HBV-SNPs showed significant association with TTD after adjustment for HBV genotype, 24 of which could also be analyzed with an extended analysis on the full-length data in conjunction with 512 partial sequences (nucleotides 2,436-1,623) from the cohort. The obtained 10 robust candidate HBV-SNPs (P ≤ 0.0304), which showed odds ratios ranging from 1.89 to 8.68, were further confirmed in 163 GenBank HBV-HCC sequences from nine Asia regions, assayed after HCC diagnosis, representing the end stage of progressive hepatic diseases. The prevalence of these HBV-SNPs and their cumulative number, presented in terms of mutation score, increased with time approaching HCC diagnosis, with an odds ratio of 2.17, 4.21, 8.15, and 19.15, respectively, for the mutation score of 1, 2, 3, and ≥4 versus 0. The mutation score for predicting short-term HCC risk outperformed other factors, including HBV-DNA levels, viral genotype, and various combinations of risk factors, and revealed increasing accuracy with shorter TTD (<4.5 years before diagnosis: area under the curve = 0.83-0.89; sensitivity = 72.7%-94.1%; specificity = 58.3%-70.5%; conditioned on optimized cutoff for genotype B and C, respectively). CONCLUSIONS: Identifying and tracking viral mutations is important for monitoring hepatitis B progression and early detection of HCC. (Hepatology 2016;64:720-731).

Cellular effects induced by 17-ß-estradiol to reduce the survival of renal cell carcinoma cells.

BACKGROUND: Renal cell carcinoma (RCC) is an adult malignancy with 2:1 men-to-women ratio, which implies the possible role of sex hormones in RCC carcinogenesis. One of the predominant sex hormones in women before menopause, 17-ß-estradiol (or E2), may regulate RCC growth by cellular mechanisms that are still not fully understood. METHODS: The expression levels of E2 receptors (ER1 and ER2) were determined in different RCC cell lines. The DNA damage response induced by E2 was determined by a DNA double-strand break marker γH2AX. To study the possible effect of E2 on oxidative stress response, RCC cells were stained with 2,7-dichlorofluorescein diacetate and analyzed by flow cytometry. Upregulation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) ser40 phosphorylation in response to oxidative stress was detected by immunoblotting. Finally, annexin V/propidium iodide (PI) double staining assay was used to determine E2-induced cellular apoptosis. RESULTS: Variable expression of ER1 and ER2 were found in the RCC cell lines studied (786-O, A498, and ACHN), in which ACHN and A498 showed highest and lowest ER expression, respectively. In A498 cells, E2 induced DNA double-strand breaks with positive staining of γH2AX. On the other hand, the level of reactive oxidative species were elevated in ACHN cells after E2 treatment. The E2-induced oxidative stress also induced the Ser40 phosphorylation and nuclear translocation of Nrf2. Finally, we also demonstrated that E2 induced apoptosis as revealed by annexin V/PI double staining. CONCLUSIONS: In this study, we demonstrated the cellular effects of E2 on DNA repair, ROS production as well as Nrf2 activation, and apoptosis in RCC cell lines. Together these cellular alterations may contribute to the reduced viability of RCC cells following E2 treatment.

Demethylation of HIN-1 reverses paclitaxel-resistance of ovarian clear cell carcinoma through the AKT-mTOR signaling pathway.

BACKGROUND: Methylation of HIN-1 is associated with poor outcomes in patients with ovarian clear cell carcinoma (OCCC), which is regarded to be an aggressive, chemo-resistant histological subtype. This study aimed to evaluate whether 5-aza-2-deoxycytidine (5-aza-2-dC) can reverse methylation of the HIN-1 gene to restore chemo-sensitivity of OCCC and the possible mechanism. METHODS: In vitro flow cytometric analysis and evaluation of caspase-3/7 activity of paclitaxel-sensitive and resistant OCCC cell lines were performed. Methylation status and expression changes of HIN-1 in the OCCC cell lines treated with 5-aza-2-dC were evaluated, and immunohistochemical staining of HIN-1 in OCCC tissues was performed. In vivo tumor growth with or without 5-aza-2-dC treatment was analyzed, and Western blotting of AKT-mTOR signaling-related molecules was performed. RESULTS: G2-M phase arrest was absent in paclitaxel-resistant OCCC cells after treatment with the cytotoxic drug. The caspase activities of the chemo-resistant OCCC cells were lower than those of the chemo-sensitive OCCC cells when treated with paclitaxel. Methylation of HIN-1 was noted in paclitaxel-resistant OCCC cell lines and cancerous tissues. 5-aza-2-dC reversed the methylation of HIN-1, re-activated the expression of HIN-1, and then suppressed the in vivo tumor growth of paclitaxel-resistant OCCC cells. Immunoblotting revealed that phospho-AKT473 and phospho-mTOR were significantly increased in HIN-1-methylated paclitaxel-resistant OCCC cell lines. However, the expressions of phospho-AKT at Ser473 and Thr308 and phospho-mTOR decreased in the OCCC cells with a high expression of HIN-1. CONCLUSIONS: Demethylating agents can restore the HIN-1 expression in paclitaxel-resistant OCCC cells through the HIN-1-AKT-mTOR signaling pathway to inhibit tumor growth.