Shiga toxin 2 A-subunit induces mitochondrial damage, mitophagy and apoptosis via the interaction of Tom20 in Caco-2 cells.

Shiga toxin type 2 (Stx2) is the primary virulence factor produced by Shiga toxin-producing enterohemorrhagic Escherichia coli (STEC), which causes epidemic outbreaks of gastrointestinal sickness and potentially fatal sequela hemolytic uremic syndrome (HUS). Most studies on Stx2-induced apoptosis have been performed with holotoxins, but the mechanism of how the A and B subunits of Stx2 cause apoptosis in cells is not clear. Here, we found that Stx2 A-subunit (Stx2A) induced mitochondrial damage, PINK1/Parkin-dependent mitophagy and apoptosis in Caco-2 cells. PINK1/Parkin-dependent mitophagy caused by Stx2A reduced apoptosis by decreasing the accumulation of reactive oxidative species (ROS). Mechanistically, Stx2A interacts with Tom20 on mitochondria to initiate the translocation of Bax to mitochondria, leading to mitochondrial damage and apoptosis. Overall, these data suggested that Stx2A induces mitochondrial damage, mitophagy and apoptosis via the interaction of Tom20 in Caco-2 cells and that mitophagy caused by Stx2A ameliorates apoptosis by eliminating damaged mitochondria. These findings provide evidence for the potential use of Tom20 inhibition as an anti-Shiga toxin therapy.

MicroRNA-31 induced by Fusobacterium nucleatum infection promotes colorectal cancer tumorigenesis.

Persistent Fusobacterium nucleatum infection is associated with the development of human colorectal cancer (CRC) and promotes tumorigenicity, but the underlying mechanisms remain unclear. Here, we reported that F. nucleatum promoted the tumorigenicity of CRC, which was associated with F. nucleatum-induced microRNA-31 (miR-31) expression in CRC tissues and cells. F. nucleatum infection inhibited autophagic flux by miR-31 through inhibiting syntaxin-12 (STX12) and was associated with the increased intracellular survival of F. nucleatum. Overexpression of miR-31 in CRC cells promoted their tumorigenicity by targeting eukaryotic initiation factor 4F-binding protein 1/2 (eIF4EBP1/2), whereas miR-31 knockout mice were resistant to the formation of colorectal tumors. In conclusion, F. nucleatum, miR-31, and STX12 form a closed loop in the autophagy pathway, and continuous F. nucleatum-induced miR-31 expression promotes the tumorigenicity of CRC cells by targeting eIF4EBP1/2. These findings reveal miR-31 as a potential diagnostic biomarker and therapeutic target in CRC patients with F. nucleatum infection.

Isolation of Fusobacterium nucleatum from human feces using immunomagnetic separation coupled with fastidious anaerobe agar.

AIM: Fusobacterium nucleatum (F. nucleatum) is associated with the initiation, development, and metastasis of colorectal cancer. However, it is difficult to isolate F. nucleatum from clinical specimens. In this study, we aimed to develop an effective and rapid method for isolating F. nucleatum from human feces using polyclonal antibody (PAB)-coated immunomagnetic beads (IMBs) with selective media. METHODS AND RESULTS: IMBs conjugated with PAB were prepared and used to isolate F. nucleatum from human feces, and the bacteria were cultured with selective culture media (fastidious anaerobe agar + nalidixic acid + vancomycin). Under optimized experimental conditions, IMBs could selectively recover F. nucleatum from fecal microbiota samples spiked with Peptostreptococcus or Bacteroides fragilis. In artificial fecal samples, the detection sensitivity of IMBs for F. nucleatum was 103 CFU mL-1. In addition, IMBs combined with selective media could rapidly isolate F. nucleatum from human feces. CONCLUSIONS: This study successfully established an effective method for the rapid isolation of F. nucleatum from human feces by IMBs. The whole procedure requires 2-3 days, and has a sensitivity of 103 CFU mL-1 feces.

Fusobacterium nucleatum induces excess methyltransferase-like 3-mediated microRNA-4717-3p maturation to promote colorectal cancer cell proliferation.

Fusobacterium nucleatum infection plays vital roles in colorectal cancer (CRC) progression. Overexpression of microRNA-4717-3p (miR-4717) was reported to be upregulated in F. nucleatum positive CRC tissues, however, the underlying mechanism is unknown. In this study, we found that miR-4717 promoted CRC cell proliferation in vitro and growth of CRC in vivo following F. nucleatum infection. MicroRNA-4717 suppressed the expression of mitogen-activated protein kinase kinase 4 (MAP2K4), a tumor suppressor, by directly targeting its 3'-UTR. Furthermore, we confirmed that methyltransferase-like 3 (METTL3)-dependent m6 A methylation could methylate primary (pri)-miR-4717, which further promoted the maturation of pri-miR-4717, and METTL3 positively regulated CRC cell proliferation through miR-4717/MAP2K4 pathways. In conclusion, F. nucleatum-induced miR-4717 excessive maturation through METTL3-dependent m6 A modification promotes CRC cell proliferation, which provides a potential therapeutic target and diagnostic biomarker for CRC.

Circ_0001666 affects miR-620/WNK2 axis to inhibit breast cancer progression.

BACKGROUND: Circular RNAs (circRNAs) play important roles in the progression of various cancers, including breast cancer (BC). However, the role of circ_0001666 in BC remains unclear. OBJECTIVE: To explore the role of circ_0001666 in the progression of BC and reveal its potential molecular mechanism. METHODS: Real-time polymerase chain reaction was conducted to determine the expression of circ_0001666, miR-620 and with-no-lysine kinase 2 (WNK2). Cell counting kit 8 assay, flow cytometry and transwell assay were used to measure cell proliferation, apoptosis, migration and invasion. Western blot was utilized to examine the level of protein. Dual-luciferase reporter assay and RNA immunoprecipitation assay were used to verify the interaction between miR-620 and circ_0001666 or WNK2. Mice xenotransplantation models were built to explore the effect of circ_0001666 on BC tumor growth in vivo. RESULTS: Circ_0001666 was downregulated in BC tumor tissues and cells. Overexpressed circ_0001666 inhibited the proliferation, migration, invasion, while promoted apoptosis and tumor growth of BC in vitro or in vivo. Furthermore, circ_0001666 could serve as a sponge of miR-620. MiR-620 inhibitor hindered BC cell progression, which was similar to the effect of circ_0001666 overexpression. WNK2 was a target of miR-620, and circ_0001666 could sponge miR-620 to positive regulate WNK2. The knockdown of WNK2 reversed the effect of circ_0001666 overexpression on BC progression. CONCLUSION: Circ_0001666 hindered the progression of BC via miR-620/WNK2 axis.

S100 family members: potential therapeutic target in patients with hepatocellular carcinoma: A STROBE study.

ABSTRACT: Proteins in S100 family exhibit different expressions patterns and perform different cytological functions, playing substantial roles in certain cancers, carcinogenesis, and disease progression. However, the expression and role of S100 family members in the prognosis of hepatocellular carcinoma (HCC) remains unclear. To investigate the effect of S100 family members for the prognosis of liver cancer, we assessed overall survival (OS) using a Kaplan-Meier plotter (KM plotter) in liver cancer patients with different situation. Our results showed that 15 members of the S100 family exhibited high levels of expression and these levels were correlated with OS in liver cancer patients. The higher expression of S100A5, S100A7, S100A7A, S100A12, S100Z, and S100G was reflected with better survival in liver cancer patients. However, worse prognosis was related to higher levels of expression of S100A2, S100A6, S100A8, S100A9, S100A10, S100A11, S10013, S100A14, and S100P. We then evaluated the prognostic values of S100 family members expression for evaluating different stages of AJCC-T, vascular invasion, alcohol consumption, and the presence of hepatitis virus in liver cancer patients. Lastly, we studied the prognostic values of S100 family members expression for patients after sorafenib treatment. In conclusion, our findings show that the proteins of S100 family members exhibit differential expression and may be useful as targets for liver cancer, facilitating novel diagnostic and therapeutic strategies in cancer.

Non-invasive markers of cardiovascular risk in patients with subclinical hypothyroidism: A systematic review and meta-analysis of 27 case control studies.

It has been reported that subclinical hypothyroidism (SCH) is closely related to subclinical atherosclerosis. According to the impact of SCH on noninvasive markers of cardiovascular risk, we fulfilled a meta-analysis of included studies to provide an integrated overview. We searched electronic databases and included all relevant studies involving SCH and epicardial adipose tissue (EAT), carotid intima-media thickness (CIMT), pulse wave velocity (PWV), flow-mediated dilation (FMD) and glyceryl trinitrate-induced dilation (GNT- induced dilation). The result was calculated in a meta-analysis to assess the impact of SCH on these markers. A total of 27 studies were entered in the final analysis. Compared with euthyroid subjects, SCH patients exhibited a significantly increased CIMT (SMD: 0.369 mm; 95%CI: 0.038, 0.700; P = 0.029) and EAT (SMD: 1.167 mm; 95%CI: 0.869, 1.466; P = 0.000) and increased PWV (SMD: 3.574 m/s; 95%CI: 0.935, 6.213, P = 0.008). We also found significantly lower FMD (SMD: -1.525%, 95%CI: -2.156, -0.894, P = 0.000) and lower GNT-induced dilation (SMD: -0.384%, 95%CI: -0.625, -0.142, P = 0.002). Sensitivity analysis and subgroup analysis confirmed the above results. Our meta-analysis confirmed a significant association of SCH and cardiovascular risk with arterial wall thickening and stiffening and endothelial dysfunction. These findings will help to establish detailed cardiovascular prevention strategies for SCH patients.

Effect of Probiotics on Glucose and Lipid Metabolism in Type 2 Diabetes Mellitus: A Meta-Analysis of 12 Randomized Controlled Trials.

BACKGROUND It has been unclear whether supplemental probiotics therapy improves clinical outcomes in type 2 diabetic patients. This meta-analysis aimed to summarize the effect of probiotics on glucose and lipid metabolism and C-reactive protein (CRP) from 12 randomized controlled trials (RCTs). MATERIAL AND METHODS An up-to-date search was performed for all relevant RCTs up to April 2016 from PubMed, Embase, and Cochrane Library. Standardized mean difference (SMD) and weighted mean difference (WMD) were calculated for a fixed-effect and random-effect meta-analysis to assess the impact of supplemental probiotics on fasting plasma glucose (FPG), glycated hemoglobin (HbA1c), fasting insulin, homeostasis model assessment of insulin resistance (HOMA-IR), lipid profile, and CRP level. RESULTS A total of 12 studies (684 patients) were entered into the final analysis. The effect of probiotics was significant on reducing HbA1c level (standardized mean difference [SMD], -0.38; confidence interval [CI], -0.62 to -0.14, P=0.002; I²=0%, P=0.72 for heterogeneity), fasting insulin level (SMD, -0.38; CI -0.59 to -0.18, P=0.0003; I²=0%, P=0.81 for heterogeneity), and HOMA-IR (SMD, -0.99; CI -1.52 to -0.47, P=0.0002; I²=86%, P<0.00001 for heterogeneity). Pooled results on effects of probiotics on FPG, CRP, or lipid profile were either non-significant or highly heterogeneous. CONCLUSIONS This meta-analysis demonstrated that probiotics supplementation was associated with significant improvement in HbA1c and fasting insulin in type 2 diabetes patients. More randomized placebo-controlled trials with large sample sizes are warranted to confirm our conclusions.

[Effects of adenosine on hMLH1 methylation of human colorectal cancer cells].

OBJECTIVE: To explore the effects of adenosine on hMLH1 methylation of human colorectal cancer cells. METHODS: The SW480 cells were treated with adenosine at the concentrations of 0, 1.5, 3.0, 4.5 mmol/L for 72 h. The hMLH1 methylation levels of CpG islands were detected by bisulfite sequencing polymerase chain reaction (BSP), hMLH1 mRNA expression levels by reverse transcription-polymerase chain reaction (RT-PCR), the expression levels of hMLH1 protein by Western blot and the apoptotic rates by flow cytometry (FCM). The cells were treated with adenosine at the concentrations of 0, 1.5, 3.0, 4.5 mmol/L for 24, 48, 72, 96 h. And their proliferation rates were detected by methyl thiazolyl tetrazolium (MTT). RESULTS: After a 72 h treatment of adenosine, the hMLH1 promoter methylation levels of 1.5, 3.0 and 4.5 mmol/L groups were 65% ± 4%, 45% ± 11% and 16% ± 4% respectively and were all significantly lower than that of the control group (80% ± 4%, all P < 0.01). The mRNA expression levels, hMLH1 protein expression levels and apoptotic rates were all significantly higher than that of the control group (0.230 ± 0.032, 0.359 ± 0.029 and 0.570 ± 0.019 vs 0.079 ± 0.010; 0.353 ± 0.016, 0.654 ± 0.018 and 0.854 ± 0.014 vs 0.126 ± 0.016; 11.9% ± 0.6%, 20.0% ± 1.8% and 35.8% ± 1.8% vs 3.9% ± 1.4%, all P < 0.01). MTT showed that the proliferation rates of SW480 cells were lower than that of the control group and a time-dosage dependence existed (all P < 0.05). CONCLUSION: Adenosine can reverse the abnormal methylation of hMLH1 CpG island and promote the expression of hMLH1 so as to restrain the proliferation and promote the apoptosis of colocectal cancer cells.