Susceptibility-guided sequential strategy versus empirical therapy for Helicobacter pylori infection: study protocol for a randomised controlled trial.

BACKGROUND: New treatment strategies are required against infections caused by Helicobacter pylori, which grows increasingly resistant to antibiotics. Polymerase chain reaction-based methods for antibiotic susceptibility testing are available for detecting H. pylori-specific mutations that confer resistance to clarithromycin and levofloxacin. Several meta-analyses have compared eradication rates for susceptibility-guided versus empirical therapy for H. pylori treatment; however, all have significant limitations and high heterogeneity, and the results are contradictory. The main objective of this trial is to assess whether a sequential strategy based on molecular susceptibility testing-guided therapy for H. pylori has a better eradication rate than empirical therapy. METHODS: This trial is designed as a prospective, randomised, open-label, active-controlled and single-centre study. Men and women who are H. pylori-positive, naïve to treatment, and aged 18-65 years will be recruited. A total of 500 participants will be randomised to receive either empirical therapy or a susceptibility-guided sequential strategy. Bismuth quadruple therapy will be the empirical first-line therapy, and in case of failure, high-dose dual (proton-pump inhibitor + amoxicillin) treatment will be the rescue therapy. For the susceptibility-guided sequential strategy, regimen selection will be based on H. pylori susceptibility to clarithromycin (first-line) and levofloxacin (rescue). A first-line treatment of clarithromycin triple therapy will be selected for clarithromycin-sensitive strains. For clarithromycin resistance, a high-dose dual therapy will be selected. During the rescue treatment, a levofloxacin quadruple regimen will be selected for levofloxacin-sensitive strains, and a furazolidone quadruple regimen will be selected for others. The primary outcome is the first-line eradication rate in both groups, and the overall (including first and rescue therapies) H. pylori eradication rate in both groups is one of the secondary outcomes. The eradication rates of H. pylori will be analysed by intention-to-treat analysis, modified intention-to-treat analysis, and per-protocol analysis. DISCUSSION: This randomised controlled trial will provide objective and valid evidence about the value of polymerase chain reaction-based molecular methods for antibiotic susceptibility testing in guiding H. pylori eradication. TRIAL REGISTRATION: Clinicaltrials.gov NCT05549115. Released on 18 September 2022. First posted on 22 September 2022. Enrolment of the first participant on 20 September 2022. The study is retrospectively registered.

miRNA-99a inhibits cell invasion and migration in liver cancer by directly targeting HOXA1.

Liver cancer is a malignant tumor that threatens human health worldwide. It has poor prognosis rates and ineffective therapeutic options. Recently, various miRNAs have been proven to exert promoting or inhibiting functions in different malignancies. However, the definitive mechanisms of miR-99a in liver cancer remain unclear. In the current study, we explored the relationships between the expression of miR-99a and HOXA1 in liver cancer tissues and cells to explore their combined effects on the occurrence and metastasis of liver cancer. The expression of miR-99a and HOXA1 in liver cancer tissue samples and cells was measured by RT-qPCR. Following transfection, transwell assays were conducted to assess the invasion and migration capacities of liver cancer cells. Subsequently, western blots and luciferase reporter assays were performed in liver cancer cells to identify the target of miR-99a. The data indicated that miRNA-99a expression was significantly reduced in both liver cancer tissue samples and cells compared with normal tissues and normal liver cells respectively. By contrast, the HOXA1 expression levels in liver cancer tissues and cells were significantly increased in contrast to the control group. The findings also revealed that the miR-99a expression was negatively correlated with HOXA1 expression in liver cancer tissue samples and miR-99a could suppress cell invasion and migration by targeting HOXA1 in liver cancer.

Therapeutic efficacy and safety of umbilical cord mesenchymal stem cell transplantation for liver cirrhosis in Chinese population: A meta-analysis.

BACKGROUND AND OBJECTIVE: Mesenchymal stem cells transfusion has been considered as a promising option for liver cirrhosis (LC). The aim of this study was to systematically evaluate the efficacy and safety of umbilical cord mesenchymal stem cells (UMSC) combined with traditional supportive therapy (TST) for the treatment of patients with LC. METHODS: Data was extracted from clinical trials published on Web of Science, PubMed, EMBASE, Cochrane Library, Wanfang and CNKI database. The evaluated outcome measurements included liver function, coagulation function, liver fibrosis indexes, clinical symptoms, quality of life (QOL) and adverse events. RESULTS: A total of 14 trials including 717 LC patients met our selection criteria were involved. The liver function of LC patients was significantly improved after combined therapy (UMSC plus TST), indicated by decreased total bilirubin, alanine aminotransferase and prothrombin time, and increased serum albumin, cholinesterase and prothrombin activity. The QOL of patients was also improved after UMSC therapy. Compared with TST alone, the combined therapy showed better treatment effect based on measurements of hyaluronic acid (OR=-143.20, CI=-181.58 to -104.82, P<0.00001), laminin (OR=-50.65, CI=-53.70 to -47.61, P<0.00001), type III procollagen (OR=-8.68, CI=-9.00 to -8.36, P<0.00001), type IV collagen (OR=-105.79, CI=-132.44 to -79.14, P<0.00001) and plasma prolidase (OR=-876.54, CI=-911.89 to -840.56, P<0.00001). Moreover, the patients' clinical symptoms including fatigue (4th, P=0.003; 8th, P=0.01), appetite (4th, P<0.0001; 8th, P=0.06), ascites (4th, P=0.03; 8th, P=0.17), and abdominal distension (4th, P=0.0008; 8th, P=0.64) were also improved in patients treated by combined therapy without adverse events observed. CONCLUSION: UMSC and TST combined therapy for LC patients improved their liver function, clinical symptoms and QOL without severe adverse events, therefore is safe and effective in LC therapy.

Serum omentin-1 as a disease activity marker for Crohn's disease.

BACKGROUND AND AIM: It remains challenging to determine the inflammatory activity in Crohn's disease (CD) for lack of specific laboratory markers. Recent studies suggest that serum omentin-1 is associated with inflammatory response. We aimed to assess the potential of serum omentin-1 as a marker of disease activity in CD patients. METHODS: Serum omentin-1 concentrations were determined by enzyme-linked immunosorbent assay (ELISA) in patients with CD (n = 240), functional gastrointestinal disorders (FGDs, n = 120), and healthy controls (HC, n = 60) and evaluated for correlation with disease activity. Expression of omentin-1 in colonic tissues from patients with CD was also analyzed by real-time PCR and Western blotting. Serum omentin-1 levels as an activity index were evaluated using a receiver operating characteristic (ROC) curve. RESULTS: Serum omentin-1 concentrations were significantly decreased in active CD patients compared with patients in remission, FGDs, and HC (all P < 0.001). Expression of omentin-1 was decreased at mRNA and protein levels in inflamed colonic tissues in active CD than that in noninflamed colonic tissues. Serum omentin-1 levels were negatively correlated with disease activity in CD, better than C-reactive protein (CRP). CONCLUSION: Our results indicate that serum and colonic omentin-1 expressions are decreased in active CD patients. The correlation of serum omentin-1 with disease activity in CD is superior to that of CRP. Serum omentin-1 is a potential marker for CD disease activity.

Down-regulation of NAMPT expression by miR-182 is involved in Tat-induced HIV-1 long terminal repeat (LTR) transactivation.

Tat's transactivating activity is controlled by sirtuin 1 (SIRT1) that connects HIV transcription with the metabolic state of the cell. Nicotinamide phosphoribosyltransferase (NAMPT) is a key enzyme in the salvaging pathway for the synthesis of nicotinamide adenine dinucleotide (NAD(+)) that is involved in energy metabolism. Host encoded microRNAs (miRNAs) may influence viral replication. In this study, our goal was aimed to investigate the regulation of miR-182 in TZM-bl cells and explore the mechanisms by which miR-182 influenced Tat-induced HIV-1 transactivation through targeting at down-regulation of NAMPT expression. We showed that miR-182 was up-regulated when Tat was expressed in TZM-bl cells. MiR-182 significantly inhibited NAMPT protein expression by acting on the 3'-UTR of the NAMPT mRNA. MiR-182 was involved in Tat-induced NAD(+) depletion, down-regulation of SIRT1 protein expression and activity, increased acetylation of p65. Forced expression of "miR-182 mimics" increased Tat-induced LTR transactivation. Our results uncover previously unknown links between Tat and a specific host cell miRNA that targets NAMPT. Our results suggest that strategies to augment NAMPT protein expression by down-regulation of miR-182 may have therapeutic benefits to prevent HIV-1 replication.

MiR-34a is involved in Tat-induced HIV-1 long terminal repeat (LTR) transactivation through the SIRT1/NFκB pathway.

MicroRNAs (miRNAs) regulate gene expression and may contribute to HIV-1 infection. In this study, our goal was to investigate the mechanisms by which miR-34a influenced Tat-induced HIV-1 transactivation through the SIRT1/NFκB pathway. We showed that Tat induced up-regulation of miR-34a expression in TZM-bl cells. MiR-34a significantly inhibited SIRT1 expression. Overexpression of miR-34a increased Tat-induced LTR transactivation. Forced expression of miR-34a decreased SIRT1 protein expression and consequently diminished Tat-induced acetylation of p65, while treatment with a miR-34a inhibitor had the opposite effect. These results suggest that regulating SIRT1 by down-regulation of miR-34a levels may be a therapeutic strategy against HIV-1 replication.

EGCG inhibits Tat-induced LTR transactivation: role of Nrf2, AKT, AMPK signaling pathway.

AIMS: Transcription is a crucial step for human immunodeficiency virus 1 (HIV-1) gene expression in infected host cells. The HIV-1 Tat activates the nuclear factor-kappa B (NF-κB) signaling transduction pathway, which is necessary for viral replication. Epigallocatechin-3-gallate (EGCG) has antioxidant, anti-inflammatory, and anti-viral properties. In this study, we investigated the effects of EGCG on Tat-induced HIV-1 transactivation and potential mechanisms by which EGCG inhibited activation of NF-κB pathway. MAIN METHODS: HeLa-CD4-long terminal repeat (LTR)-ß-gal (MAGI) cells were transfected with Tat plasmid. Tat-induced HIV-1 LTR transactivation was determined by MAGI cell assay. The reactive oxygen species (ROS) levels and glutathione (GSH) levels were measured. In addition, the protein expressions were assayed by western blotting. KEY FINDINGS: Tat caused a significant decrease in the intracellular glutathione (GSH) levels, a mild increase in the expression of nuclear levels of NF-E2-related factor-2 (Nrf2), a significant increase in the levels of NF-κB (phosphorylation of p65 and IKK) and a significant increase in ROS production. EGCG supplementation significantly improved the changes associated with Tat-induced oxidative stress by increasing nuclear levels of Nrf2, decreasing levels of NF-κB and ROS production. EGCG reversed Tat-mediated AKT activation and AMPK inhibition in MAGI cells. EGCG inhibited Tat-induced LTR transactivation in a dose-dependent manner. SIGNIFICANCE: The results suggest that Nrf2 signaling pathway may be the primary target for prevention of Tat-induced HIV-1 transactivation by EGCG, and EGCG also reduce NF-κB activation by inhibiting AKT signaling pathway and activating AMPK signaling pathway.

MiR-217 is involved in Tat-induced HIV-1 long terminal repeat (LTR) transactivation by down-regulation of SIRT1.

MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression and may contribute to the development and progression of many infective diseases including human immunodeficiency virus 1 (HIV-1) infection. The Tat protein is fundamental to viral gene expression. In this study, our goal was to investigate the regulation of a specific miRNA (known as miR-217) in multinuclear activation of galactosidase indicator (MAGI) cells and explore the mechanisms by which miR-217 influenced Tat-induced HIV-1 transactivation through down-regulation of SIRT1 expression. We showed that miR-217 was up-regulated when Tat was expressed in multinuclear activation of galactosidase indicator cells. Forced expression of "miR-217 mimics" increased Tat-induced LTR transactivation. In addition, miR-217 significantly inhibited SIRT1 protein expression by acting on the 3'-UTR of the SIRT1 mRNA. In turn, the decrease in SIRT1 protein abundance provoked by miR-217 affected two important types of downstream signaling molecules that were regulated by Tat. Lower expression of SIRT1 caused by miR-217 enhanced Tat-induced phosphorylation of IKK and p65-NFkB and also exacerbated the loss of AMPK phosphorylation triggered by Tat. Our results uncover previously unknown links between Tat and a specific host cell miRNA that targets SIRT1. We also demonstrate that this regulatory mechanism impinges on p65-NFkB and AMPK signaling: two important host cell pathways that influence HIV-1 pathogenesis. Our results also suggest that strategies to augment SIRT1 protein expression by down-regulation of miR-217 may have therapeutic benefits to prevent HIV-1 replication.

HDAC1/NFκB pathway is involved in curcumin inhibiting of Tat-mediated long terminal repeat transactivation.

Chromatin remodeling, especially in relation to the transactivator Tat, is an essential event for human immunodeficiency virus-1 (HIV-1) transcription. Curcumin has been shown to suppress pathways linked to HIV-1 replication. We investigated whether curcumin had the potential to inhibit Tat-induced long terminal repeat region (LTR) transactivation. As we shown, curcumin inhibited Tat-induced LTR transcativation, while knockdown of histone deacetylase 1 (HDAC1) by siRNA potentiated Tat-induced HIV-1 transcativation. Curcumin reversed Tat-induced down-regulation of HDAC1 expression in multinuclear activation of galactosidase indicator (MAGI) cells. Treatment with curcumin reversed Tat-induced dissociation of HDAC1 from LTR; and curcumin caused a decline in the binding of p65/NFκB to LTR promoters stimulated by Tat. Curcumin attenuated Tat-induced p65 phosphorylation and IKK phosphorylation. Curcumin reversed Tat-mediated reduction in AMPK activation and downstream acetyl-CoA carboxylase (ACC) activation. Collectively, our data provide new insights into understanding of the molecular mechanisms of curcumin inhibited Tat-regulated transcription, suggesting that targeting AMPK/HDAC1/NFκB pathway could serve as new anti-HIV-1 agents.

Akt/Nox2/NF-κB signaling pathway is involved in Tat-induced HIV-1 long terminal repeat (LTR) transactivation.

Human immunodeficiency virus (HIV) regulatory protein Tat has pro-oxidant property, which might contribute to Tat-induced long terminal repeat region (LTR) transactivation. However, the intracellular mechanisms whereby Tat triggers ROS production, and the relationship between Tat-induced ROS production and LTR transactivation, are still subject to debate. The present study was undertaken to evaluate the specific effects of Tat on nicotinamide adenine denucleotide phosphate (NADPH) oxidase in MAGI cells, and to determine the specific role of NADPH oxidase in Tat-induced LTR transactivation. Application of Tat to MAGI cells caused increases in ROS formation that were prevented by both pharmacologic NADPH oxidase inhibitors and by siRNA Nox2, but not by other inhibitors of pro-oxidant enzymes or siRNA Nox4. Furthermore, inhibition of NADPH oxidase by both pharmacologic NADPH oxidase inhibitors and by siRNA Nox2 attenuated Tat-induced p65 phosphorylation and IKK phosphorylation. Phosphatidylinositol 3-kinase/Akt signaling pathway was involved in Tat-induced NADPH oxidase stimulation. Finally, NADPH oxidase inhibitors or Nox2 siRNA, but not control siRNA, inhibited Tat-induced LTR transactivation. Tat-induced HIV-1 LTR transactivation was inhibited in wortmannin or LY294002 treated cells compared to control cells. Together, these data describe a specific and biologically significant signaling component of the MAGI cells response to Tat, and suggest the PI3K/Akt signaling pathway might originate in part with Tat-induced activation of NADPH oxidase and LTR transactivation.

Nicotinamide phosphoribosyltransferase/sirtuin 1 pathway is involved in human immunodeficiency virus type 1 Tat-mediated long terminal repeat transactivation.

Tat is a multifunctional transactivator encoded by human immunodeficiency virus type 1 (HIV-1). Tat transactivating activity is controlled by nicotinamide adenine nucleotide(+) (NAD(+))-dependent deacetylase sirtuin 1 (SIRT1). Nicotinamide phosphoribosyltransferase (Nampt) is a rate-limiting enzyme in the conversion of nicotinamide into NAD(+), which is crucial for SIRT1 activation. Thus, the effect of Nampt on Tat-regulated SIRT activity was studied in Hela-CD4-beta-gal (MAGI) cells. We demonstrated that Tat caused NAD(+) depletion and inhibited Nampt mRNA and protein expression in MAGI cells. Resveratrol reversed Tat-induced NAD(+) depletion and inhibition of Nampt mRNA and protein expression. Further investigation revealed that Tat-induced inhibition of SIRT1 activity was potentiated in Nampt-knockdown by Nampt siRNA compared to treatment with Tat alone. Nampt siRNA potentiated Tat-induced HIV-1 transactivation in MAGI cells. Altogether, these results indicate that Nampt is critical in the regulation of Tat-induced inhibition of SIRT1 activity and long terminal repeat (LTR) transactivation. Nampt/SIRT1 pathway could be a novel therapeutic tool for the treatment of HIV-1 infection.