Ginsenoside Rg_3 inhibit hepatocellular carcinoma growth via intrinsic apoptotic pathway

AIM:To investigate the anti-tumor function of ginsenoside Rg3 on hepatocellular carcinoma(HCC) in vitro and in vivo,and its mechanism.METHODS:Hep1-6 and HepG2 cells were treated by Rg3 in different concentrations(0,50,100 and 200 μg/mL) in vitro.After incubation for 0,6,12,24 and 48 h,cell viability was measured by 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide assay.Apoptosis was identified by terminal deoxynucleotidyl transferasemediated dUTP-biotin nick end labeling.Caspase-3 activity was measured by chromophore p-nitroanilide and flow cytometry.Bcl-2 family proteins were ascertained by Western-blotting.Mitochondria membrane potentialwas detected by 5,5',6' 6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide.Forty liver tumor-bearing C57Bl6 mice were divided randomly into 4 groups for intra-tumor injection of saline,ginsenoside Rg3,cyclophosphamide(CTX) and ginsenoside Rg3 + CTX combination.RESULTS:The survival time was followed up to 102 d.The mice in the Rg3 + CTX group showed significant increased survival time compared with those in the control group(P < 0.05).Rg3 could inhibit HCC cell proliferation and induce cell apoptosis in vitro in the concentration and time dependent manner.It also induced mitochondria membrane potential to decrease.Caspase-3 activation can be blocked by the inhibitor z-DEVD-FMK.Bax was up-regulated while Bcl-2 and Bcl-XL were down-regulated after Rg3 treatment.CONCLUSION:Our data suggested that Rg3 alone or combined with CTX inhibited tumor growth in vivo and prolonged mouse survival time by inducing HCC cell apoptosis via intrinsic pathway by expression alterations of Bcl-2 family proteins.

Application of metagenomics in the human gut microbiome

There are more than 1000 microbial species living in the complex human intestine.The gut microbial community plays an important role in protecting the host against pathogenic microbes,modulating immunity,regulating metabolic processes,and is even regarded as an endocrine organ.However,traditional culture methods are very limited for identifying microbes.With the application of molecular biologic technology in the field of the intestinal microbiome,especially metagenomic sequencing of the next-generation sequencing technology,progress has been made in the study of the human intestinal microbiome.Metagenomics can be used to study intestinal microbiome diversity and dysbiosis,as well as its relationship to health and disease.Moreover,functional metagenomics can identify novel functional genes,microbial pathways,antibiotic resistance genes,functional dysbiosis of the intestinal microbiome,and determine interactions and co-evolution between microbiota and host,though there are still some limitations.Metatranscriptomics,metaproteomics and metabolomics represent enormous complements to the understanding of the human gut microbiome.This review aims to demonstrate that metagenomics can be a powerful tool in studying the human gut microbiome with encouraging prospects.The limitations of metagenomics to be overcome are also discussed.Metatranscriptomics,metaproteomics and metabolomics in relation to the study of the human gut microbiome are also briefly discussed.

Gut microbial dysbiosis associates hepatocellular carcinoma via the gut-liver axis

Background: Hepatocellular carcinoma(HCC) is one of the most common malignancies in the world. Gut microbiota has been demonstrated to play a critical role in liver inflammation, chronic fibrosis, liver cirrhosis, and HCC development through the gut-liver axis. Data sources: Recently there have been several innovative studies investigating gut microbial dysbiosismediated enhancement of HCC through the gut-liver axis. Literatures from January 1998 to January 2018 were searched in the Pub Med database using the keywords "gut microbiota" and "hepatocellular carcinoma" or "liver cancer", and the results of experimental and clinical studies were analyzed. Results: Gut microbial dysbiosis accompanies the progression of alcoholic liver disease, non-alcoholic fatty liver disease and liver cirrhosis, and promotes HCC progression in an experimental mouse model. The immune system and key factors such as Toll-like receptor 4 are involved in the process. There is evidence for gut microbial dysbiosis in hepatitis virus-related HCC patients. Conclusions: Gut microbial dysbiosis is closely associated with hepatic inflammation disease and HCC through the gut-liver axis. With the enhanced understanding of the interactions between gut microbiota and liver through the gut-liver axis, new treatment strategies for HCC are being developed.

Optimal immunosuppressor induces stable gut microbiota after liver transplantation

AIM To study the influence of different doses of tacrolimus(FK506)on gut microbiota after liver transplantation(LT)in rats.METHODS Specific pathogen-free Brown Norway(BN)rats and Lewis rats were separated into five groups:(1)Tolerance group(BN-BN LT,n=8);(2)rejection group(Lewis-BN LT,n=8);(3)high dosage FK506(FK506-H)group(Lewis-BN LT,n=8);(4)middle dosage FK506(FK506-M)group(Lewis-BN LT,n=8);and(5)low dosage FK506(FK506-L)group(LewisBN LT,n=8).FK506 was administered to recipients at a dose of 1.0 mg/kg,0.5 mg/kg,and 0.1 mg/kg body weight for 29 d after LT to the FK506-H,FK506-M,and FK506-L groups,respectively.On the 30^(th) day after LT,all rats were sampled and euthanized.Blood samples were harvested for liver function and plasma endotoxin testing.Hepatic graft and ileocecal tissues were collected for histopathology observation.Ileocecal contents were used for DNA extraction,Real-time quantitative polymerase chain reaction(RT-PCR)and digital processing of denaturing gradient gel electrophoresis(DGGE)profiles and analysis.RESULTS Compared to the FK506-H and FK506-L groups,FK506-M was optimal for maintaining immunosuppression and inducing normal graft function;the FK506-M maintained gut barrier integrity and low plasma endotoxin levels;furthermore,DGGE results showed that FK506-M induced stable gut microbiota.Diversity analysis indicated that FK506-M increased species richness and rare species abundance,and cluster analysis confirmed the stable gut microbiota induced by FK506-M.Phylogenetic tree analysis identified crucial bacteria associated with FK506-M;seven of the nine bacteria that were decreased corresponded to Bacteroidetes,while increased bacteria were of the Bifidobacterium species.FK506-M increased Faecalibacterium prausnitzii and Bifidobacterium spp.and decreased Bacteroides-Prevotella and Enterobacteriaceae,as assessed by RT-PCR,which confirmed the crucial bacterial alterations identified through DGGE.CONCLUSION Compared to the low or high dosage of FK506,an optimal dosage of FK506 induced immunosuppression,normal graft function and stable gut microbiota following LT in rats.The stable gut microbiota presented increased probiotics and decreased potential pathogenic endotoxin-producing bacteria.These findings provide a novel strategy based on gut microbiota for immunosuppressive dosage assessment for recipients following LT.

Chronic bile duct hyperplasia is a chronic graft dysfunction following liver transplantation

AIM: To investigate pathological types and influential factors of chronic graft dysfunction (CGD) following liver transplantation (LT) in rats. METHODS: The whole experiment was divided into three groups: (1) Normal group (n = 12): normal BN rats without any drug or operation; (2) SGT group (syngeneic transplant of BN-BN, n = 12): both donors and recipients were BN rats; and (3) AGT group (allogeneic transplant of LEW-BN, n = 12): Donors were Lewis and recipients were BN rats. In the AGT group, all recipients were subcutaneously injected by Cyclosporin A after LT. Survival time was observed for 1 year. All the dying rats were sampled, biliary tract tissues were performed bacterial culture and liver tissues for histological study. Twenty-one d after LT, 8 rats were selected randomly in each group for sampling. Blood samples from caudal veins were collected for measurements of plasma endotoxin, cytokines and metabonomic analysis, and faeces were analyzed for intestinal microflora. RESULTS: During the surgery of LT, no complications of blood vessels or bile duct happened, and all rats in each group were still alive in the next 2 wk. The long term observation revealed that a total of 8 rats in the SGT and AGT groups died of hepatic graft diseases, 5 rats in which died of chronic bile duct hyperplasia. Compared to the SGT and normal groups, survival ratio of rats significantly decreased in the AGT group (aP < 0.01, bP < 0.001, respectively). Moreover, liver necrosis, liver infection, and severe chronic bile duct hyperplasia were observed in the AGT group by H and E stain. On 21 d after LT, compared with the normal group (25.38 ± 7.09 ng/L) and SGT group (33.12 ± 10.26 ng/L), plasma endotoxin in the AGT group was remarkably increased (142.86 ± 30.85 ng/L) (both P < 0.01). Plasma tumor necrosis factor-α and interleukin-6 were also significantly elevated in the AGT group (593.6 ± 171.67 pg/mL, 323.8 ± 68.30 pg/mL) vs the normal (225.5 ± 72.07 pg/mL, 114.6 ± 36.67 pg/mL) and SGT groups (321.3 ± 88.47 pg/mL, 205.2 ± 53.06 pg/mL) (P < 0.01). Furthermore, Bacterial cultures of bile duct tissues revealed that the rats close to death from the SGT and AGT groups were strongly positive, while those from the normal group were negative. The analysis of intestinal microflora was performed. Compared to the normal group (7.98 ± 0.92, 8.90 ± 1.44) and SGT group (8.51 ± 0.46, 9.43 ± 0.69), the numbers of Enterococcus and Enterobacteria in the AGT group (8.76 ± 1.93, 10.18 ± 1.64) were significantly increased (both aP < 0.01, bP < 0.05, respectively). Meanwhile, compared to the normal group (9.62 ± 1.60, 9.93 ± 1.10) and SGT group (8.95 ± 0.04, 9.02 ± 1.14), the numbers of Bifidobacterium and Lactobacillus in the AGT group (7.83 ± 0.72, 8.87± 0.13) were remarkably reduced (both aP < 0.01, bP < 0.05, respectively). In addition, metabonomics analysis showed that metabolic profiles of plasma in rats in the AGT group were severe deviated from the normal and SGT groups. CONCLUSION: Chronic bile duct hyperplasia is a pathological type of CGD following LT in rats. The mechanism of this kind of CGD is associated with the alterations of inflammation, intestinal barrier function and microflora as well as plasma metabolic profiles.

Diagnostic and prognostic role of neutrophil-to-lymphocyte ratio(NLR) in sepsis

Objective: To evaluate neutrophil-to-lymphocyte ratio(NLR) as diagnostic and prognostic role in sepsis. Methods: It was a prospective, observational study, conducted in Intensive Care Unit of Mianyang Central Hospital , from August 2017 to August 2018. A total of 37 cases of newly diagnosed cases of sepsis were included in the study and 20 healthy adults were taken as controls. According to the mortality within 30 d,patients with sepsis were divided into survival group (n=15) and death group (n=22) . The white blood cell (WBC), neutrophils count (NEU), lymphocyte count (LYM), and NLR in peripheral blood were recorded at 1, 3,5,7 days after admission for patients . Logistic regression analysis was used to evaluate the risk factors for predicting the outcome, and receiver-operating characteristic curve (ROC) was plotted for evaluating the value of these factors on the 30-day prognosis. Results: NLR on day 1 (NLR1) of sepsis was signifcantly higher as compared to controls (P<0.001), with far higher diagnostic efficiency (AUC=0.959) than WBC (AUC=0.788) and equivalent to NEU% (AUC=0.942);WBC and NLR on day 7 (NLR7) is independent risk factors for 30-day mortality of sepsis patients and is helpful to predict the prognosis of sepsis. Conclusion: NLR can be a convenient and useful diagnostic and prognostic marker in sepsis and is of great clinical applicative value for primary hospitals without ability to detect other costly biomarkers and for emergency department.