Analysis of factors related to recanalization of portal vein thrombosis in liver cirrhosis: a retrospective cohort study.

BACKGROUND: Portal vein thrombosis (PVT) is a common complication of liver cirrhosis, yet there are fewer studies about predictors of PVT recanalization. We aimed to further explore the predictors of recanalization in cirrhotic PVT to facilitate accurate prediction of patients' clinical status and timely initiation of appropriate treatment and interventions. To further investigate the benefits and risks of anticoagulant therapy in cirrhotic PVT patients. METHODS: A retrospective cohort study of patients with cirrhotic PVT in our hospital between January 2016 and December 2022, The primary endpoint was to analyze predictors of PVT recanalization by COX regression. Others included bleeding rate, liver function, and mortality. RESULTS: This study included a total of 82 patients, with 30 in the recanalization group and 52 in the non-recanalization group. Anticoagulation therapy was the only independent protective factor for portal vein thrombosis recanalization and the independent risk factors included massive ascites, history of splenectomy, Child-Pugh B/C class, and main trunk width of the portal vein. Anticoagulation therapy was associated with a significantly higher rate of PVT recanalization (75.9% vs. 20%, log-rank P < 0.001) and a lower rate of PVT progression (6.9% vs. 54.7%, log-rank P = 0.002). There was no significant difference between different anticoagulation regimens for PVT recanalization. Anticoagulation therapy did not increase the incidence of bleeding complications(P = 0.407). At the end of the study follow-up, Child-Pugh classification, MELD score, and albumin level were better in the anticoagulation group than in the non-anticoagulation group. There was no significant difference in 2-year survival between the two groups. CONCLUSION: Anticoagulation, massive ascites, history of splenectomy, Child-Pugh B/C class, and main portal vein width were associated with portal vein thrombosis recanalization. Anticoagulation may increase the rate of PVT recanalization and decrease the rate of PVT progression without increasing the rate of bleeding. Anticoagulation may be beneficial in improving liver function in patients with PVT in cirrhosis.

Flap endonuclease 1 silencing is associated with increasing the cisplatin sensitivity of SGC­7901 gastric cancer cells.

Flap endonuclease 1 (FEN1), which is key in DNA replication and repair, has been demonstrated to be intimately involved in the development and progression of cancer. Our previous study determined that the downregulation of FEN1 can suppress the proliferation of, and induce apoptosis in, gastric cancer SGC­7901 cells. In addition, several FEN1 inhibitors have been identified to increase sensitisation to DNA injury agents. These results may provide a promising treatment method to enhance the traditional chemotherapeutics used for the treatment of gastric cancer. Thus, the aim of the present study was to determine the role of FEN1 in the chemosensitivity of SGC­7901 cells. The protein expression levels of FEN1 in cisplatin (CDDP)­treated SGC­7901 cells were detected using western blot analysis. FEN1 was silenced via specific FEN1­targeted small interfering RNAs (siRNA). The survival and apoptotic rates of the SGC­7901 cells were assessed using an MTT assay and flow cytometry, respectively. Relevant apoptotic factors were detected using western blotting. The results showed that the expression of FEN1 was significantly induced by CDDP in a dose­ and time­dependent manner. The targeting of FEN1 in SGC­7901 cells, in combination with CDDP treatment, significantly inhibited their proliferation and effectively increased their apoptotic rate. In addition, in the cells targeted with FEN1­siRNA and exposed to CDDP, the levels of Bcl­2­associated X protein were significantly increased, whereas the expression levels of Bcl­2 and Bcl­extra large were effectively decreased, compared with the cells exposed to negative control­siRNA and CDDP. These results suggest a potential chemotherapeutic target, which exhibits enhanced sensitivity to CDDP following FEN1 silencing in SGC­7901 cells via decreased survival and increased apoptosis.

Probiotics for the prevention of antibiotic-associated diarrhoea in older patients: a systematic review.

OBJECTIVE: Here, we evaluated the efficacy of probiotic interventions in prevention of antibiotic-associated diarrhoea (AAD) and Clostridium difficile diarrhoea (CDD) in older patients. METHODS: PubMed, Embase, CENTRAL, CINAHL, and Web of Science were comprehensively searched from their dates of inception to May 2014. Only randomised controlled trials reporting data on probiotics including Lactobacillus, Bifidobacterium, Saccharomyces, Streptococcus, Enterococcus, and Bacillus-alone or in combination-versus placebo or absence of treatment in older patients (age ≥65 years) were included. Risk ratios (RRs) for AAD and CDD relative to placebo or absence of treatment were estimated. RESULTS: Six trials with a total of 3562 patients were included. Only one trial showed that Bacillus licheniformis was effective for preventing AAD in older patients. However, there was no preventive effect for AAD and CDD with Lactobacillus acidophilus (Florajen), Lactobacillus casei Shirota, Saccharomyces cerevisiae (boulardii) lyo, mixture of Lactobacillus acidophilus and Bifidobacterium bifidum (Cultech strains), and mixture of Lactobacillus acidophilus CUL60, CUL21, Bifidobacterium bifidum CUL20 and B. lactis CUL34. CONCLUSIONS: Our findings indicate that probiotics may not reduce the risk of AAD and CDD in older patients. However, with current published data, it is difficult to draw concrete conclusions. To confirm these findings, sample sizes, multi-centre, double-blind studies that consider factors such as probiotic strains and types of antibiotics are required.

Flap endonuclease 1 is a promising candidate biomarker in gastric cancer and is involved in cell proliferation and apoptosis.

As a DNA repair protein, flap endonuclease 1 (FEN1), a structure-specific 5' nuclease, plays pivotal roles in the maturation of Okazaki fragments, long-patch base excision repair, restarting of stalled replication forks and telomere maintenance. FEN1 possesses 5' endonuclease, 5' exonuclease and gap-endonuclease activities, which render it an essential node in maintaining genome fidelity. The aim of this study was to investigate the association between the expression level of FEN1 and gastric cancer and to explore the role of FEN1 in carcinogenesis and the progression of gastric cancer. The mRNA and protein expression of FEN1 in 42 matched pairs of human gastric tumor tissues and corresponding normal tissues were measured by semiquantitative reverse transcription-PCR and immunohistochemical staining. FEN1 expression was downregulated in the SGC-7901 gastric cancer cells following transfection with siRNA targeting the FEN1 gene. Western blot analysis was used to evaluate the protein expression of FEN1 in SGC-7901 human gastric cancer cells in order to verify the transfection efficiency of FEN1 siRNA. Moreover, cell proliferation was analyzed by MTS assay. The apoptosis of the cells was determined by flow cytometry. Our results revealed that FEN1 was overexpressed in gastric cancer in comparison to the corresponding normal gastric tissues (P<0.01). We further confirmed that FEN1 expression has a positive correlation with the degree of differentiation (P=0.027), lymphatic metastasis (P=0.001), tumor size (P=0.026) and TNM stage (P=0.020) of gastric cancer. A high FEN1 expression in SGC-7901 cells can be effectively downregulated by siRNA constructed to target the FEN1 gene. Moreover, the inhibition of FEN1 expression suppressed the proliferation and induced the apoptosis of SGC-7901 cells. Taken together, our results indicate that FEN1 may be a promising biomarker for the diagnosis of gastric cancer and individual therapy.

[Effects of pshRNA-DNMT1 on the proliferation and apoptosis of gastric cancer: experiments in vitro and in vivo].

OBJECTIVE: To investigate the effects of pshRNA-DNMT1 on the proliferation and apoptosis of gastric cancer. METHODS: Recombinant eukaryotic expression plasmid pshRNA-DNMT1 containing the sequence of the gene of DMNT1 that methylates the specific pyrimidine residue in the DNA promoter region was constructed. Human gastric cells of the line AGS were cultured and transfected with pshRNA-DNMT1. Western blotting was used to detect the protein expression of DNMT1 of the AGS cells, and RT-PCR was used to detect the mRNA expression of DNMT1 of the AGS cells. MTT method was used to dynamically monitor the surviving cells and the cell apoptotic was observed by electron microscopy and TUNEL method. Forty nude the mice were inoculated with suspension of AGS cells. When the tumor reached the size of 5 - 6 mm in diameter the mice were randomly divided into 5 equal groups to be injected intravenously with PBS, liposome, pTZU6 + 1, pshRNA-DNMT1 of medium dose, and pshRNA-DNBMT1 of large dose for 4 times with an interval of 3 days. The tumor size was measured every day. Three days after the last injection the mice were killed and the tumors were taken out to undergo light and electron microscopy and TUNEL method to detect the cell apoptosis. Immunohistochemistry was used to detect the proliferating cell nuclear antigen (PCNA) of the cells. RESULTS: The protein and mRNA expression levels of DNMT1 in the cultured AGS cells 24, 48, and 72 hours after transfection of the pshRNA-DNMT1 group were all lower than those of the control group. The numbers of surviving AGS cells of the pshRNA-DNMT1 group became significantly gradually lower than those of the liposome and pTZU6 + 1 groups since 24 hours after transfection (all P < 0.05). The apoptotic rate of AGS cells in the pshRNA-DNMT1 group was 34.78% +/- 0.52%, significantly higher than those of the liposome and pTZU6 + 1 groups (4.86% +/- 0.17% and 5.12% +/- 0.76% respectively, both P < 0.05). The subcutaneous tumors of the mice of the PGS, liposome, and pTZU6 + 1 groups augmented along with time without significant differences among these 3 groups (all P > 0.05). The tumor of the 2 pshDNMT1 groups began to augment since the 5(th) day and began to be reduced in size since the 10(th) day in comparison with the other 3 groups (all P < 0.05), and the tumor size of the pshRNA-DNMT1 (large dose) group was significantly smaller than that of the pshRNA-DNMT1 (medium dose) group 15 days after the injection (P < 0.05). The rates of cell apoptosis of the pshRNA-DNMT1 (large dose) and pshRNA-DNMT1 (medium dose) groups were both significantly higher than those of the other 3 groups (all P < 0.05) and with a sufficient difference between these 2 pshRNA-DNMT1 groups (P < 0.05). PCNA analysis showed that the proliferation activity of the cells in the pshRNA-DNMT1 groups was significantly suppressed. CONCLUSION: The recombinant plasmid pshRNA-NMT1 effectively and specifically inhibits the expression of the gene DNMT1, thus inhibiting the proliferation and inducing the apoptosis of gastric cancer cells.

[Effects of dnmt1 gene silencing on cell cycle, proliferation, and apoptosis of gastric cancer cell line AGS].

BACKGROUND & OBJECTIVE: Dnmt1, a major DNA methyltransferase gene, is highly expressed in many cancers and lowly expressed in normal adult cells, therefore, its overexpression is closely related to tumorigenesis. This study was to assess effects of dnmt1 gene silencing on cell cycle, proliferation, and apoptosis of gastric cancer cell line AGS. METHODS: The eukaryotic expression plasmid pshRNA-dnmt1, containing the sequence of short hairpin RNA (shRNA) targeting dnmt1, was constructed and transfected into AGS cells. PBS-treated cells and pTZU6+1-transfected cells were set as control. The expression levels of dnmt1 were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. Cell cycle was analyzed by flow cytometry; cell survival was analyzed by MTT assay; cell apoptosis was evaluated by AO/EB double staining, electron microscopy, and TUNEL. RESULTS: pshRNA-dnmt1 targeting dnmt1 was successfully constructed, and confirmed by sequencing. Relative to control, 24, 48, and 72 h after transfection of pshRNA-dnmt1, the inhibitory rates of dnmt1 protein levels in AGS cells were 28.24%, 68.54%, and 81.47%, respectively, and those of dnmt1 mRNA levels were 21.63%, 52.97%, and 72.06%, respectively. The growth of AGS cells was suppressed 72 h after transfection: S phase cells were reduced from (36.58+/-1.76)% to (18.54+/-6.59)% (P<0.05), and G(2)/M phase cells were increased from (6.18+/-0.32)% to (18.53+/-1.42)% (P<0.05). Cell survival rates were 79.49%, 51.63%, and 39.16%, respectively, 24, 48, and 72 h after transfection of pshRNA-dnmt1. A lot of apoptotic and necrosis cells were observed after transfection. CONCLUSIONS: The recombinant plasmid pshRNA-dnmt1 can efficiently and specifically inhibit the expression of dnmt1 gene and the proliferation of AGS cells, and induce cell apoptosis. It provides evidence for gene therapy of human cancers.

Cloning and sequence analysis of gene oipA encoding an outer membrane protein of human Helicobacter pylori.

AIM: To construct a recombinant E. coli strain that would highly express the proinflammatory outer membrane protein of human Helicobacter pylori (H pylori). METHODS: The oipA DNA was amplified by PCR, inserted into pET-32a, and transformed into Top10 E. coli strain. This recombinant plasmid of Top10 was sent out for nucleotide sequence analysis. Finally this sequence AF479754 was compared with HP0638 and JHP0581. RESULTS: The sequence of the aim gene was obtained. It had 924 base pairs. The identity was 95.32% against HP0638, 95.02% against JHP0581, which was higher than the identity between HP0638 and JHP0581. CONCLUSION: Although the aim gene was obtained, but it was different from the published sequence of GenBank. It is not clear what makes this difference. Maybe it is because different strain was used or because there were some variations. So more researches are required to prove it.