Objective:To explore the risk factors for mortality in pediatric acute respiratory distress syndrome (PARDS) requiring extracorporeal membrane oxygenation (ECMO) support.Methods:Clinical data of 109 patients with severe PARDS supported by ECMO, who were hospitalized in 6 ECMO centers in China from September 2012 to February 2020, were retrospectively analyzed. They were divided into survival group and death group according to the prognosis. Chi-square test and rank sum test were used to compare the variables between the two groups, including the demographic data, laboratory examination results, clinical data before and after ECMO, and other supportive treatment. Univariate and multivariate Logistic regression models were used to analyze the prognostic risk factors.Results:In these 109 cases, 54 died and 55 survived. Compared with the survival group, the death group had higher incidences of acute kidney injury (AKI) (48.1% (26/54) vs. 21.8% (12/55) , χ2=8.318, P=0.004) and coagulation dysfunction (22.2% (12/54) vs. 7.3% (4/55) , χ2=4.862, P=0.027), and higher rate of renal replacement therapy (48.1% (26/54) vs. 21.8% (12/55) , χ2=9.694, P=0.008) during ECMO support. Logistic regression analysis showed that continuous renal replacement therapy (CRRT) and AKI were independent risk factors for death in patients with severe PARDS requiring ECMO support ( HR=3.88,95% CI 1.04-14.52, HR=4.84,95% CI 1.21-19.46, both P<0.05). Conclusion:AKI and CRRT are independent risk factors for predicting mortality in patients with severe PARDS requiring ECMO support.

Objective: To investigate the effect of over-expression of paired related homoeobox 1 (PRRX1) on apoptosis of hepatocellular carcinoma SMMC7721 cells, and to explore its detailed mechanism. Methods： ：Lentivirus-mediated PRRX1 over-expression vector (pGMLV-PRRX1) and empty vector (Vector) were respectively infected into SMMC7721 cells, and the mRNA and protein expression levels of PRRX1 in infected cells were detected by qPCR and WB. The effect of PRRX1 over-expression on the cell proliferation and apoptosis of SMMC7721 cells were assessed by CCK-8 assay and Annexin-V FITC/PI double staining flow cytometry assay, respectively. The change of mitochondrial membrane potential of SMMC7721 cells was detected by mitochondrial membrane potential assay kit (JC-10 staining assay). The enzymatic activities of caspase-8 and caspase-9 in infected cells were detected by using caspase activity assay kit (spectrophotometric method). The protein expression levels of p53, Bcl-2, Bax, Fas, Cleaved-caspase-3 and Cty C expressed in mitochondria and cytosol were evaluated by WB. Results： ：PRRX1 over-expressed SMMC7721 cell line was successfully constructed, and the protein and mRNA expression levels of PRRX1 were significantly increased in lentivirus infected cells (all P< 0.01). Compared with control group and vector group, over-expression of PRRX1 significantly inhibited cell proliferation, weakened mitochondrial membrane potential, but increased the rate of apoptosis, elevated the shear level of caspase-3, promoted the release of Cyt C protein from mitochondrial into cytosol and increased the enzymatic activity of caspase-9 (all P<0.05 or P<0.01). In addition, over-expression of PRRX1 also promoted the protein expressions of p53 and Bax but inhibited the protein expression of Bcl-2 (all P< 0.05 or P<0.01); however,it had no significant effect on the expression of Fas protein and the enzymatic activity of caspase-8 (all P＞ 0.05). Conclusion： ：Over-expression of PRRX1 induces apoptosis in hepatocellular carcinoma SMMC7721 cells, which may be related to the activation of p53-mediated mitochondrial apoptotic pathway

Objective To explore the mechanism of microRNA (miRNA, miR)-155 in the rejection after liver transplantation in rats. Methods The rats were divided into two groups. In the xenograft model group (rejection group, n=10),the donors were male Lewis rats and the recipients were male BN rats.In the allograft model group(control group, n=10),both the donors and recipients were male Lewis rats.The rat models with orthotopic liver transplantation were established by two-cuff technique in two groups. At postoperative 7 d, the animals were sacrificed for the collection of blood and liver tissue samples. The serum levels of alanine aminotransferase (ALT), total bilirubin (TB), and cytokines of interleukin (IL)-2, IL-4, interferon (IFN)-γ were quantitatively measured. The pathological changes of liver tissues were observed under light microscope. In each group, three liver tissue samples were prepared and subject to high-throughput sequencing. The miRNAs related to rejection were identified for bioinformatics analysis to predict and analyze relevant signaling pathways and genes. Results In the rejection group, the serum levels of ALT and TB were significantly higher than those in the control group (both P<0.01). Compared with the control group, the levels of IL-2 and IFN-γ were considerably up-regulated (both P<0.01), whereas the level of IL-4 was dramatically down-regulated (P<0.01). Pathological examination demonstrated that more evident rejections were observed in the rejection group than the control group. High-throughput sequencing revealed that the expression level of miR-155 was significantly up-regulated in the rejection group, which was 5.89 times of that in the control group. Bioinformatics analysis demonstrated that up-regulation of miR-155 was associated with the mammalian target of rapamycin (mTOR), mitogen-activated protein kinase (MAPK) and T cell receptor signaling pathways. The genes which were probably responsible for regulation included the yeast autophagy related gene 1(ATG1) and its homologous gene ULK2, insulin-like growth factor-1 (Igf-1) and G protein-coupled receptor regulatory gene(Arrb1),etc.Conclusions miR-155 might promote the incidence and progression of rejection after liver transplantation in rats. The involved signaling pathways probably include the mTOR, MAPK signaling pathway and T cell receptor signaling pathway.ATG1,ULK2,Igf-1,and Arrb1 genes may participate in this process.

Objective: To investigate the role of bone marrow mesenchymal stem cells (BMSCs) with CTLA4Ig and CD40LIg gene modification in rejection reaction after liver transplantation in rats and possible mechanisms. Methods: The modified Kamada’s two-cuff technique was used to establish a Lewis-BN rat model of orthotopic liver transplantation, and a total of 75 rats were randomly divided into groups A, B, C, D, and E, with 15 rats in each group. The rats in group A (control group) were given infusion of isotonic saline via the portal vein during liver transplantation, those in group B (BMSC group) were given infusion of BMSCs via the portal vein during liver transplantation, those in group C (BMSCs with CTLA4Ig gene modification) were given infusion of BMSCs carrying the CTLA4Ig gene via the portal vein during liver transplantation, those in group D (BMSCs with CD40LIg gene modification) were given infusion of BMSCs carrying the CD40LIg gene via the portal vein during liver transplantation, and those in group E (BMSCs with CTLA4Ig and CD40LIg gene modification) were given infusion of BMSCs carrying CTLA4Ig and CD40LIg gene modification via the portal vein during liver transplantation. Postoperative survival and change in liver function were observed. HE staining was used to observe the pathomorphological changes of the graft liver, and ELISA was used to measure the levels of interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-10 (IL-10), and interferon-γ (IFN-γ) in peripheral blood. A one-way analysis of variance was used for comparison of means of multiple samples, and the Kaplan-Meier survival curve analysis was used for comparison of survival rates between multiple groups. Results: Group E had a significantly longer survival time after surgery than groups A, B, C, and D (P < 0.05), groups C and D had a significantly longer survival time than groups A and B (P < 0.05), and there was no significant difference between groups C and D (P > 0.05). On day 10 after surgery, group A had significantly higher levels of alanine aminotransferase and total bilirubin than the other four groups (P < 0.05). HE staining showed severe rejection reaction in group A, moderate rejection reaction in group B, and mild rejection reaction in groups C and D; pathological examination showed no marked rejection reaction in group E. Group A had significant increases in the levels of IL-2 and IFN-γ and significant reductions in the levels of IL-4 and IL-10 after surgery compared with the other four groups (all P < 0.05). Conclusion Infusion of BMSCs with modification of both CTLA4Ig and CD40LIg genes can significantly inhibit acute rejection reaction after liver transplantation in rats and effectively prolong the survival time of the graft liver, with a better effect than infusion of BMSCs alone or BMSCs with modification of CTLA4Ig or CD40LIg gene.

Objective: To investigate the efficacy and safety of the new investigational drug pegylated interferon α-2b (Peg-IFN-α-2b) (Y shape, 40 kD) injection (180 µg/week) combined with ribavirin in the treatment of patients with genotype 1/6 chronic hepatitis C (CHC), with standard-dose Peg-IFN-α-2a combined with ribavirin as a positive control. Methods: A multicenter, randomized, open-label, and positive-controlled phase III clinical trial was performed. Eligible patients with genotype 1/6 CHC were screened out and randomly divided into Peg-IFN-α-2b(Y shape, 40kD) group and Peg-IFN-α-2a group at a ratio of 2:1. The patients in both groups were given oral ribavirin for 48 weeks in addition and then followed up for 24 weeks after drug withdrawal. Abbott Real Time HCV Genotype II was used to determine HCV genotype, and Cobas TaqMan quantitative real-time PCR was used to measure HCV RNA level at 0, 4, 12, 24, 48, and 72 weeks. Adverse events were recorded in detail. The primary efficacy endpoint was sustained virological response (SVR), and a non-inferiority test was also performed. Results: A total of 561 patients with genotype 1/6 CHC were enrolled, among whom 529 received treatment; 90.9% of these patients had genotype 1 CHC. The data of the full analysis set showed that SVR rate was 69.80% (95% CI 65.00%-74.60%) in the trial group and 74.16% (95% CI 67.73%-80.59%) in the control group (P = 0.297 0). The data of the per protocol set (PPS) showed that SVR rate was 80.63% (95% CI 76.04%-85.23%) in the trial group and 81.33% (95% CI 75.10%-87.57%) in the control group (P = 0.849 8), and the 95% CI of rate difference conformed to the non-inferiority standard. The analysis of the PPS population showed that of all subjects, 47.9% achieved rapid virologic response, with a positive predictive value of 93.8%. The incidence rate of adverse events was 96.30% in the trial group and 94.94% in the control group, and the incidence rate of serious adverse events was 5.13% in the trail group and 5.06% in the control group. Conclusion In the regimen of Peg-IFN-α combined with ribavirin for the treatment of genotype 1/6 CHC, the new investigational drug Peg-IFN-α-2b(Y shape, 40 kD) has comparable clinical effect and safety to the control drug Peg-IFN-α-2a.

Objective To explore the effects of the Chinese medicine, modified Erchen decoction, on the serum lipid spectrum of ApoE-/-mice, and to explore its possible anti-atherosclerotic mechanism.Methods Forty-four male 7-8-week old ApoE-/-mice were used in this experiment.ApoE-/-mouse models of atherosclerosis were generated by high-cholesterol diet for 4 weeks.And then, they were given simvastatin or modified Erchen decoction by gavage.The body weight of mice was recorded every week, The mice were sacrificed after treated with the drugs for 8 weeks continuously, and the plasma lipid was determined by enzymatic method.The aortic valves and arches were stained with oil red O to depict atherosclerotic plaques and liver structural changes of the mice were examined by pathology.Results Modified Erchen decoction lowered plasma lipid ( including TCHOL and LDL-C ) significantly ( P<0.01 ) .The body weight was increased in the mice of all groups, but it was more pronounced in the mice of model group than in the blank and modified Erchen decoction groups.The serum CHOL and LDL-C levels were significantly lowered in the modified Erchen decoction group (P<0.01).The area of atherosclerotic plaques in the aortic wall was significantly reduced in the mice of modified Erchen decoction group as shown by oil red O staining.The pathological changes of hepatocytes were less severe and the structure of hepatic lobules was better preserved in the mice of modified Erchen decoction group.Conclusions The Chinese medicin modified Erchen decoction can effectively reduce serum lipids, regulate lipid metabolism, and ameliorate the process of atherosclerosis in ApoE-/-mice.

AIM:To investigate the expression of long non-coding RNA maternally expressed gene 3 (MEG3) in colorectal cancer ( CRC) cells, and to observe the effect of MEG 3 on the invasion and migration of CRC cells .METH-ODS:The levels of MEG3 in human normal colon cell NCM 460 and CRC cells SW48 and LoVo were detected by real-time PCR.MEG3 was over-expressed by plasmid transfection , and the effects of MEG 3 on the invasion and migration of SW 48 and LoVo cells were analyzed by Transwell assay and wound healing assay .The expression of matrix metalloproteinase ( MMP) family proteins was determined by Western blotting .RESULTS:The level of MEG3 was down-regulated in CRC cells compared with normal colon cell NCM 460.The invasion and migration of CRC cells were reduced after MEG 3 over-ex-pression.Transwell invasion and migration assays showed that the numbers of transmembrane SW 48 and LoVo cells were smaller in MEG3 over-expression group than control group (P<0.05).The cell spaces were broader after MEG3 over-ex-pression in the wound healing assay , indicating that MEG3 over-expression inhibited the mobility of CRC cells .Meanwhile, over-expression of MEG3 reduced the expression of MMP-2 and MMP-9, and elevated the expression of tissue inhibitor of metalloproteinase-2 (TIMP-2).CONCLUSION:The expression of MEG3 is down-regulated in CRC cells.Over-expres-sion of MEG3 inhibits the invasion and migration of CRC cells .TIMP-2, MMP-2 and MMP-9 might play an important role in this regulation .

To investigate the effects and mechanism of bone marrow mesenchymal stem cell (MSC)modified by cytotoxicity T lymphocyte-associated antigen 4-immunoglobulin (CTLA4-Ig)gene on the rejection of orthotopic liver transplantation (OLT)in rats. Methods MSC was infected with recombinant adenoviruses (Ad)5 containing CTLA4-Ig gene. After recombinant Ad-5 containing CTLA4-Ig infected MSC for 72 h,the total proteins were extracted. The protein expression of CTLA4-Ig was assessed by Western-blot.The suppression to lymphocyte proliferation by MSC and transgenic MSC were tested by cell counting kit (CCK)-8 analysis. Forty models of acute rejection after OLT in rats were established by modified Kamada’s two-cuff technique,with male Lewis and BN rats serving as liver donors and recipients respectively. Forty recipient rats were randomly divided into 4 groups with 10 rats in each group including control group (group A, only saline solution was injected into portal venous during transplantation),MSC group (group B,MSC was injected into portal venous during transplantation),transgenic MSC group (group C,transgenic MSC was injected into portal venous during transplantation),immunosuppressant group [group D,saline solution was injected into portal venous during transplantation,and ciclosporin (CsA)was administered intramuscularly at a dose of 1.5 mg /(kg·d) for 8 days]. On the 9 th day after operation,5 rats were killed randomly in every group,then the levels of interleukin (IL)-2,interferon (IFN)-γ,IL-4 in peripheral blood were measured and the pathological changes and rejection expression of liver tissues were observed by light microscope. The survival condition of other 5 rats in 4 groups was observed. Results After recombinant Ad-5 containing CTLA4-Ig infected MSC for 72 h,the protein expression of CTLA4-Ig gene in MSC infected with Ad5-CTLA4-Ig could be detected by Western-blot.When the ratios of MSC∶peripheral blood monouclear cell (PBMC)were 1∶10 and 1∶20,the rates of suppression to lymphocyte proliferation were 85.60% and 76.69% respectively.When the ratios of transgenic MSC∶PBMC were 1∶10 and 1∶20,the rates of suppression to lymphocyte proliferation were 90.50% and 84.20% respectively. Compared with MSC,MSC infected with Ad5-CTLA4-Ig had stronger effect on suppression to lymphocyte proliferation (P <0.05 ). The survival time after liver transplantation of rats in group A,B,C,D was (13 ±3),(41 ±6),(90 ±15),(102 ±18)d respectively.There were significant differences among group A,B,C (P<0.05 )and there was no significant difference between group C and D (P>0.05 ). Compared with group A,the serum levels of IL-4 in group B and C were significantly higher (P<0.05 ). The serum levels of IL-4 in group C were significantly higher than that in group B (P<0.05 ). There was no significant difference in the serum levels of IL-4 between group C and D (P>0.05 ). Compared with group A,the serum levels of IL-2 and IFN-γin group B and C were lower significantly (P<0.05 ). The serum levels of IL-2 and IFN-γin group C were significantly lower than those in group B (both in P<0.05 ). There were no significant differences in the serum levels of IL-2 and IFN-γbetween group C and D (P>0.05 ). The pathological result of liver tissues of rats showed that the grafts of group A developed severe rejection,and the grafts of group B developed moderate rejection. And the grafts of group C and D developed slight rejection. Conclusions MSC infected with recombinant Ad5-CTLA4-Ig can inhabit the rejection in liver transplantation,and the effect is superior to MSC alone.

AIM:To investigate the formation of membrane pore in PC 12 cells induced by exogenous adenosine triphosphate ( ATP) and to identify the key molecular targets .METHODS:PC12 cells were treated with different concen-trations of ATP to establish the injury model .The morphological change was observed under an inverted phase -contrast mi-croscope.The viability of the PC12 cells was measured by CCK-8 assay.Fluorescent dye YO-PRO-1 was used to detect the membrane permeability.The expression of P2X7 receptor and pannexin 1 (Panx1) at mRNA and protein levels was as-sessed by real-time PCR and Western blotting .RESULTS:After exposed to ATP (1 mmol/L, 3 mmol/L and 5 mmol/L) for 3 h, the PC12 cells became edematous , and the number of adherent cells decreased gradually in a dose-dependent man-ner .The cell viabilities in 3 mmol/L ATP group and 5 mmol/L ATP group were significantly decreased compared with con-trol group (P<0.05).YO-PRO-1 uptake in the PC12 cells exposed to ATP (0, 1, 3 and 5 mmol/L) for 15 min, 30 min and 60 min increased in a dose-dependent and time-dependent manner .The cell viability increased and the intracellular fluorescence intensity induced by ATP were significantly antagonized in brilliant blue G ( a P2X7 receptor inhibitor ) pre-treatment group (P<0.05), whereas it did not change in carbenoxolone (a Panx1 inhibitor) pretreatment group (P>0. 05).The expression of P2X7 receptor at mRNA and protein levels was significantly increased (P<0.05), but the expres-sion of Panx1 was not changed ( P>0.05) when PC12 cells were exposed to ATP for 3 h.CONCLUSION:Extracellular ATP at high concentration may induce membrane pore formation with the expression and activation of P 2X7 receptor in PC12 cells.

Objective To observe the effects of platelet-rich plasma (PRP) gel in the treatment of [Ⅳ stage pressure ulcers.Methods A total of 12 patients with stage Ⅳ pressure ulcers were treated with PRP gel from December 2012 to December 2013 in our department.The PRP gel was formed by autologous PRP mixed with thrombin and calcium chloride.The PRP gel was applied to the wound after dressing.Time intervals between dressing were 4 days.Overall pressure ulcer improvement was assessed every week until complete wound healing.Results All stage Ⅳ pressure ulcer wound infection was controlled and fresh granulation tissue was found after 2 times of PRP gel covering.The time of ulcer wound healing were 6-10 weeks (mean 8 weeks).The times of changing the PRP gel were 16.No patients experienced adverse reactions during treatment.Conclusions PRP gel can effectively control the pressure ulcer infection and promote ulcer wound healing.