Objective To investigate the role of Rho kinase (ROCK) in the protective effects of hydrogen on intestinal epithelial barrier function in sepsis. Methods Caco-2 cells were cultured routinely, and divided into 6 groups randomly (n=3):control group (C group), hydrogen-rich medium group (H group), lipopolysaccharide (LPS)-treatment group (L group), hy?drogen+LPS-treatment group (HL group), Rho kinase inhibitor (Y-37632) treatment group (Y group) and Rho kinase inhibi?tor Y-27632+LPS-treatment group (YL group). H group was treated with 0.6 mmol/L hydrogen-rich media. The concentra?tion of LPS and Y-27632 were 50 mg/L and 25μmol/L separately. After the Caco-2 monolayer model was established, the transepithelial electrical resistance (TEER) values were measured regularly. When the TEER value reached 800Ω·cm2, the treatment was administered. Then TEER values were measured at 6 h, 12 h and 24 h, and FITC-dextran permeability was de?tected at 24 h. Cells were seeded on 6-well plates. After cell density reached 80%-90%, treatments were given randomly. The real time-polymerase chain reaction (RT-PCR) was conducted to assess mRNA levels of ZO-1 and ROCK mRNA. ZO-1 and ROCK protein expression levels were detected by Western blot assay. Results Compared with C group, TEER values were elevated in 12 h and 24 h in H group (P<0.05). There were no statistical significances in FITC-dextran permeability,
protein expression levels of ZO-1 and ROCK between C group and H group (P>0.05). TEER values were elevated at 6 h, 12 h and 24 h in Y group (P<0.05). There was no significant difference in FITC-dextran permeability between C group and Y group (P > 0.05). The mRNA expression of ZO-1 increased and mRNA expression of ROCK decreased in Y group (P <0.05). The TEER values reduced at 6 h, 12 h and 24 h in L group. The FITC-dextran permeability increased significantly, mRNA and protein expressions of ZO-1 significantly decreased, mRNA and protein expressions of ROCK significantly in?creased in L group (all P<0.05). Compared with L group, TEER values increased significantly at 6 h, 12 h and 24 h in YL group, FITC-dextran permeability decreased, mRNA expressions of ZO-1 increased, mRNA expressions of ROCK de?creased in YL group (P<0.05). Compared with L group, TEER values increased at 6 h, 12 h and 24 h in HL group, FITC-dextran permeability reduced markedly, protein expressions of ZO-1 increased at each time point, protein expressions of ROCK decreased at each time point in HL group (P<0.05). Conclusion Hydrogen can protect intestinal barrier function against sepsis, ameliorate the integrity and permeability of intestinal epithelium and increase the expressions of intercellular tight junction proteins. The suppression of Rho kinase over-expression induced by LPS may be involved in these protective effects of hydrogen.

Objective To investigate the effect of hydrogen on endotoxin-induced expression of zonula occludens-1 (ZO-1) in human colon epithelial cells (Caco-2 cells).Methods Caco-2 cells were cultured routinely,seeded in Transwell chambers or wells,and randomly divided into 4 groups (n =45 each) using a random number table:control group (group C);hydrogen-rich culture medium group (group H);endotoxin group (group E);hydrogen-rich culture medium + endotoxin group (group HE).The cells were cultured in high-glucose DMEM culture medium in group C.The cells were incubated in hydrogen-rich culture medium containing hydrogen 0.6 mmol/L in group H.The cells were incubated in highglucose DMEM culture medium containing 50 μg/ml lipopolysaccharide in group E.The cells were incubated in hydrogen-rich culture medium containing 50 μg/ml lipopolysaccharide and 0.6 mmol/L hydrogen in group HE.Transepithelial electrical resistance (TEER) was measured before incubation or culture,and at 6,12 and 24 h of incubation or culture.The viability of Caco-2 cells was measured by methyl thiazolyl tetrazolium assay at 24 h of incubation or culture.The expression of ZO-1 mRNA in Caco-2 cells was determined using real-time reverse transcriptase polymerase chain reaction at 6,12 and 24 h of incubation or culture.The distribution of ZO-1 in Caco-2 cells was observed by immunofluorescence at 24 of incubation or culture.Results Compared with group C,TEER was significantly decreased at 6,12 and 24 h of incubation or culture,and the expression of ZO-1 mRNA was significantly down-regulated in E and HE groups (P＜0.05),and no significant change was found in the parameters mentioned above in group H (P＞0.05).Compared with group E,TEER was significantly increased at 6,12 and 24 h of incubation or culture,and the expression of ZO-1 mRNA was significantly up-regulated in group HE (P＜0.05).The distribution of ZO-1 protein in cell membrane became discontinuous,and the distribution of ZO-1 protein in cytoplasm was significantly increased in group E.Compared with group E,the distribution of ZO-1 protein in cell membrane was significantly increased and gradually became continuous,and the distribution of ZO-1 protein in cytoplasm was significantly decreased in group HE.Conclusion The mechanism by which hydrogen reduces the damage to human colon epithelial cell barrier is related to up-regulation of ZO-1 expression and improvement in the redistribution of ZO-1 protein.

Objective:To compare the treatment outcomes of neoadjoint therapy combined with liver transplantation versus radical hepatectomy for patients with surgically resectable hilar cholangiocarcinoma.Methods:A retrospective study was performed on the data of 64 patients with resectable hilar cholangiocarcinoma operated from January 2009 to December 2014 at the Organ Transplantation Department of the First Central Hospital of Tianjin. There were 43 males and 21 females, with an average age of 61.2 years. There were 45 patients who underwent radical hepatectomy in the liver resection group, and 19 patients who underwent combined neoadjuvant therapy (radiotherapy combined with 5-fluorouracil intravenous drip, transcatheter lumen radiotherapy, capecitabine oral administration) and liver transplantation in the liver transplantation group. The recurrence rates and survival rate were compared between groups.Results:The 1, 3 and 5 years cumulative survival rates of the liver transplantation group were 89.5%, 73.7% and 63.2%, respectively, which were significantly better than those of the liver resection group (80.0%, 53.3% and 35.6%) ( P<0.05). The postoperative tumor recurrence rate in the liver transplantation group was 31.6% (6/19), which was significantly lower than that in the liver resection group of 60.0% (27/45) ( P<0.05). Subgroup analysis using postoperative pathological results showed the cumulative survival rates of patients without lymph node metastasis (N 0) and those with negative resection margins (R 0) were not significantly different between groups ( P>0.05). However, for patients with regional lymph node invasion (N 1) and with R 0 resection margin, the cumulative survival rates at 1, 3 and 5 years after liver transplantation were 83.3%, 66.7% and 50.0%, respectively, which were significantly superior to the 64.3%, 28.6% and 14.3% of the liver resection group ( P<0.05). Conclusion:Hepatectomy is recommended for patients with N 0 R 0 resectable hilar cholangiocarcinoma. For patients with hilar cholangiocarcinoma with marginally resectable N 1R 0, neoadjuvant therapy combined with liver transplantation resulted in significantly better long-term overall survival than resection.

Objective To investigate the protective effects and underlying molecular mechanisms of hydrogen (H2) on high glucose-induced poly (ADP-ribose) polymerase-1 (PARP-1) dependent cell death (PARthanatos) in primary rat Schwann cells. Methods Cultured primary rat Schwann cells were randomly divided into five groups: blank control group (C group), H2 control group (H2 group), high osmotic control group (M group), high glucose treatment group (HG group), and H2 treatment group (HG+H2 group). The cells in H2 group and HG+H2 group were cultured with saturated hydrogen-rich medium containing 0.6 mmol/L of H2, and those in three control groups were cultured with low sugar DMEM medium containing 5.6 mmol/L of sugar, and the cells in HG and HG+H2 groups were given 44.4 mmol/L of glucose in addition (the medium containing 50 mmol/L of glucose), the cells in C group and H2 group were given the same volume of normal saline, and the cells in M group were given the same volume of mannitol. Cytotoxicity was evaluated using lactate dehydrogenase (LDH) release rate assays after treatment for 48 hours in each group. The contents of peroxynitrite (ONOO-) and 8-hydroxy-2-deoxyguanosine (8-OHdG) reflecting oxidative stress injury and DNA damage were detected by enzyme linked immunosorbent assay (ELISA). Poly (ADP-ribose) (PAR) protein expression was analyzed by Western Blot, and immunofluorescence staining was used to determine the nuclear translocation of the apoptosis-inducing factor (AIF). Results The cytotoxicity in HG and HG+H2 groups was significantly increased as compared with that of C group [LDH release rate: (61.40±2.89)%, (42.80±2.32)% vs. (9.92±0.38)%, both P < 0.01], the levels of ONOO- and 8-OHdG were markedly elevated [ONOO- (ng/L): 853.58±51.00, 553.11±38.66 vs. 113.56±14.22; 8-OHdG (ng/L): 1 177.37±60.97, 732.06±54.29 vs. 419.67±28.77, all P < 0.01], and the PAR protein expression was up-regulated (A value: 0.603±0.028, 0.441±0.010 vs. 0.324±0.021, both P < 0.01). The cytotoxicity, the levels of ONOO- and 8-OHdG, and PAR expression in HG+H2 group were significantly lower than those of the HG group (all P < 0.01). There were no significant differences in above parameters between H2 group as well as M group and C group. It was shown by immunofluorescence that AIF was expressed in the cytoplasm in C group, H2 group and M group, AIF was expressed in the whole cell in HG group, and the expression in the nucleus was particularly increased. A small amount of AIF expression was found in the nucleus of HG+H2 group, which indicated that high glucose could promote the AIF nuclear translocation, and that hydrogen-rich medium could prevent the process of translocation. Conclusions High glucose levels could enhance DNA damage that enhance PARthanatos in primary rat Schwann cells. However, H2 can not only reduce DNA damage of injured cells, but also inhibit the special death process, reduce the cell toxicity, all of which have protective effects.

Objective To compare the difference of clinical efficacy between surgical magnifying glass and surgical microscope assisted hepatic artery reconstruction in living donor liver transplantation (LDLT). Methods Clinical data of 272 donors and recipients undergoing LDLT were retrospectively analyzed. According to different patterns of hepatic artery reconstruction, all recipients were divided into the magnifying glass group (n=189) and microscope group (n=83). The operation time, intraoperative blood loss, hepatic artery reconstruction site, diameter of anastomosis, incidence of postoperative complications and survival rate of recipients were statistically compared between two groups. Results Compared with the microscope group, the operation time, hepatic artery reconstruction time and intraoperative blood loss were significantly less in the magnifying glass group (all P < 0.001). The most common site of hepatic artery reconstruction was the right hepatic artery in two groups, and the diameter of anastomosis was (2.1±0.9) mm in the magnifying glass group and (2.1±0.8) mm in the microscope group, with no statistical significance between two groups (P > 0.05). The 1-, 2- and 3-year survival rates of recipients in the magnifying glass group were 88%, 86% and 85%, which did not significantly differ from 89%, 87% and 86% in the microscope group (all P > 0.05). The incidence of postoperative complications did not significantly differ between two groups (all P > 0.05). Conclusions The efficacy and safety of hepatic artery reconstruction in LDLT under surgical magnifying glass are equivalent to those under surgical microscope, with less operation workload and intraoperative blood loss. For experienced transplantation surgeons, it is recommended to perform hepatic artery reconstruction assisted by surgical magnifying glass.

Objective To explore the teaching methodologies to improve the training of the Key Health professional scientific and technological personnel of ethnic minorities in Xinjiang.Methods To analyze the impact factors of the training effectiveness,developed training models that mainly aimed for research capacity building,which combined with training purposes to empower their learning and research in practice.Results The main impact factors affecting the training effectiveness of the Key Health professional scientific and technological personnel of ethnic minorities in Xinjiang include relatively poor understanding and expression of language,limited education and academic training,undeveloped economic status and weak sense of scientific research.Pre and post training tests were conducted by the clinical departments to evaluate and grade the learning ability and scientific research practice ability of these trainees.The final scores will be statistically analyzed according to three categories,namely 60 points,60 80 points and 80-100 points.The statistical results showed that after the training for all the 115 trainees,their language comprehension,imagination,perception,observation,memory,thinking and knowledge integration abilities have all been significantly improved in learning ability,with statistically significant differences (P＜0.01).In addition to the literature reading and comprehensive ability in scientific research and practical ability,identification evaluation ability,the sensibility of problems,information collection and selection,prediction,hands-on,analysis abilities,flexible thinking,logic reasoning,expression,organization and coordination,independent decision-making ability were significantly increased,the difference is statistically significant(P＜0.05).Conclusions Designing a practical training program for particular scientific research ability,targeted innovation,training curriculum content,methods and methods are effective measures to improve the research ability of the Key Health professional scientific and technological personnel of ethnic minorities in Xinjiang.

0.05). Conclusion After heated, the physical stability of UAE and UAS is reduced, the viscosity become lower, ADM releasing rate is fell. The heated Lipiodol-Adriamycin pharmaceutics had advantage in the interventional embolization chemotherapy of the neoplasm.

Objective:To investigate the effects of gelatin methacrylate anhydride (GelMA) hydrogel loaded with small extracellular vesicles derived from human umbilical cord mesenchymal stem cells (hUCMSCs-sEVs) in the treatment of full-thickness skin defect wounds in mice.Methods:This study was an experimental study. hUCMSCs-sEVs were extracted by ultracentrifugation, their morphology was observed through transmission electron microscope, and the expression of CD9, CD63, tumor susceptibility gene 101 (TSG101), and calnexin was detected by Western blotting. The human umbilical vein endothelial cells (HUVECs), the 3 rd and 4 th passages of human epidermal keratinocytes (HEKs) and human dermal fibroblasts (HDFs) were all divided into blank control group (routinely cultured) and hUCMSC-sEV group (cultured with the cell supernatant containing hUCMSCs-sEVs). The cell scratch test was performed and the cell migration rates at 6, 12, and 24 h after scratching were calculated, the cell Transwell assay was performed and the number of migration cells at 12 h after culture was calculated, and the proportion of proliferating cells was detected by 5-acetylidene-2'-deoxyuridine and Hoechst staining at 24 h after culture, with sample numbers being all 3. The simple GelMA hydrogel and the GelMA hydrogel loaded with hUCMSCs-sEVs (hereinafter referred to as hUCMSC-sEV/GelMA hydrogel) were prepared. Then the micromorphology of 2 kinds of hydrogels was observed under scanning electron microscope, the distribution of hUCMSCs-sEVs was observed by laser scanning confocal microscope, and the cumulative release rates of hUCMSCs-sEVs at 0 (immediately), 2, 4, 6, 8, 10, and 12 d after soaking hUCMSC-sEV/GelMA hydrogel in phosphate buffer solution (PBS) were measured and calculated by protein colorimetric quantification ( n=3). Twenty-four 6-week-old male C57BL/6J mice were divided into PBS group, hUCMSC-sEV alone group, GelMA hydrogel alone group, and hUCMSC-sEV/GelMA hydrogel group according to the random number table, with 6 mice in each group, and after the full-thickness skin defect wounds on the back of mice in each group were produced, the wounds were performed with PBS injection, hUCMSC-sEV suspenson injection, simple GelMA coverage, and hUCMSC-sEV/GelMA hydrogel coverage, respectively. Wound healing was observed on post injury day (PID) 0 (immediately), 4, 8, and 12, and the wound healing rates on PID 4, 8, and 12 were calculated, and the wound tissue was collected on PID 12 for hematoxylin-eosin staining to observe the structure of new tissue, with sample numbers being both 6. Results:The extracted hUCMSCs-sEVs showed a cup-shaped structure and expressed CD9, CD63, and TSG101, but barely expressed calnexin. At 6, 12, and 24 h after scratching, the migration rates of HEKs (with t values of 25.94, 20.98, and 20.04, respectively), HDFs (with t values of 3.18, 5.68, and 4.28, respectively), and HUVECs (with t values of 4.32, 19.33, and 4.00, respectively) in hUCMSC-sEV group were significantly higher than those in blank control group ( P<0.05). At 12 h after culture, the numbers of migrated HEKs, HDFs, and HUVECs in hUCMSC-sEV group were 550 ±23, 235 ±9, and 856 ±35, respectively, which were significantly higher than 188 ±14, 97 ±6, and 370 ±32 in blank control group (with t values of 22.95, 23.13, and 17.84, respectively , P<0.05). At 24 h after culture, the proportions of proliferating cells of HEKs, HDFs, and HUVECs in hUCMSC-sEV group were significantly higher than those in blank control group (with t values of 22.00, 13.82, and 32.32, respectively, P<0.05). The inside of simple GelMA hydrogel showed a loose and porous sponge-like structure, and hUCMSCs-sEVs was not observed in it. The hUCMSC-sEV/GelMA hydrogel had the same sponge-like structure, and hUCMSCs-sEVs were uniformly distributed in clumps. The cumulative release rate curve of hUCMSCs-sEVs from hUCMSC-sEV/GelMA hydrogel tended to plateau at 2 d after soaking, and the cumulative release rate of hUCMSCs-sEVs was (59.2±1.8)% at 12 d after soaking. From PID 0 to 12, the wound areas of mice in the 4 groups gradually decreased. On PID 4, 8, and 12, the wound healing rates of mice in hUCMSC-sEV/GelMA hydrogel group were significantly higher than those in the other 3 groups ( P<0.05); the wound healing rates of mice in GelMA hydrogel alone group and hUCMSC-sEV alone group were significantly higher than those in PBS group ( P<0.05). On PID 8 and 12, the wound healing rates of mice in hUCMSC-sEV alone group were significantly higher than those in GelMA hydrogel alone group ( P<0.05). On PID 12, the wounds of mice in hUCMSC-sEV/GelMA hydrogel group showed the best wound epithelization, loose and orderly arrangement of dermal collagen, and the least number of inflammatory cells, while the dense arrangement of dermal collagen and varying degrees of inflammatory cell infiltration were observed in the wounds of mice in the other 3 groups. Conclusions:hUCMSCs-sEVs can promote the migration and proliferation of HEKs, HDFs, and HUVECs which are related to skin wound healing, and slowly release in GelMA hydrogel. The hUCMSC-sEV/GelMA hydrogel as a wound dressing can significantly improve the healing speed of full-thickness skin defect wounds in mice.

【Objective】  To predict the active components and action targets of Wuyao (Linderae Radix) in the treatment of chronic pelvic inflammatory disease (CPID) based on network pharmacology, explore possible mechanisms of the treatment through animal experiments, and provide a scientific basis for clinical applications of Wuyao (Linderae Radix). 【Methods】  Possible active components and targets of Wuyao (Linderae Radix) in the treatment of CPID were obtained applying network pharmacology and molecular docking technology. CPID rat models were established using the mixed Escherichia coli, Staphylococcus aureus, and Ureaplasma urealyticum plus the performance of mechanical injury. Hematoxylineosin (HE) staining was applied to observe the pathological changes in the uterus, fallopian tube, and spleens of rat models. The contents of nitric oxide (NO), superoxide dismutase (SOD), and malondialdehyde (MDA) in the serum of rats were determined with the use of corresponding detection kits. Enzyme-linked immunosorbent assay (ELISA) test was used to measure the expression of interleukin (IL)-6 and IL-10 in the serum of rat models. Flow cytometry was used to determine the percentage of CD4+ and CD8a+ T cells as well as CD4+ CD25+ regulatory T cells (Tregs) in the spleen of rat models. 【Results】  A total of nine potential active components and four core therapeutic targets related to inflammatory response in Wuyao (Linderae Radix) were obtained. The animal experiments showed that Wuyao (Linderae Radix) markedly inhibited uterus swelling, regulated morphological changes in the fallopian tube and spleen,  effectively reduced inflammatory infiltration and injuries in the uterus and fallopian tube, and improved spleen functions in CPID rats. Moreover, Wuyao (Linderae Radix) markedly reduced the levels of NO, IL-6, and MDA, and increased the levels of IL-10 and SOD in the serum of rats. Wuyao (Linderae Radix) also elevated the percentage of CD4+T cells and the CD4+ T/CD8a+ T cell ratio, reduced the percentage of CD8a+ T cells, and raised the percentage of CD4+ CD25+ Tregs that had been abnormally decreased in rat models (P < 0.05). 【Conclusion】  Wuyao (Linderae Radix) could have therapeutic effects on CPID rats by relieving oxidative stress, mitigating inflammatory levels, and regulating the immuno-function of T cell subgroups to improve the pathological changes in CPID rats. It is a medicinal herb worth being further explored for its clinical values.

ERα-36 is a novel subtype of estrogen receptor α which promotes tumor cell proliferation, invasion and drug resistance, and it serves as a therapeutic target. However, only small-molecule compounds targeting ERα-36 are under development as anticancer drugs at present. Gene therapy approach targeting ERα-36 can be explored using recombinant adenovirus armed with decoy receptor. The recombinant shuttle plasmid pDC316-Ig κ-ERα-36-Fc-GFP was constructed via genetic engineering to express an Ig κ-signaling peptide-leading secretory recombinant fusion protein ERα-36-Fc. The recombinant adenovirus Ad-ERα-36-Fc-GFP was subsequently packaged, characterized and amplified using AdMax^TM adenovirus packaging system. The expression of fusion protein and functional outcome of Ad-ERα-36-Fc-GFP transduction were further analyzed with triple-negative breast cancer MDA-MB-231 cells. Results showed that the recombinant adenovirus Ad-ERα-36-Fc-GFP was successfully generated. The virus effectively infected MDA-MB-231 cells which resulted in expression and secretion of the recombinant fusion protein ERα-36-Fc, leading to significant inhibition of EGFR/ERK signaling pathway. Preparation of the recombinant adenovirus Ad-ERα-36-Fc-GFP provides a basis for further investigation on cancer gene therapy targeting ERα-36.
Adenoviridae/genetics* ; Cell Proliferation ; Estrogen Receptor alpha/metabolism* ; Recombinant Proteins ; Transfection