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Abstract@#As the largest human microecosystem, intestinal microorganisms participate in human material and energy metabolisms and pose a significant impact on human health. Diabetes mellitus is likely to cause imbalance of abundance and component alterations in intestinal microorganisms, and reduce the diversity and balance, leading to intestinal microflora dysregulation. It has been shown that intestinal microflora dysregulation may promote diabetes development and progression through the reduction of intestinal microbial metabolites, inflammatory reaction and insulin resistance. This review summarizes the involvement of intestinal microorganisms in the pathogenesis of diabetes through metabolites including short-chain fatty acid, bile acid and lipopolysaccharide, and describes the current status of intestinal microorganisms-mediated treatments for diabetes, so as to provide the theoretical basis for the researches on diabetes and intestinal microorganisms.
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Objective@#To investigate the protective effects and mechanism of keratinocyte growth factor (KGF) combined with hypoxia inducible factor-1α (HIF-1α) on intestinal crypt epithelial cells (IEC-6) of rats with hypoxia stress.@*Methods@#(1) The routinely cultured IEC-6 of rats were collected and divided into normoxia blank group, normoxia KGF group, normoxia HIF-1α group, and normoxia combine group, according to the random number table, and then the previous mediums were respectively replaced with dulbecco′s modified eagle medium (DMEM), medium with 0.5 ng/mL KGF, medium with 10.0 ng/mL HIF-1α, and medium with 0.5 ng/mL KGF and 30.0 ng/mL HIF-1α. And the cells were cultured in cell incubator with oxygen volume fraction of 21% for 24 hours. (2) Another batch of routinely cultured IEC-6 were collected and divided into normoxia control group, hypoxia control group, hypoxia KGF group, hypoxia HIF-1α group, and hypoxia combine group, according to the random number table. The previous mediums were replaced with DMEM, DMEM, medium with 0.5 ng/mL KGF, medium with 10.0 ng/mL HIF-1α, and medium with 0.5 ng/mL KGF and 30.0 ng/mL HIF-1α respectively. And then, the cells in normoxia control group were cultured routinely for 24 hours, and cells in the other 4 groups were cultured in cells incubator of 3 gases, with oxygen volume fraction of 5% for 24 hours. Cells cultured in normoxic and hypoxic incubators were collected, with 3 samples in each group, and morphological changes of cells were observed with optical microscope. Cells cultured in normoxic and hypoxic incubators were collected, with 3 samples in each group, and survival rates of cells were detected by cell count kit 8. Cells in normoxia control group and cells cultured in hypoxic incubator were collected, with 3 samples in each group. The cell cycle changes and apoptosis rates were detected by flow cytometer, the content of adenosine triphosphate (ATP) was detected by ultraviolet spectrophotometer, and protein expression of p53 was detected by Western blotting. Data were processed with one-way analysis of variance and least significant difference test.@*Results@#(1) After being cultured for 24 h, cells cultured in normoxic incubator grew well with oval or round shapes and clear cytoplasm, and cells cultured in hypoxic incubator showed irregular shapes such as fusiform or starlike shape, with black particle in cytoplasm. (2) After being cultured for 24 h, cell survival rates of normoxia blank group, normoxia KGF group, normoxia HIF-1α group, and normoxia combine group were (107.4±8.7)%, (109.8±2.9)%, (115.8±7.4)%, and (112.8±10.6)% respectively. There was no significantly statistical difference in general comparison of cell survival rates among the above groups (F=0.685, P=0.586). After being cultured for 24 h, cell survival rates of hypoxia control group, hypoxia KGF group, hypoxia HIF-1α group, and hypoxia combine group were (35.1±4.6)%, (52.9±6.8)%, (56.2±3.1)%, and (71.2±9.6)% respectively, which were significantly lower than (106.3±12.3)% of normoxia control group (P<0.001). Survival rates of cells in hypoxia KGF group, hypoxia HIF-1α group, and hypoxia combine group were significantly higher than the rate of cells in hypoxia control group (P=0.023, 0.009, <0.001). Survival rate of cells in hypoxia combine group was significantly higher than the rates of cells in hypoxia KGF group and hypoxia HIF-1α group (P=0.017, 0.045). (3) After being cultured for 24 h, percentage of cells in G1 phase in hypoxia control group was significantly higher than that of cells in normoxia control group (P=0.030), percentages of cells in S phase in hypoxia control group, hypoxia KGF group, and hypoxia HIF-1α group were obviously lower than the percentage of cells in normoxia control group (P=0.020, 0.031, 0.026), and percentages of cells in different phases in other groups were close to those of cells in normoxia control group (P=0.516, 0.107, 0.052, 0.985, 0.637, 0.465, 0.314, 0.591). After being cultured for 24 h, percentages of cells in G1 phase in hypoxia control group, hypoxia KGF group, and hypoxia HIF-1α group were obviously higher than the percentage of cells in hypoxia combine group (P=0.001, 0.030, 0.014), and percentages of cells in S phase in the above 3 groups were obviously lower than the percentage of cells in hypoxia combine group (P=0.001, 0.012, 0.010). (4) After being cultured for 24 h, compared with that of cells in normoxia control group, apoptosis rate of cells in hypoxia control group obviously increased (P=0.018), and apoptosis rate of cells in hypoxia combine group obviously decreased (P=0.008). After being cultured for 24 h, compared with that of cells in hypoxia control group, apoptosis rates of cells in hypoxia KGF group and hypoxia combine group obviously decreased (P=0.004, 0.001). Apoptosis rate of cells in hypoxia combine group was obviously lower than those of cells in hypoxia KGF group and hypoxia HIF-1α group (P=0.032, 0.002). (5) After being cultured for 24 h, compared with that of cells in normoxia control group, the content of ATP of cells in hypoxia combine group changed unobviously (P=0.209), and content of ATP of cells in the other groups obviously decreased (P= <0.001, 0.001, 0.002). Content of ATP of cells in hypoxia HIF-1α group and hypoxia combine group was obviously higher than that of cells in hypoxia control group (P=0.044, 0.001). Content of ATP of cells in hypoxia combine group was obviously higher than that of cells in hypoxia KGF group and hypoxia HIF-1α group (P=0.011, 0.020). (6) After being cultured for 24 h, protein expressions of p53 of cells in hypoxia control group, hypoxia KGF group, and hypoxia HIF-1α group were obviously higher than that of cells in normoxia control group (P<0.001), and protein expression of p53 of cells in hypoxia combine group was obviously lower than those of cells in hypoxia control group, hypoxia KGF group, and hypoxia HIF-1α group (P=0.001, 0.001, 0.002).@*Conclusions@#KGF combined with HIF-1α have significant protective effects on IEC-6 of rats with hypoxia stress, and can improve its survival in hypoxic environment by inhibiting cell cycle arrest, reducing the level of apoptosis, and increasing level of energy metabolism.
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Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer, and its mortality ranks fourth in global malignant tumors. Early diagnosis of HCC has an important impact on the survival rate of patients. Exosomal microRNA (miRNA) is closely related to the occurrence and development of HCC. It can be secreted and transferred to recipient cells through the corresponding target genes, and play a role of regulating cancer progression. Exosomal miRNAs have great differences in the expression of HCC patients and in vitro cell lines, and it has the advantages of high content, specificity and stability relative to circulating miRNAs. It not only can better distinguish between HCC and healthy patients, but also can be further differentiated from hepatitis or cirrhosis, and it also has certain value in the diagnosis of recurrent HCC.
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Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer, and its mortality ranks fourth in global malignant tumors. Early diagnosis of HCC has an important impact on the survival rate of patients. Exosomal microRNA (miRNA) is closely related to the occurrence and development of HCC. It can be secreted and transferred to recipient cells through the corresponding target genes, and play a role of regulating cancer progression. Exosomal miRNAs have great differences in the expression of HCC patients and in vitro cell lines, and it has the advantages of high content, specificity and stability relative to circulating miRNAs. It not only can better distinguish between HCC and healthy patients, but also can be further differentiated from hepatitis or cirrhosis, and it also has certain value in the diagnosis of recurrent HCC.
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Insulin resistance (IR) is a complex metabolic disorder related to several diseases including type 2 diabetes (T2DM), hypertension and dyslipidemia. These diseases are all independent risk factors for cardiovascular disease. Lipid metabolism disorder has toxic effects on cells and may cause or aggravate IR in performance of elevated plasma levels of triglyceride (TG) and free fatty acids (FFA), the last one is an independent risk factor for IR. It has been clear that FFA may induce IR by endoplasmic reticulum (ER) stress, oxidative stress, apoptosis and inflammation, although the specific mechanisms remained unknown. The present paper summarizes the related molecules involved in the pathogenic process of IR and its mechanism, might provide a theoretical basis for the molecular mechanism of IR caused by FFA, and therapeutic reference for clinical treatment of IR and prevention of T2DM.
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The present study developed an oral hepatocyte growth factor (HGF) gene therapy strategy for gastric ulcers treatment. An attenuated Salmonella typhimurium that stably expressed high HGF (named as TPH) was constructed, and the antiulcerogenic effect of TPH was evaluated in a rat model of gastric ulcers that created by acetic acid subserosal injection. From day 5 after injection, TPH (1 × 10⁹ cfu), vehicle (TP, 1 × 10⁹ cfu), or sodium bicarbonate (model control) was administered orally every alternate day for three times. Then ulcer size was measured at day 21 after ulcer induction. The ulcer area in TPH-treated group was 10.56 ± 3.30 mm², which was smaller when compared with those in the TP-treated and model control groups (43.47 ± 4.18 and 56.25 ± 6.38 mm², respectively). A higher level of reepithelialization was found in TPH-treated group and the crawling length of gastric epithelial cells was significantly longer than in the other two groups (P < 0.05). The microvessel density in the ulcer granulation tissues of the TPH-treated rats was 39.9 vessels/mm², which was greater than in the TP-treated and model control rats, with a significant statistical difference. These results suggest that TPH treatment significantly accelerates the healing of gastric ulcers via stimulating proliferation of gastric epithelial cells and enhancing angiogenesis on gastric ulcer site.
Subject(s)
Animals , Rats , Acetic Acid , Epithelial Cells , Genetic Therapy , Granulation Tissue , Hepatocyte Growth Factor , Hepatocytes , Microvessels , Models, Animal , Salmonella typhimurium , Salmonella , Sodium Bicarbonate , Stomach Ulcer , UlcerABSTRACT
Objective This study was provided the evidence of infection respiratory tract clinical treatment through examining 695 cases of pathogen .Methods Using indirect immunofluorescence(IFA) IgM antibody with 9 respiratory pathogen in serum of 695 patients from August 2015 to March 2016 with respiratory tract infection from espiratory ,pediatrician ,thoracic surgery clinics were detected in this paper in order to provide the basis for clinical treatment .Results The results showed that single pathogen de‐tection positive rate was 33 .9% in 695 cases ,The top three of positive rate were MP(14 .1% ) ,IFB(9 .6% ) ,RSV(4 .8% );the posi‐tive rate of two mixed infection was 7 .79% ;The postive rate of MP ,IFB have obvious difference in seasons ,but which of Coxiella burneti was no obvious seasonal difference;there was an obvious difference between the sex ratio .Conclusion MP ,IFB ,RSV infec‐tion were given priority to detection rate in our region .
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Silent mating type information regulation 2 homolog-1 ( SIRT1 ) is a nicotinamide adenine dinucleotide ( NAD)-dependent deacetylase, which can deacetylate histone and non-histone proteins and is involved in many life proces-ses, such as energy metabolism, cell senescence and apoptosis.SIRT1 can enhance the cellular energy supply, inhibit cell apoptosis, alleviate inflammation, enhance the resistance of oxidative stress and improve the function of endothelial cells by deacetylating relevant factors in many vascular-related diseases.This article summarizes the current research progress in the function and signal pathway of SIRT1 in ischemic stroke, atherosclerosis and tumors.
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BACKGROUND:At present, there are few reports about construction of the model of acute high altitude stress-induced gastrointestinal injury. Construction of a stable high altitude stress-induced gastrointestinal injury model has become a premise of the current studies on acute stress-induced gastrointestinal injury and related diseases. OBJECTIVE: To establish rat models of acute high altitude stress-induced gastrointestinal ischemia/reperfusion injury. METHODS:Twenty-four male Wistar rats were randomly divided into normal control (lower limit of high altitude) , sham (high altitude) and ischemia/reperfusion (high altitude) groups (n=8 rats/group). Rats from al groups underwent a rush for high altitude region. The root of the superior mesenteric artery of rats in the ischemia/reperfusion group was clamped by a vascular clip to completely block blood flow for 60 minutes, and then intestinal blood flow was recovered for reperfusion for 60 minutes. The rats in the sham group were only
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<p><b>OBJECTIVE</b>To construct a recombinant adenovirus (pAdxsi-GFP-HIF) encoding human hypoxia inducible factor 1 α gene (HIF-1 α) and to express it in endothelial cells.</p><p><b>METHODS</b>HIF-1 α gene was obtained from human lung cancer cell line A549, which was cultured in hypoxia condition, by RT-PCR. The HIF-1 α gene was subcloned into shuttle vector p Shuttle-CMV-EGFP at KpnI and BamHI sites. After identified with restriction enzymes, plasmid p Shuttle-GFP-HIF was linearized by digestion with restriction endonuclease I-CeuI and I-SceI, and subsequently cotransformed into E.coli DH5a with adenoviral backbone plasmid pAdxsi to make homologous recombination. After linearized by PacI, the homologous recombinant adenovirus plasmid was transfected into 293 cells to package and amplify. The recombinant adenovirus was infected with human umbilical vein endothelial cells (ECV304), and the expression level of HIF-1 α protein was evaluated by ELISA.</p><p><b>RESULTS</b>The recombinant adenovirus vector containing HIF-1 α gene (pAdxsi-GFP-HIF) was successfully constructed and amplified with titer of 3.38 X 10(10) pfu/mL. The green fluorescence protein was detected under fluorescent microscope in ECV304 at 24h after transfection and with a stronger degree after 48h. The concentration of HIF-1 protein was (48.93 ±3.86)ng/mL in supernatant at 48 h after transfection.</p><p><b>CONCLUSION</b>A recombinant adenovirus vector pAdxsi-GFP-HIF, encoding human hypoxia inducible factor 1 α gene, has been constructed in vitro and expressed successfully in ECV304 cells.</p>
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Humans , Adenoviridae , Genetics , Cells, Cultured , Genetic Vectors , Human Umbilical Vein Endothelial Cells , Metabolism , Hypoxia-Inducible Factor 1, alpha Subunit , Genetics , Plasmids , GeneticsABSTRACT
ObjectiveTo investigate the effect of mesenchymal stem cells (MSCs) transfected with hepatic growth factor (HGF) gene on the survival volume of free grain fat grafts.Methods MSCs of male Wistar rats were transfected with Ad-GFP or Ad-HGF.The transfection infectivity of Ad-GFP to MSCs,the expression of HGF were measured using ELISA assay.150 rats were randomly divided into 5 groups:control group (group A),MSCs group (group B),Ad HGF group (group C),MSCs transplanted with Ad-GFP group (group D),and MSCs transplanted with Ad-HGF (MSCsHGF) group (group E).Then,the same volume of frain fat graft,mixed with DMEM LG andMSCs,Ad HGF,MSCs-GFP,MSCs HGF respectively,were transplanted to rats back,but control group were only mixed with DMEM-LG.Fat graft was obtained on days 3,5,7,14,28,and 60 after implantation.The volume of fat graft was measured by messcylinder,and the expression of HGF and CD34 in transplanted fat tissue were detected by immunohistochemistry.ResultsThe transfection infectivity of Ad-GFP to MSCs was 89.6 ％ at 100 MOI,the expression of HGF in MSCs culture medium reached to the level after being transfected with Ad-HGF for 48 h.Compared with other 4 groups,at days 3,5,7,and 14 post-transplantation,the expression of HGF in E group transplanted fat of group E had statistics significance (P＜0.05).The persentage of survival volume of fat graft in group E was significantly higher than that of other group ( P＜0.05) at days 28,and 60 post transplantation.ConclusionsMSCs transplanted with Ad-HGF could secrete HGF and increase the survival volume of fat grafts.
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<p><b>OBJECTIVE</b>To evaluate the effects of hepatocyte growth factor (HGF) on the prevention of scar formation and the promotion of wound healing by gene transfer.</p><p><b>METHODS</b>A total of 12 female New Zealand rabbits were used in this study. Rabbits were anesthetized with an intravenous injection of sodium pentobarbital, and identical wounds were made over the ventral surface of each ear. Five circular wounds, 7 mm in diameter, were created in each ear by excision through the skin to the underlying cartilage using sterile technique. After the surgical procedures, 10 of the rabbits were randomly allocated to five groups, with 2 rabbits in each group: Ad-HGF group 1, Ad-HGF group 2, Ad-HGF group 3, Ad-GFP (a reporter gene) group and the solvent group. Immediately after surgery, 6 x 10(7) pfu Ad-HGF, 6 x 10(8) pfu Ad-HGF, 6 x 10(9) pfu of Ad-HGF, 6 x 10(9) pfu of Ad-GFP, or same volume of solvent (PBS, pH 7.2) was applied once to each wound in groups 1 to 5, respectively. One additional rabbit was used to evaluate the transfer efficiency of the adenovirus vector by transferring Ad-GFP (6 x 10(9) pfu) into its wounds. Ice slides of wounds from this animal were observed under fluorescence microscopy. Another additional rabbit was used to evaluate the expression of HGF and TGFbeta1 after transferring Ad-HGF (6 x 10(9) pfu) into each of its wound. Immunohistochemistry was used for detection.</p><p><b>RESULTS</b>The effect of HGF on reducing excessive dermal scarring was observed by adenovirus-mediated gene transfer. Transfection of the human HGF cDNA into skin wounds through an adenoviral vector suppressed the over-expression of TGFbeta1, which plays an essential role in the progression of dermal fibrogenesis. Application of HGF to the wounds significantly enhanced wound healing and inhibited over scarring.</p><p><b>CONCLUSION</b>HGF gene therapy could be a new approach for preventing excessive dermal scarring in wound healing.</p>