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Objective:To observe the expression levels of Toll-like receptor 4 (TLR4) signaling pathway-related proteins and their phosphorylation in the liver tissues of rats with inorganic arsenic poisoning, and to explore the role of TLR4-mediated inflammatory signaling pathway in arsenic-induced liver fibrosis injury.Methods:Eighteen healthy weanling SD rats were divided into 3 groups according to their body weight (80 - 100 g) using a random number table (6 rats in each group, half males and half females). The control group was given 10 ml/kg of normal saline by gavage. The sodium arsenite (NaAsO 2) exposure group was given 10 mg/kg of NaAsO 2 by gavage. The TAK-242 intervention group was given 10 mg/kg of NaAsO 2 by gavage, and 0.5 mg/kg of TAK-242 was also administered intraperitoneally to inhibit TLR4 after 12 weeks. All rats were administered 6 days a week for 36 weeks. At the end of the treatment, the liver tissues and serum of the rats in each group were collected. HE and Masson staining were used to observe the pathological and fibrotic changes of the liver tissues. Automatic biochemical analyzer was used to detect serum liver function indexes of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP). Western blot was used to detect the expression changes of rat liver fibrosis protein α-smooth muscle actin (α-SMA), transforming growth factor-β1 (TGF-β1), Vimentin and TLR4 signaling pathway-related proteins TLR4, nuclear factor κB (NF-κB)-p65 subunit (p65), NF-κB-p50 subunit (p50) and their phosphorylation p-p65 and p-p50 expression levels. Enzyme-linked immunosorbent assay (ELISA) was used to detect the secretion levels of inflammatory related factors interleukin (IL)-6, tumor necrosis factor-α (TNF-α) and IL-10. Results:HE and Masson staining results showed that compared with the control group, the NaAsO 2 exposure group showed significant inflammatory cell infiltration, hepatocyte necrosis and collagen fibrous deposition, while the TAK-242 intervention group showed improvement of the inflammatory cell infiltration and reduction of collagen fibrous deposition compared with the NaAsO 2 exposure group. The results of serum liver function indexes showed that ALT, AST and ALP in NaAsO 2 exposure group were increased compared with the control group, but the TAK-242 intervention group was significantly decreased compared with the NaAsO 2 exposure group ( P < 0.05). Western bolt results showed that in NaAsO 2 exposure group, the expression levels of fibrosis protein α-SMA, TGF-β1 and Vimentin (1.04 ± 0.19, 0.92 ± 0.14, 1.20 ± 0.21) and TLR4 signaling pathway-related proteins and their phosphorylation TLR4, p50, p-p50 and p-p65 (1.16 ± 0.21, 0.95 ± 0.16, 1.24 ± 0.23, 1.56 ± 0.25) were higher than the control group (0.44 ± 0.08, 0.42 ± 0.08, 0.72 ± 0.07, 0.69 ± 0.15, 0.71 ± 0.11, 0.46 ± 0.07, 0.54 ± 0.11, P < 0.05), and the TAK-242 intervention group (0.60 ± 0.13, 0.59 ± 0.16, 0.49 ± 0.11, 0.47 ± 0.08, 0.86 ± 0.09, 0.79 ± 0.14, 1.02 ± 0.17) were lower than the NaAsO 2 exposure group ( P < 0.05). There was no significant difference in the expression level of TLR4 signal pathway-related protein p65 among the three groups ( F = 14.29, P = 0.053). ELISA results showed that the secretion levels of IL-6 and TNF-α [(98.89 ± 4.58), (83.25 ± 4.57) ng/g] in rats liver tissues of the NaAsO 2 exposure group were higher than the control group [(27.30 ± 3.92), (27.77 ± 1.83) ng/g, P < 0.05], while the secretion level of IL-10 [(36.88 ± 3.86) ng/g] was lower than the control group [(77.96 ± 7.87) ng/g, P < 0.05]. In TAK-242 intervention group, IL-6 and TNF-α secretion levels [(44.32 ± 3.60), (36.51 ± 2.93) ng/g] were lower and IL-10 secretion level [(60.40 ± 4.94) ng/g] was higher compared with the NaAsO 2 exposure group ( P < 0.05). Conclusion:TLR4-mediated inflammatory signaling pathway-related proteins and their phosphorylation are highly expressed in the liver tissues of rats with inorganic arsenic poisoning, and inhibition of TLR4 signaling pathway could significantly reduce the degree of liver fibrosis injury caused by inorganic arsenic in rats.
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Background Long-term exposure to sodium arsenite leads to its accumulation in the liver and liver injury as a result. Previous studies showed that mesenchymal cells play an important role in hepatic fibrosis, and epithelial-mesenchymal transformation (EMT) is considered to be a main source of mesenchymal cells. Objective To investigate the effects of sodium arsenite at different doses on liver fibrosis and EMT-related protein expressions in SD rats. Methods Twenty-four healthy weaned SD rats, half male and half female, were randomly divided into four groups according to body weight, with 6 rats in each group. The four groups were control group (gavage with 10.0 mL·kg−1 physiological saline), 2.5 mg·kg−1 sodium arsenite group, 5.0 mg·kg−1 sodium arsenite group, and 10.0 mg·kg−1 sodium arsenite group. All rats were gavaged 6 d per week for 36 weeks and weighed once a week, the serum and liver tissues of rats were collected and weighed, then the organ coefficient was calculated. Hematoxylin-eosin staining and Masson's trichrome staining were used to determine the pathological changes of hepatic fibrosis in rats. The serum secretion levels of hyaluronic acid (HA), laminin (LN), procollagen Ⅲ N-terminal propeptide (PⅢNP), and collagen Ⅳ (COL-Ⅳ) in rats were detected by enzyme-linked immunosorbent assay (ELISA). The protein expressions of HSCs activation-related proteins, such as α-smooth muscle actin (α-SMA) and transforming growth factor-β1 (TGF-β1), as well as EMT-related markers, such as E-cadherin, N-cadherin, Vimentin, and Snail, were detected by Western blotting. Results Compared with the control group, the 10.0 mg·kg−1 sodium arsenite group showed decreased body weight (P<0.05) and increased liver coefficient (P<0.05) of female and male rats. The pathological staining showed that, compared with the control group, a large number of inflammatory cells were observed in liver tissue of rats exposed to sodium arsenite, liver parenchymal cells were also liquefied, necrotic, and denatured, and the collagen positive staining area of liver tissue showed an upward trend along with the increase of arsenic exposure dose (P<0.05). The results of ELISA and Western blotting showed that the serum secretion levels of HA, LN, PⅢNP, and COL-Ⅳ in the 5.0 and 10.0 mg·kg−1 sodium arsenite groups were higher than those in the control group and the 2.5 mg·kg−1 sodium arsenite group (P<0.05). Compared with the control group, the expressions of α-SMA and TGF-β1 proteins in liver tissue were increased in each sodium arsenite exposure group (P<0.05), the expression levels of E-cadherin protein were decreased (P<0.05), and the expression levels of N-cadherin, Vimentin, and Snail were increased (P<0.05). Conclusion Sodium arsenite exposure can induce HSCs activation and liver fibrosis injury in SD rats, resulting in increased extracellular matrix secretion levels, accompanied by EMT in liver tissue, suggesting that EMT is closely related to the process of liver fibrosis caused by arsenic.
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Background Arsenic is a well-known environmental toxicant. Hepatic fibrosis could occur dueto excessive or long-term exposure to arsenic, while associated molecular mechanisms remain undefined. Mitogen-inducible gene 6 (Mig-6) exhibits a protective effect on numerous diseases or cancers. However, the specific role of Mig-6 in the mechanisms of arsenite-induced hepatic fibrosis remains indistinct. Objective To investigate the specific role of Mig-6 in the activation of hepatic stellate cells (HSC) and the deposition of extracellular matrix (ECM) induced by sodium arsenite (NaAsO2). Methods Human hepatic stellate cells (Lx-2) were treated with 0, 1.875, 3.75, 7.5, and 15 μmol·L−1 of NaAsO2 for 24 h, or with 7.5 μmol·L−1 NaAsO2 for 0, 12, 24, 48, and 72 h. Additionally, Lx-2 cells were transfected by pcDNA3.1(+)/Mig-6, then treated with 7.5 μmol·L−1 NaAsO2 for 24 h; a blank control group, a pcDNA3.1(+)-control group, a pcDNA3.1(+)/Mig-6 group, and an arsenic (7.5 μmol·L−1 NaAsO2) group were also set up. After transfection, the cells and culture supernatants were collected, and the protein levels of Mig-6, α-smooth muscle actin (α-SMA), and transforming growth factor-β1 (TGF-β1) in Lx-2 cells were identified by Western blotting analysis; moreover, the secretion levels of main ECM components in supernatants such as hyaluronic acid (HA), laminin (LN), collagens IV (COL-IV), and procollagen-III (PIIINP) were tested by ELISA. Results The Mig-6 expression decreased in the 3.75, 7.5, and 15 μmol·L−1 NaAsO2 groups (0.561±0.095, 0.695±0.048, and 0.401±0.030) compared to the control group (1.000±0.000) in Lx-2 cells (P<0.05). After administration with 7.5 μmol·L−1 of NaAsO2 for 24, 48, and 72 h, the Mig-6 expression (0.856±0.036, 0.515±0.077, 0.491±0.060) decreased compared with the 0 h group (1.000±0.000) (P<0.05). After over-expression of Mig-6, the results of Lx-2 activation related protein levels showed that compared to the control group, the α-SMA and TGF-β1 expression were up-regulated in the arsenic group (P<0.05); meanwhile, the α-SMA and TGF-β1 in the Mig-6 over-expression combined arsenic exposure group reduced compared to the arsenic (7.5 μmol·L−1) group (P<0.05). The results of ELISA showed that compared with the control group, the HA, LN, PIIINP, COL-IV in the arsenic group were up-regulated (P<0.05); while compared to the arsenic group, the HA, LN, PIIINP, and COL-IV in the Mig-6 over-expression combined with arsenic exposure group were decreased (P<0.05). Conclusion Arsenic down-regulates Mig-6 expression in HSC, and over-expression of Mig-6 can reverse the activation of HSC and ECM deposition induced by arsenic exposure. It suggests that Mig-6 plays a protective role in arsenic-induced HSC activation and ECM deposition.
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Objective To observe the effect of different doses of atorvastatin in the treatment of chronic heart failure of coronary heart disease (CHD), and to provide scientific reference for the choice of clinical medication. Methods 54 cases from June 2015 ~2017 year in April in our hospital with 20mg/d atorvastatin therapy regimen in the treatment of chronic heart failure of coronary heart disease patients as the study group, 54 cases of chronic heart failure of coronary heart disease patients with another force selected by 40mg/d atorvastatin therapy regimen as study group. LDL-C, LVESD, LVEDD, LVEF, hs-CRP, 6MWT, NT-proBNP and other indicators as evaluation basis, through the observation of the two groups before and after treatment the indexes to evaluate clinical efficacy, adverse events were observed after treatment in two groups. Results There was no significant difference between the groups before and after treatment, and there was no statistical significance(P<0.05). The indexes of clinical curative effect were improved in different degree after treatment, and the improvement effect in observation group was better than that in control group (P<0.05). Conclusion Atorvastatin 40mg daily oral drug treatment of chronic heart failure in patients with coronary heart disease is the best choice of atorvastatin dose, the dose range of atorvastatin treatment effectiveness and safety protection, improve clinical symptoms, promote the improvement of the quality of life, worthy of clinical application.
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Objective To compare the reliability and validity of 2 incontinence- associated dermatitis (IAD) risk assessment tools (IAD risk assessment scale and PAT) in patients with incontinence, and to search for the best risk assessment tool. Methods A total of 101 IAD cases were chosen from the Affiliated Hospital of Nantong University.Reliability was evaluated by the inter- rater reliability,internal consistency,item sensitivity analysis and retest reliability; validity was evaluated by the structure validity and predictive validity. Results Cronbach α was 0.313 (IAD risk assessment scale) and 0.421(PAT);Cronbach α coefficient was the highest after the items that Change sheets and mats and relevant factors were removed respectively in two tools,which were 0.431, 0.428; The correlation between the total scores were 0.711 (P<0.01)(IAD risk assessment scale)and 0.498 (PAT)(P<0.01);Area under curve were 0.661 (IAD risk assessment scale),0.864 (PAT); the best cut-off value to forecast a poor prognosis was 53.5 (IAD risk assessment scale),6.5 (PAT). Conclusions The reliability and validity of IAD risk assessment scale and PAT are low. The change of sheets and mats and Related influencing factors are needed to be adjusted and revised.
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Objective: To categorize and summarize the clinical and mechanism studies of the past 30 years on the treatment of Hashimoto’s thyroiditis (HT) with moxibustion, moxibustion plus medication, and acupuncture plus medication, etc., and to analyze the current problems. Methods: The clinical and laboratory studies related to the treatment of HT with acupuncture-moxibustion therapies published before June 2015 were retrieved from MEDLINE, Excerpta Medica Database (EMBASE), China National Knowledge Infrastructure (CNKI), Wanfang Academic Journal Full-text Database (Wanfang) and Chongqing VIP Database (CQVIP). Results:Moxibustion, moxibustion plus medication, and acupuncture plus medication can produce certain therapeutic effects in treating HT. Conclusion:The research on the treatment of HT with acupuncture-moxibustion therapies is rather limited in the amount and content. In the future, standardization should be fortified, specific moxibustion research needs deepening, and the action mechanism of moxibustion should be emphasized.
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BACKGROUND:The form and structure of antigen-extracted xenogeneic cancel ous bone through series of physical and chemical treatment are similar to human tissue. OBJECTIVE:To detect the biocompatibility of antigen-extracted xenogeneic cancel ous bone matrix prepared by three different ways. METHODS:The antigen-extracted xenogeneic cancel ous bone scaffold materials which were prepared through physical, chemical and physical-chemical combined methods and hydroxy apatite biological ceramic materials were implanted into the dorsum subcutaneous tissue. Histological observation was done at 4, 8 and 12 weeks after surgery. The antigen-extracted xenogeneic cancel ous bone scaffold materials which were prepared through physical, chemical and physical-chemical combined methods respectively was used to culture sheep bone marrow mesenchymal stem cells for 7 days. Cel adhesion, growth, proliferation and stroma secretion were observed. RESULTS AND CONCLUSION:At 4 weeks after surgery, a strong inflammatory reaction was detected around materials in four groups. At 12 weeks, the xenogeneic bone materials prepared through physical and physical-chemical combined methods and hydroxy apatite biological ceramic materials internal pore and surrounding tissue inflammation disappeared basical y, with the presence of thimbleful inflammation cells. The material degradation was more than at 8 weeks. The xenogeneic bone materials prepared through chemical methods material internal pore and surrounding tissue inflammation stil existed, suggesting that the xenogeneic bone materials prepared through physical and physical-chemical combined methods exhibited good histocompatibility. A smal amount of orderly osteoblasts existed around hydroxy apatite biological ceramic materials and physical-chemical prepared materials, with a smal amount of bone. These suggested that there was a tendency for ectopic bone formation. The xenogeneic cancel ous bone materials prepared through physical or physical-chemical combined methods have better cytocompatibility. However, scaffold materials prepared through chemical method have poor cytocompatibility and they are not qualified for the safety standards of biological materials.
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The aim of the study was to construct a recombinant adenovirus overexpression vector for Sterol Regulatory Element Binding Protein-1 (SREBP-1) of Xinong Saanen dairy goat, and to detect its effect on genes related to fatty acid metabolism in goat mammary epithelial cells, to establish foundation for further study of its roles in metabolism of fatty acid synthesis and lactation. First, we designed primers based on the SREBP-1 gene sequence in GenBank for PCR amplification and inserted the sequence into shuttle vector pAdTrack-CMV. The recombinant plasmid pAdTrack-CMV-SREBP-1 linearized by Pme I was transformed into E. coli BJ5183 competence cell containing the backbone vector pAdEasy-1 to obtain recombinant vector pAd-SREBP-1 by homologous recombination. pAd-SREBP-1 was linearized by Pac I and transfected into HEK 293 cell. Then we infected goat mammary epithelial cells with recombinant adenovirus which was packaged in HEK 293 cell line. The results showed that the recombinant adenovirus vector containing SREBP-1 was successfully constructed, and the titer of virus was 10(9) U/mL. Compared with the control group, mRNA level of SREBP-1 increased by about 15 times after infected for 48 h and 30 times after infected for 72 h. Fatty acid synthase (FASN) and Acetyl-CoA carboxylase (ACC) was upregulated by almost 2 times. The expression level of Peroxisome proliferator activated receptorgamma (PPARgamma) increased by 1.5 times. Liver X receptoralpha (LXRalpha) and Adipose triglyceride lipase (ATGL) upregulated by 1.2 times compared with that of control. But Stearoyl-coenzyme A desaturase (SCD) had no obvious change. In conclusion, SREBP-1 can activate the expression of genes related to fatty acid synthesis in mammary epithelial cells of Xinong Saanen dairy goat, demonstrated a regulatory function on the fatty acid metabolism in goat mammary gland.
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Animaux , Femelle , Humains , Adenoviridae , Génétique , Métabolisme , Cellules épithéliales , Biologie cellulaire , Métabolisme , Acides gras , Métabolisme , Régulation de l'expression des gènes , Capra , Génétique , Cellules HEK293 , Métabolisme lipidique , Génétique , Glandes mammaires animales , Biologie cellulaire , Protéine-1 de liaison à l'élément de régulation des stérols , Génétique , Métabolisme , TransfectionRésumé
Objective To study the relationship between cleft palate and secretory otitis media(SOM)and the change of hearing of cleft palate patients with SOM before and after tube insertion.Methods 203 cleft palate patients(406 ears)were investigated by pure tone audiometry,acoustic immitance audiometry and auditory brainstem response(ABR).21 cleft palate patients with SOM underwent palatoplasty and myringotomy with tube insertion at the same time.Results The incidence of middle ear disfunction was 68.5% in 203 cases with cleft palate,which in the cases under 6 years the incidence was 80%.After operation,21 cases were followed up one to three years,their average hearing improvement was 17dB by tube insertion( P