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Objective:To evaluate the relationship between intestinal mucosal mast cells and intestinal flora during intestinal ischemia-reperfusion(I/R) in mice.Methods:Forty-eight healthy male C57BL/6 mice, aged 9-12 weeks, weighing 20-25 g, were divided into 4 groups ( n=12 each) using a random number table method: sham operation group (group Sham), sham operation plus mast cell membrane stabilizer cromolyn sodium (CS) group (group Sham+ CS), I/R group and I/R+ CS group.The intestinal I/R injury was induced by clamping the superior mesenteric artery for 45 min followed by 4 h of reperfusion in I/R and I/R+ CS groups.The superior mesenteric artery was only exposed but not occluded in Sham and Sham+ CS groups.CS 50 mg/kg was intraperitoneally injected once a day for two weeks before surgery in Sham+ CS and I/R+ CS groups.The equal volume of normal saline was intraperitoneally injected once a day for two weeks before surgery in Sham and I/R groups.Mice were then sacrificed and intestinal tissues were harvested for examination of the pathological changes of intestinal mucosa with a light microscope after HE staining, and the damage to intestinal tissues was assessed and scored according to Chiu.The expression of mast cell tryptase was detected using the immunohistochemical SP staining method, and the mast cells were counted.The intestinal contents were collected, the total amount of bacteria in intestinal flora was detected by 1% agarose gel electrophoresis, and the diversity (Chao1 index and Shannon index) and relative abundance of intestinal microbial bacteria were detected by 16S rDNA sequencing method. Results:Compared with Sham group, the Chiu′s score and mast cell count were significantly increased, the expression of mast cell tryptase was up-regulated, the total amount of intestinal flora bacteria was increased, Chao1 index and Shannon index were decreased, the relative abundance of Bacteroidetes, Proteobacteria, Bacteroides and Enterobacteriaceae were increased, and the relative abundance of Firmicutes, Lactobacillus and Bifidobacteria was reduced in I/R group ( P<0.05 or 0.01), and no significant change was found in the parameters mentioned above in Sham+ CS group ( P>0.05). Compared with I/R group, the Chiu′s score and mast cell count were significantly decreased, the expression of mast cell tryptase was down-regulated, the total amount of intestinal flora bacteria was decreased, Chao1 index and Shannon index were increased, the relative abundance of Bacteroidetes, Proteobacteria, Bacteroides and Enterobacteriaceae were reduced, and the relative abundance of Firmicutes, Lactobacillus and Bifidobacteria was increased in I/R+ CS group ( P<0.05 or 0.01). Conclusion:The activation of intestinal mucosal mast cell can lead to imbalance of intestinal flora, decrease the number of probiotics and increase the number of potential pathogenic bacteria, and thus be involved in the pathophysiological mechanism of intestinal I/R injury in mice.
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Objective To evaluate the role of tryptase in intestinal ischemia-reperfusion (Ⅰ/R) injury in rats and the relationship with inflammatory responses,lipid peroxidation and cell apoptosis.Methods Thirty clean-grade healthy male Sprague-Dawley rats,aged 2-3 months,weighing 150-270 g,were divided into 3 groups (n =10 each) using a random number table method:sham operation group (Sham group),intestinal Ⅰ/R group (Ⅱ/R group) and tryptase inhibitor group (PRTM group).Intestinal Ⅰ/R was produced by occlusion of the superior mesenteric artery for 75 min followed by 4-h reperfusion in Ⅱ/R and PRTM groups.The superior mesenteric artery was only exposed but not occluded in Sham group.Protamine 2.5 mg/kg was injected via the caudal vein at 5 min of before reperfusion in PRTM group.The rats were sacrificed at the end of reperfusion,and the small intestinal tissues 1 cm in length 5 cm away from the terminal ileum were removed for examination of the pathological changes (with a light microscope) and for determination of intestinal mucosal mast cell (IMMC) count and expression of tryptase in mast cells (by immunohistochemical SP staining),malondialdehyde (MDA) content (using thiobarbituric acid assay),superoxide dismutase (SOD) activity (by xanthine oxidase method),expression of caspase-3 (by Western blot) and contents of tumor necrosis factor alpha (TNF-ot) and interleukin-6 (IL-6) (by enzyme-linked immunosorbent assay).The degree of intestinal tissue damage was graded using Chiu's scoring system.Results Compared with Sham group,the pathological changes were significantly accentuated,Chiu's score was increased,the expression of tryptase was up-regulated,the IMMC count and contents of TNF-α,IL-6 and MDA were increased,the activity of SOD was decreased,and the expression of caspase-3 was up-regulated in Ⅱ/R and PRTM groups (P＜0.05).Compared with Ⅱ/R group,the pathological changes were significantly attenuated,Chiu's score was decreased,the expression of tryptase was down-regulated,the IMMC count and contents of TNF-α,IL-6 and MDA were decreased,the activity of SOD was increased,and the expression of caspase-3 was down-regulated in PRTM group (P＜0.05).Conclusion Tryptase is involved in the process of intestinal Ⅰ/R injury though promoting inflammatory responses,lipid peroxidation and cell apoptosis in rats.
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Anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) is a group of autoimmune diseases with rapid progession and poor prognosis.The histopathological hallmark in the kidneys of patients with AAV is " pauci-immune" necrotizing crescentic glomerulonephritis,therefore complement was previously suggested not to play a major role in the disease development.However,in the recent years,accumulating evidence from both experimental and clinic study has strongly incriminated alternative complement pathway activation as critically important in the pathogenesis of AAV.In patients with AAV,plasma levels of Bb and FH,components of the alternative complement pathway,are associated with disease activity and prognosis,which might be useful biomarkers in monitoring systemic disease activity and renal disease activity in AAV.The complement activation product C5a and its interaction with C5a receptor play a central role.It is suggested that neutrophils,ANCA and complement system form a positive feedback loop contributing to the development of AAV.Preliminary data of two clinical trials have demonstrated effectiveness and safety of C5a receptor blockade in patients with AAV.Therefore,measurement of complement biomarkers will be helpful in assessing the disease activity of patients and be of great importance for monitoring the efficiency of complement-targeting therapy in the future.
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Objective To explore the relationship between serum osteocalcin levels and glucolipid metabolism in elderly type 2 diabetic patients with non-alcoholic fatty liver disease (NAFLD).Methods Data collected from 97 patients with type 2 diabetes mellitus(T2DM)admitted to the Department of Geriatric Endocrinology of the First Affiliated Hospital of Zhengzhou University from June 2014 to April 2015 were retrospectively analyzed.Patients were divided into the T2DM group(type 2 diabetic patients without NAFLD,n＝ 47)and the NAFLD group(T2DM patients with NAFLD,N ＝ 50).Healthy elderly subjects (n ＝ 30)from the same period served as the control group.Body mass index(BMl),osteocalcin,fasting blood glucose,fasting insulin,homeostasis model assessment for insulin secretion index (HOMA-β) and insulin resistance (HOMA-IR),glycosylated hemoglobin(HbA1c),total cholesterol,triglyceride,high-density lipoprotein cholesterol(HDL-C)and low-density lipoprotein cholesterol(LDL C)were compared between the 3 groups.Results Levels of fasting blood glucose,fasting insulin,HbAlc,total cholesterol,triglyceride,LDL-C and HOMA IR were higher,while levels of HDL-C,HOMA-β and osteocalcin were lower in the T2DM and NAFLD groups than in the control group(all P＜＜0.05).Levels of BMI,fasting glucose,fasting insulin,HbAlc,total cholesterol,triglyceride,LDL-C and HOMA-IR were higher and levels of osteocalcin were lower in the T2DM group than in the NAFLD group(all P＜0.05).Pearson correlation analysis showed that the osteocalcin level was negatively correlated with fasting blood glucose,HbA1C,HOMA IR and BMI(r＝-0.701,0.442,-0.337 and 0.543,P＜0.05 or P＜0.01),and positively correlated with HOMA-β (r ＝ 0.341,P ＜ 0.05) in the NAFLD group.With serum osteocalcin as the dependent variable,multiple linear regression results showed that fasting blood glucose was an independent influencing factor for serum osteocalcin(β＝-1.57,P＜0.05)in the fatty liver group.Conclusions Serum osteocalcin levels significantly decrease in elderly T2DM patients with NAFLD,are closely correlated with glucolipid metabolism,and may have some important clinical significance in the prevention and treatment of NAFLD in elderly patients with type 2 diabetes.
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BACKGROUND:Mechanical properties of the blood vessels include axial and radial tension which is important to guarantee the tissue perfusion and its pressure. OBJECTIVE:To test and analyze stretching and puncture parameters that can ful y characterize the performance of blood vessels, so as to predict the health condition of the vessels transplanted into recipients. METHODS:Mechanical properties as circumferential tension, axial tension and puncture were tested on the porcine aorta, and then pathological and staining analysis was done to explore whether freeze-dried vessels can withstand the blood pressure in al directions after transplanted into the body. RESULTS AND CONCLUSION:Specific trend on the curves of freeze-dried vessels during circumferential tension, axial tension and puncture process was closer to that of fresh vessel curves, suggesting that structural components and arrangement of the freeze-dried vessels were not changed greatly. Meanwhile, the microscopic observations of slice and tensile section were taken into account so that the reason was studied and explained for the mechanical properties of the experimental data and results. The macroscopic mechanical result was analyzed based on the change of microstructure. These findings indicate that the vacuum freeze-drying is a better method to maintain the performance of blood vessels.
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To predict and identify the dominant B‐cell epitopes of conserved region of Treponema pallidum repeat protein F (TprFN ) and provide the basis for development of polyvalent epitope‐based syphilis vaccine ,the amino acid sequence of TprFN was obtained from GenBank and analyzed with comprehensive meta‐analysis Mobyle ,ABCpred and IEDB online software .The peptides containing predicted epitopes were artificially synthesized . To obtain and measure the titers of antibodies against TprFN ,New Zealand rabbits were immunized with recombinant protein TprFN expressed in E .coli and identified by Western blot (WB) .Sera from TprFN‐immunized rabbits ,syphilis patients ,and normal human and normal rabbits were used to deter‐mine the immunoreactivity and specificity of 7 predicted peptides of TpFN by indirect ELISA .Comprehensive meta‐analysis of online software showed that P1 (43‐62AA) ,P2(57‐71AA) ,P3(81‐88AA) ,P4(89‐103AA) ,P5(125‐138AA) ,P6(231‐251AA) and P7(268‐279AA) might be the B‐cell epitopes .A protein was expressed in a soluble form and identified as TpFN by WB .The ELISA indicated that P1 and P3 were active with TprFN‐immunized rabbit sera and syphilis patient sera but not with negative control sera .These results indicate that P1 and P3 are the potential dominant B‐cell epitopes .
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ObjectiveTo observe the effect and safety of arsenic trioxide (ATO) combined with all-trans retinoic acid (ATRA) in treating patients with acute promyelocytic leukemia (APL).Methods Eighty-three cases with APL treated for the first time were divided into two groups by random digits table method:observation group with 48 cases was received combination induction treatment of ATO and ATRA,control group with 35 cases was treated with combination induction treatment of ATRA and chemotherapy.The clinical effect and adverse reaction between two groups were compared.ResultsThe effective rate and early death rate were 100.0％( 48/48 ) and 0 in observation group,97.1％(34/35 ) and 2.9％( 1/35 ) in control group,which had no significant difference between two groups(P ＞ 0.05 ).The incidences of bone marrow suppression,infection,liver and kidney damage,cardiac toxicity and gastrointestinal symptoms were 8.3％ (4/48),10.4％ (5/48),12.5％ (6/48),6.2％ (3/48) and 18.8％ (9/48) in observation group,while 97.1％(34/35),65.7％(23/35),45.7％(16/35),37.1％(13/35) and 100.0％(35/35) in control group,which had significant differences between two groups (P ＜ 0.05).ConclusionCombination treatment of ATO and ATRA in APL has an obvious effect and few adverse reaction,which can be applied in clinic.
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ObjectiveTo determine the subcellular localization of exogenous human papillomavirus type 16 E6 protein(HPV16 E6) and hDaxx in HeLa cells and their effects on tumor necrosis factor (TNF)-α-induced apoptosis.MethodsHeLa cells were transfected with plasmids pDsRed-monomer-C1/HPV16 E6,pEGFP-CI/hDaxx,pEGFP-C1 and pDsRed-monomer-C1 respectively.Subsequently,Western blot was carried out to quantify the expression of fusion proteins DsRed-HPV16E6 and EGFP-hDaxx in transfected cells,and laser scanning confocal microscopy to observe the subcellular distribution of HPV16 E6 protein and hDaxx.Some HeLa cells were divided into 5 groups:untransfected (control group),untransfected and treated with TNF-α(TNF-ot group),transfected with pcDNA3.1 (-) and treated with TNF-α(empty vector group),transfected with pcDNA3.1 (-)/HPV16 E6 and treated with TNF-α (HPV16 E6 group),cotransfected with pcDNA3.1(-)/HPV16 E6 and pcDNA3.1 (-)/hDaxx and treated with TNF-α (cotransfected group).After additional culture,the cells were collected and subjected to flow cytometry(FCM) to evaluate the apoptosis of cells as well as spectrophotometry to determine the relative activity of Caspase-8 and Caspase-3.ResultsWestern blot showed that both DsRed-HPV16 E6 and EGFP-hDaxx were expressed in HeLa cells.In Hela cells transfected with pDsRedmonomer-C1/HPV16 E6 or pEGFP-C1/hDaxx alone,the red fluorescence of HPV16 E6 was observed in the nucleus and cytoplasm,while the green fluorescence of hDaxx only in the nucleus; in those cotransfected with pDsRed-monomer-C1/HPVl6 E6,HPV16 E6 and hDaxx proteins were regionally aggregated near the nuclear membrane in nuclei,and hDaxx was partly translocated from the nucleus to the cytoplasm.The apoptosis rate and relative activity of Caspase-8 and Caspase-3 were statistically lower in HPV16 E6 group than in the empty vector group and cotransfected group(21.4％ ± 1.1％ vs.27.0％ ± 0.9％ and 32.5％ ± 2.1％,0.057 ± 0.003 vs.0.092 ±0.012 and 0.109 ± 0.013,0.054 ± 0.006 vs.0.093 ± 0.005 and 0.110 ± 0.004,all p＜ 0.01).Conclusions HPV16 E6 protein induces the partial translocation of hDaxx from the nucleus into the cytoplasm and colocalizes with hDaxx in the cells.The apoptosis of HeLa cells induced by TNF-α can be suppressed by HPV16 E6 protein,while the overexpression of hDaxx can attenuate the suppressing effect of HPV16 E6 protein on apoptosis in Hela cells.
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Objective To investigate the prevalence and features of thyroid nodules in middle and aged patients with type 2 diabetes mellitus(T2DM). Methods High-resolution ultrasonography was used to detect thyroid nodules in 132 cases middle and aged patients with type 2 diabetes and 89 patients without diabetes.The nodule features and its relationships with related indicators in diabetic patients were analyzed. Results The prevalence of thyroid nodules in middle and aged patients with type 2 diabetes was higher than that without diabetes (67.4％ vs. 53.9％,P＜0.05),and most occurred in 50 to 59 age group (66.7％ vs. 42.9％) without dependence on changes in thyroid functions and volumes.In diabetes group,the prevalence of thyroid nodules were 59.5％ in male and 81.3％ in female (P＜0.05),no obvious difference was observed in the size and number of thyroid nodules between male and female,multiple nodules and micronodule (＜ 1.0 cm) had the higher incidences in both sexes.The prevalence of thyroid nodules was increased with aging,but not with diabetes duration and glycosylated hemoglobin (HbAlc) level (x2 =0.797,P=0.372; x2 =1.078,P =0.229). Conclusions It is common that thyroid nodules combined with diabetes in middle and aged patients,thyroid ultrasound screening and regular following-up of patients aged ≥50 years have important clinical significance.
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Objective To investigate the characteristics of the blood glucose fluctuation in elderly patients with type 2 diabetes mellitus (T2DM). Methods The 92 elderly patients with T2DM (the elderly group) and 58 young and middle-aged patients with T2DM (the non-elderly group) were monitored using the continuous glucose monitoring system(CGMS). The characteristics of glucose profiles of the two different age groups, and of the different glycosylated hemoglobin (HbA1c) level groups in the elderly were comparatively analyzed. Results (1)There was no significant difference in HbA1c level between the elderly group and the non-elderly group. Compared with the non-elderly group, the elderly group showed the increases in blood glucose fluctuant coefficient [BGFC, (2.68±1.00) mmol/L vs. (2.12±0.74) mmol/L, t=-3.691, P＜0.001], in postprandial glucose excursion (PPGE) of breakfast and supper [(5.96±2.47) mmol/L vs. (5.11±2.44) mmol/L, t=-2.058, P＜0.05; (5.17±2.15) mmol/L vs. (4.16±2.28) mmol/L, t=-2.730, P＜0.01], in the time to postprandial glucose peak of breakfast and lunch [(112.5±29.7) min vs. (97.0±27.2) min, t=-3.225, P＜0.01; (140.0±39.7) min vs. (118.1±42.6) min, t=-3.195, P＜0.01], in the frequency of hypoglycemia (26.3% vs. 5.5%, P＜0.05), and showed the largest amplitude of glycemic excursions [LAGE, (9.66±2.48) mmol/L vs.(8.40±3.13) mmol/L, t=-2.720, P＜0.01]. (2)In the elderly, along with decreased HbA1c, the incidence of hypoglycaemia increased (P＜0.05); And along with increased HbA1c, the amplitude of blood glucose fluctuation increased. There were significant differences in BGFC, PPGE of breakfast and lunch, and LAGE among different HbA1c level groups (P＜0.01, P＜0.05, P＜0.05, P＜0.001). (3)HbA1c was positively correlated with FBG, mean blood glucose (MBG), percentage of time at glycemia (PT7.8, PT11.1), the lowest blood glucose (LBG), the highest blood glucose (HBG), BGFC, PPGE and LAGE (r=0.899-0.289, all P＜0.001). Multiple stepwise regression analysis indicated that MBG, FBG and PT7.8 was the independent influential factor of HbA1c (adjusted R2=0.807, P＜0.05). Conclusions The elderly patients with T2DM are at a particularly high risk for postprandial hyperglycemia and nocturnal hypoglycemic episodes, CGMS could show glucose fluctuation characters of T2DM patients diurnally, and provide a clinical basis for reasonable therapy.