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Objective To establish a quantitative polymerase chain reaction(PCR)method for the analysis of human-derived SRY DNA in mouse tissues,and to study the tissue distribution of human umbilical cord mesenchymal stem cells(HUCMSCs)in immunodeficient NOG mice after a single intravenous injection.Methods We established a quantitative PCR method for the analysis of human SRY DNA in mouse tissues,and validated the standard curve,linear range,accuracy,precision,and stability.Thirty-six NOG mice(18 male,18 female)were administered 3.5×107 HUCMSCs/kg by single intravenous injection.Six mice were then anesthetized and dissected after blood collection(EDTA anticoagulation)at 6,12,24,and 72 h,and at 1 and 2 weeks,respectively.DNA was extracted from lung,kidney,heart,liver,brain,spinal cord,stomach,small intestine,fat,skin,spleen,testis,uterus,and ovary tissues,and the distribution of HUCMSCs in each tissue was determined by the validated quantitative PCR method for detecting the human-derived SRY gene in mouse tissues.In addition,18 NOG mice(9 male,9 female)were divided into control(n = 6)and treatment groups(n = 12)injected intravenously with 0.9%sodium chloride and 3.5×107 cells/kg,respectively.Acute toxic reactions were observed during the administration period,and four animals were dissected at 72 h and at 2 and 4 weeks after administration to observe the gross organs.Mitochondrial protein expression was detected in paraffin sections of lung tissues by immunohistochemistry to analyze the colonization of HUCMSCs in lung tissues.Results The established RT-qPCR method for human-derived SRY DNA in mouse tissues met the validation criteria for each index.After a single intravenous injection in NOG mice,HUCMSCs were mainly distributed in the lungs and blood within 1 week after administration,with higher concentrations in lung tissues than in blood.The concentrations of HUCMSCs in lung tissue and blood remained relatively stable within 6～24 h and 6～72 h,respectively,and then decreased over time.The distribution of HUCMSCs in other tissues was not measured at all sampling points.The colonization result showed that HUCMSCs were detected in lungs 72 h after intravenous injection,but not at 2 and 4 weeks.No obvious acute toxicity was observed in NOG mice after single intravenous administration of HUCMSCs.Conclusions The above method for analyzing the distribution of HUCMSCs in mouse tissue is reliable and feasible.HUCMSCs were mainly distributed in lung and blood in NOG mice within 1 week after a single intravenous injection,and mainly colonized lung tissue at 72 h.A single intravenous administration of HUCMSCs has a good safety profile.
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Objective In this study,we explored the role of combination of autophagy inhibition and anti-VEGF in proliferation,migration and tube formation of mouse retinal vascular endothelial cells(RVECs). Methods Well cultured mouse RVECs were randomly divided into four groups:autophagy inhibition group(add-ing autophagy inhibitor 3-MA),anti-VEGF group(adding anti-VEGF-A neutralized antibody),autophagy inhibi-tion+anti-VEGF group(adding the two reagents)and the control group.All cells were then cultured in the hypoxic condition. The cell proliferation,migration and tube formation were detected by EdU,transwell and matrigel as-say,respectively. Results The cell proliferation rate,number of migrated cells and number of tube formation of the other three groups decreased when compared with the control group.These data above in autophagy inhibition+anti-VEGF group were all significantly less than 3-MA group and anti-VEGF group. Conclusion Combination of autophagy inhibition and anti-VEGF may be more effective than simple anti-VEGF in inhibition of retinal neovascu-larization.
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Objective To study the association of the serum alanine aminotransferase (ALT) level with cardiovascular risks in patients with type 2 diabetes mellitus (T2DM).Methods A total of 676 patients with T2DM who excluded liver disease due to chronic viral hepatitis,autoimmune,drugs,alcohol,and other hereditary were selected.According to the ALT level,676 patients were assigned into elevated ALT group (ALT ＞40 U/L,126 cases) and normal ALT group (ALT 5-40 U/L,550 cases).And normal ALT group were divided into 4 groups on the interquartile distance:group 1 (ALT 5-11 U/L,108 cases),group 2 (ALT 12-15 U/L,114 cases),group 3 (ALT 16-21 U/L,127 cases),group 4 (ALT 22-40 U/L,201cases).The age,duration of diabetes,body mass index (BMI),waist-to-hip ratio (WHR),blood pressure,triglyceride (TG),total cholesterol (TC),high density lipoprotein cholesferol (HDL-C),low density lipoprotein cholesferol (LDL-C),fasting plasma glucose (FPG),2 h postprandial blood glucose (2 hPG),glycosylated hemoglobin (HbA1c),uric acid (UA),fasting insulin and homeostasis model assessment insulin resistance index (HOMA-IR) was compared among these groups.Results The levels of BMI,WHR,diastolic blood pressure,FPG,2 hPG,HbA1c,fasting insulin,TG,HOMA-IR and UA in elevated ALT group were higher than those in normal ALT group [(26.80 ± 3.60) kg/m2 vs.(23.40 ± 3.50) kg/m2,0.95 ±0.05 vs.0.91 ± 0.05,(75.3 ± 9.3) mm Hg (1 mm Hg =0.133 kPa) vs.(69.2 ± 9.5)mm Hg,(9.24 ± 2.56)mmol/L vs.(8.53 ± 2.78) mmol/L,(15.25 ± 4.62) mmol/L vs.(14.37 ± 5.06) mmoUL,(8.31 ± 1.76)％ vs.(7.78 ± 1.94)％,(10.76 ± 6.78) mU/L vs.(7.85 ± 7.04) mU/L,(2.73 ± 2.40) mmol/L vs.(2.20 ± 2.78)mmol/L,4.13 ± 2.76 vs.3.06 ± 2.63,(332.42 ± 95.12) μ mol/L vs.(285.14 ± 79.53) μ mol/L] (P ＜ 0.05),and HDL-C was lower than that in normal ALT group [(1.06 ± 0.27) mmol/L vs.(1.15 ± 0.57) mmol/L] (P ＜0.05).Compared with that in normal ALT group,the age was younger,and the duration of diabetes was shorter in elevated ALT group [(49.6 ± 10.5) years vs.(57.5 ± 11.3) years,(37.9 ±42.0) months vs.(54.5 ± 57.8) months] (P ＜ 0.05).The levels of FPG,2 hPG,HbA1c and UA in group 4 were higher than those in group 1,2,3 [(9.18 ± 3.01) mmol/L vs.(8.33 ± 2.57),(8.38 ± 2.52),(8.45 ± 2.73) mmol/L;(14.56 ± 4.80) mmol/L vs.(13.15 ± 5.72),(13.42 ± 4.56),(13.50 ± 5.17) mmol/L; (7.81 ± 1.98)％ vs.(6.76 ± 1.84)％,(6.79 ± 2.10)％,(6.88 ± 2.43)％; (288.24 ± 78.26) μ mol/L vs.(271.15 ± 75.43),(273.27 ± 71.25),(275.56 ± 69.57) μ mol/L] (P ＜ 0.05) ; HOMA-IR was lower than that in group 1,2,3(2.67 ± 2.65 vs.2.84 ± 2.53,2.98 ± 3.10,3.12 ± 2.57,P ＜ 0.05).Conclusion TheincreaseofALTlevel is associated with the clustered cardiovascular risk factors and insulin resistance in patients with T2DM.
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Encouraging voluntary donation of citizens' organ is the best institutional solution for the severe shortage of organ transplantation.The policy incentives should allow donors to rescind their donation of their organ anytime before the organ is removed from their body.As long as the donor does not commit subjective offence,he/she shall be released from any legal responsibilities.This arrangement is highly significant legally and practically.