Objective To establish the sperm specific Sleeping Beauty ( SB ) transposase-expression transgenic mouse for the study of the genetic modification mediated by transposon system in mouse .Methods Prm1 promoter was cloned from mouse genomic DNA to drive the expression of SB transposase .The transgenic mice were generated by microinjection .The gene type of transgenic line was identified by PCR .The expressing level in testis was determined by western blot and immunohistochemistry (IHC) staining.Results Five lines of transposase transgenic mice were obtained by microinjection and three can be germline .One mouse line with higher expression level of transposase in the testis was obtained.Conclusion One transgenic mouse model with Sleeping Beauty transposase - expression was successfully established .This model will greatly contribute to the research of genetic modification mediated by transposon in mouse.

Objective To study the relationship of insulin receptor substrate-1 (Irs1) and metabolic disease, we generated Irs1 gene knockout rat by CRISPR/Cas9 system.Methods Two sgRNA targeting sites were designed for Irs1 targeting.The Cas9 and sgRNAs were transcribed by T7 RNA polymerase in vitro.Cas9 mRNA and sgRNA mixtures were pooled and microinjected into one-cell fertilized eggs of SD rats to generate rats with targeted mutation .Results Five rats with the mutations were detected with the efficiency of 83%.Conclusion The Irs1 gene knockout rats generated in this study can be transmitted by germline .

Objective To study the effects of NRG2 on cardiac structure and function , we established the cardiac-specific human NRG2 transgenic mice and investigate the effect of NRG2 on cardiac structure and function under pressure overload situation .Methods The transgenic vector was constructed by insertion of the human NRG2 gene under the α-MHC promoter.The transgenic mice were generated by microinjection and were all maintained on a C57BL/6J genetic background .The genotype of transgenic mice was identified by PCR and the expression level of target gene was determined by western blot .Transverse aortic constriction ( TAC) was applied to prepare the pressure overload induced cardiomyopathy mice model .The cardiac structure and function of the transgenic mice were compared and analysized by echocardiographic and pathological observation .Results Transgenic mice with high level of NRG2 in heart tissues were established.The left ventricular wall thickness (LVPWD) was increased, and to 15.6% at 3 months old compared with that of the non transgenic ( NTG) mice.The hypertrophy of left ventricular wall caused by pressure overload was removed due to the expression of NRG2 .Meanwhile, cardiac disarray and fibrosis were increased obviously compared with that of the NTG mice.Conclusion The transgenic expression of NRG2 in heart tissues could shorten the pathological process of hypertrophy, but accelerated the process of heart failure (HF).

Objective To knock out the Abcb1 gene of rat,and establish the Abcb1 humanized rat model based on the Abcb1 knock out rat.Methods The animal model was established using BAC and CRISPR/Cas9 technology,and was analyzed by PCR, RT-PCR and real-time PCR.Results Establishing a rat model expressing human Abcb1 stably by transfer the 153 kb BAC containing human Abcb1 promoter and cDNA into rat genome, and establishing the Abcb1 knock out rat at the same time.Establishing the Abcb1 humanized model by crossing these two strains together.The expression pattern of Abcb1 in Abcb1 humanized rat is different from the wild type rat.The Abcb1 humanized model express not only the human Abcb1 gene but also has similar expression pattern as human.Conclusions The Abcb1 knock out rat and the Abcb1 humanized rat were successfully established, and this model is close to human concerning about the drug metabolism related to Abcb1.

bThis study explored whether the transplantation of modified marrow stromal cells (MSCs) has angiogenic effects in a left middle cerebral artery occlusion infarction/reperfusion (MCAO I/R) rat model and preliminarily examined the mechanism of angiogenesis following cerebral infarction. MSCs were isolated by using a direct adherent method and cultured. Vascular endothelial growth factor (VEGF) was transfected into MSCs by employing the liposome transfection. The transfection efficiency was measured by the optical density method. The protein expression of VEGF gene before and after transfection was measured by Western blotting. SD rat model of transient occlusion of the left middle cerebral artery was established by using an approach of intra-luminal occlusion. Tetrazolium (TTC) and HE staining were performed to observe the cerebral infarction. ELISAs were used to measure the levels of VEGF in the rat cerebral tissues. The expression patterns of angiopoietin-2 (Ang-2) and CD34 in cells surrounding the area of infarction were immunohistochemistrically observed. Ang-2 protein expression in the tissue surrounding the area of infarction was measured by Western blotting. VEGF expression in the MSCs increased after transfection at a rate of approximately 28%±3.4%. ELISA showed that the expression of VEGF in the cerebral tissue was significantly increased after induction of infarction, peaking on the 4th day and decreasing to the levels of the sham surgery group (normal) within 7 to 10 days. The VEGF level was significantly higher at each time point in the VEGF-MSC and MSC groups compared to the model group. Moreover, the VEGF level was higher in the VEGF-MSC group than in the MSC group and stayed relatively high until the 10th day. The immunohistochemical results showed that 10 days after the infarction, the number of Ang-2 and CD34-expressing cells in the area surrounding the infarction was significantly higher in the VEGF-MSC group and the MSC group compared to the model group. Moreover, the VEGF level was higher in the VEGF-MSC group than the MSC group. A similar trend in Ang-2 protein expression was revealed by Western blotting. In the MCAO rat model transfected with modified MSCs over-expressing VEGF, compared to the MSC transplantation group, the concentration of VEGF was significantly increased in the brain tissue after cerebral infarction. In addition, the level of Ang-2 was up-regulated, with angiogenesis promoted, the blood supply to the areas surrounding the cerebral infarction increased, and neurological function improved. We are led to speculate that the synergistic effects of VEGF and Ang-2 may be responsible for the angiogenesis following cerebral infarction.

bThis study explored whether the transplantation of modified marrow stromal cells (MSCs) has angiogenic effects in a left middle cerebral artery occlusion infarction/reperfusion (MCAO I/R) rat model and preliminarily examined the mechanism of angiogenesis following cerebral infarction. MSCs were isolated by using a direct adherent method and cultured. Vascular endothelial growth factor (VEGF) was transfected into MSCs by employing the liposome transfection. The transfection efficiency was measured by the optical density method. The protein expression of VEGF gene before and after transfection was measured by Western blotting. SD rat model of transient occlusion of the left middle cerebral artery was established by using an approach of intra-luminal occlusion. Tetrazolium (TTC) and HE staining were performed to observe the cerebral infarction. ELISAs were used to measure the levels of VEGF in the rat cerebral tissues. The expression patterns of angiopoietin-2 (Ang-2) and CD34 in cells surrounding the area of infarction were immunohistochemistrically observed. Ang-2 protein expression in the tissue surrounding the area of infarction was measured by Western blotting. VEGF expression in the MSCs increased after transfection at a rate of approximately 28%±3.4%. ELISA showed that the expression of VEGF in the cerebral tissue was significantly increased after induction of infarction, peaking on the 4th day and decreasing to the levels of the sham surgery group (normal) within 7 to 10 days. The VEGF level was significantly higher at each time point in the VEGF-MSC and MSC groups compared to the model group. Moreover, the VEGF level was higher in the VEGF-MSC group than in the MSC group and stayed relatively high until the 10th day. The immunohistochemical results showed that 10 days after the infarction, the number of Ang-2 and CD34-expressing cells in the area surrounding the infarction was significantly higher in the VEGF-MSC group and the MSC group compared to the model group. Moreover, the VEGF level was higher in the VEGF-MSC group than the MSC group. A similar trend in Ang-2 protein expression was revealed by Western blotting. In the MCAO rat model transfected with modified MSCs over-expressing VEGF, compared to the MSC transplantation group, the concentration of VEGF was significantly increased in the brain tissue after cerebral infarction. In addition, the level of Ang-2 was up-regulated, with angiogenesis promoted, the blood supply to the areas surrounding the cerebral infarction increased, and neurological function improved. We are led to speculate that the synergistic effects of VEGF and Ang-2 may be responsible for the angiogenesis following cerebral infarction.
Angiopoietin-2 ; genetics ; metabolism ; Animals ; Bone Marrow ; metabolism ; pathology ; Cerebral Infarction ; genetics ; metabolism ; pathology ; Male ; Neovascularization, Pathologic ; genetics ; pathology ; Rats ; Rats, Sprague-Dawley ; Stromal Cells ; metabolism ; pathology ; Vascular Endothelial Growth Factor A ; genetics ; metabolism

Objective To examine the proportion and reasons of drop-out from antiretroviral therapy(ART)among 8 367 adult HIV-infected individuals in Dehong prefecture,Yunnan province. Methods All adult HIV-infected patients receiving ART before September 30 of 2014 were examined for the situation of drop-out from ART. Results The proportion of drop-out from ART among adult HIV-infected patients in Dehong prefecture was 14.4%(1 202/8 367). Results from the univariate logistic regression analyses indicated that drop-out from ART was significantly correlated with factors as:living area,gender,age,marital status,HIV transmission route,baseline CD4+T cell counts and initial treatment regimen of the patients. After adjusted for potential confounding variables by multiple logistic regression model,drop-out from ART was significantly correlated with residential area,marital status,HIV transmission route,baseline CD4+T cell count and initial treatment regimen of the patients. HIV-infected patients who were living in Mangshi city,Lianghe county or Yingjiang county,being married or living with partner,HIV infection through sexual contact,with baseline CD4+T cell counts≤200 cells/mm3,and ART included in the initial treatment regimen etc.,were less likely to drop out from ART. The proportion of drop out from ART was significantly decreasing along with the increasing time of ART. Data from specific investigation revealed that among the 1 202 patients who dropped out from ART,704(58.6%)were lost to follow-up,303(25.2%) did not adhere to treatment,74(6.2%)moved out the region,64(5.3%)were Burmese that had returned to Burma,29 (2.4%) stopped the treatment according to doctors’advice,18(1.5%)were incarcerated and 10 (0.8%) were under other reasons. Reasons for the drop-out varied,according to the situation of patients. Conclusion The proportion of drop-out from ART varied significantly according to the characteristics of HIV-infected patients in Dehong prefecture that underscoring the needs for tailored responses to reduce drop-out of ART. Focus should be targeted on reducing the loss to follow-up and improving the treatment adherence.

Objective To examine the proportion and reasons of drop-out from antiretroviral therapy(ART)among 8 367 adult HIV-infected individuals in Dehong prefecture,Yunnan province. Methods All adult HIV-infected patients receiving ART before September 30 of 2014 were examined for the situation of drop-out from ART. Results The proportion of drop-out from ART among adult HIV-infected patients in Dehong prefecture was 14.4%(1 202/8 367). Results from the univariate logistic regression analyses indicated that drop-out from ART was significantly correlated with factors as:living area,gender,age,marital status,HIV transmission route,baseline CD4+T cell counts and initial treatment regimen of the patients. After adjusted for potential confounding variables by multiple logistic regression model,drop-out from ART was significantly correlated with residential area,marital status,HIV transmission route,baseline CD4+T cell count and initial treatment regimen of the patients. HIV-infected patients who were living in Mangshi city,Lianghe county or Yingjiang county,being married or living with partner,HIV infection through sexual contact,with baseline CD4+T cell counts≤200 cells/mm3,and ART included in the initial treatment regimen etc.,were less likely to drop out from ART. The proportion of drop out from ART was significantly decreasing along with the increasing time of ART. Data from specific investigation revealed that among the 1 202 patients who dropped out from ART,704(58.6%)were lost to follow-up,303(25.2%) did not adhere to treatment,74(6.2%)moved out the region,64(5.3%)were Burmese that had returned to Burma,29 (2.4%) stopped the treatment according to doctors’advice,18(1.5%)were incarcerated and 10 (0.8%) were under other reasons. Reasons for the drop-out varied,according to the situation of patients. Conclusion The proportion of drop-out from ART varied significantly according to the characteristics of HIV-infected patients in Dehong prefecture that underscoring the needs for tailored responses to reduce drop-out of ART. Focus should be targeted on reducing the loss to follow-up and improving the treatment adherence.

OBJECTIVETo examine the proportion and reasons of drop-out from antiretroviral therapy (ART) among 8 367 adult HIV-infected individuals in Dehong prefecture, Yunnan province.

METHODSAll adult HIV-infected patients receiving ART before September 30 of 2014 were examined for the situation of drop-out from ART.

RESULTSThe proportion of drop-out from ART among adult HIV-infected patients in Dehong prefecture was 14.4% (1 202/8 367). Results from the univariate logistic regression analyses indicated that drop-out from ART was significantly correlated with factors as: living area, gender, age, marital status, HIV transmission route, baseline CD4⁺ T cell counts and initial treatment regimen of the patients. After adjusted for potential confounding variables by multiple logistic regression model, drop-out from ART was significantly correlated with residential area, marital status, HIV transmission route, baseline CD4⁺ T cell count and initial treatment regimen of the patients. HIV-infected patients who were living in Mangshi city, Lianghe county or Yingjiang County, being married or living with partner, HIV infection through sexual contact, with baseline CD4⁺ T cell counts ≤ 200 cells/mm³, and ART included in the initial treatment regimen etc., were less likely to drop out from ART. The proportion of drop out from ART was significantly decreasing along with the increasing time of ART. Data from specific investigation revealed that among the 1 202 patients who dropped out from ART, 704 (58.6%) were lost to follow-up, 303 (25.2%) did not adhere to treatment, 74 (6.2%) moved out the region, 64 (5.3%) were Burmese that had returned to Burma, 29 (2.4%) stopped the treatment according to doctors' advice, 18 (1.5%) were incarcerated and 10 (0.8%) were under other reasons. Reasons for the drop-out varied, according to the situation of patients.

CONCLUSIONThe proportion of drop-out from ART varied significantly according to the characteristics of HIV-infected patients in Dehong prefecture that underscoring the needs for tailored responses to reduce drop-out of ART. Focus should be targeted on reducing the loss to follow-up and improving the treatment adherence.

Adult ; Anti-Retroviral Agents ; therapeutic use ; China ; HIV Infections ; drug therapy ; Humans ; Patient Dropouts ; statistics & numerical data

OBJECTIVETo understand HIV rival suppression and drug resistance (HIVDR) among AIDS patients who were receiving antiretroviral treatment (ART) in Dehong prefecture, Yunnan province.

METHODSAll AIDS patients who were aged over 15 years and with experience more than six months on ART by the end of 2012 in Dehong prefecture, were enrolled to receive testing for HIV viral load in plasma and genetic mutations associated with HIVDR.

RESULTSA total of 4 390 AIDS patients were qualified for the study according to the selection criteria, of whom 3 964 (90.3%) finally participated in the study. Among them, 2 307(58.2%) had CD₄(+) cell counts more than 350 cells/mm³. 3 169 (79.9%) patients showed undetectable plasma HIV viral load which was lower than the detection threshold. Those who had the following factors as:resided in Ruili city, being female, older than 45 years of age, married, heterosexually infected with HIV, having received ART more than 5 years, and CD₄(+) cell counts >500 cells/mm³, were more likely to have undetectable plasma virus load, with the differences statistically significant. 402 (10.1%) patients had plasma viral load ≥ 1 000 copies/ml, of whom 353 (87.8%) were successfully amplified and examined for HIVDR. Among them, 198 (56.1% ) were identified to bear genetic mutations associated with HIVDR. Most mutations were related to the resistance to nucleotide reverse transcriptase inhibitors (NNRTIs) or non-nucleotide reverse transcriptase inhibitors (NNRTIs), with M184V and K103N most frequently seen. 12 patients (3.4%) were found to have mutations resistant to protease inhibitors (PI). Data from multiple logistic regression analysis indicated that the period of receiving ART and the initial ART regimen could both significantly predict the occurrence of HIV resistance.

CONCLUSIONViral suppression was highly achieved among ART-prescribed AIDS patients in Dehong prefecture,Yunnan province. However, among those who did not show effective viral suppression, the proportion of HIVDR was high, underscoring the needs for health education so as to improve the adherence to drugs as well as for improving testing for viral load and HIVDR among AIDS patients.

Acquired Immunodeficiency Syndrome ; drug therapy ; Adolescent ; Adult ; Anti-HIV Agents ; therapeutic use ; Drug Resistance, Viral ; genetics ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Mutation ; Viral Load ; drug effects ; Young Adult