Usually it is rather difficult to obtain and retain an ideal phenotype in an animal breeding program.Inspired by the success in transplanting cellular nuclei of amphibian and fish species since the 1950s,scientists be...Usually it is rather difficult to obtain and retain an ideal phenotype in an animal breeding program.Inspired by the success in transplanting cellular nuclei of amphibian and fish species since the 1950s,scientists began to believe in the possibility that cloned mammals could be acquired as well in the same way.Some 20 years prior to 1986 when the British biologist S.M.Willadson succeeded in breeding the first sheep by nuclear transplant,Prof.T.C.Tung (Tong Dizhou), a famous Chinese exper-展开更多
BACKGROUND： Acute cellular rejection（ACR） after liver transplantation（LT） is one of the most common problems faced by transplant recipients in spite of advances in immunosuppressive therapy. Recently, clinical tr...BACKGROUND： Acute cellular rejection（ACR） after liver transplantation（LT） is one of the most common problems faced by transplant recipients in spite of advances in immunosuppressive therapy. Recently, clinical trials reported that ursodeoxycholic acid（UDCA） reduced the incidence of ACR significantly.However, others have shown contradictory conclusion. Therefore,we performed a meta-analysis of rigorous randomized controlled trials（RCTs） to determine the efficacy of UDCA in reducing ACR after LT.DATA SOURCES： All RCTs that evaluated efficacy of UDCA as an adjuvant treatment to prevent ACR after LT were searched from PubMed/MEDLINE, EMBASE, Cochrane Central Register of Controlled Trials, ScienceDirect databases and Web of Science（from January 1981 to March 2012）. There was no language limitation in these searches. Relevant abstracts of international meetings were also searched. References of each included study were searched manually.RESULTS： A total of 234 patients from four high-quality RCTs（Jadad score 4 to 5） were included in this meta-analysis.Prophylactic use of UDCA did not decrease the incidence of ACR（RR： 0.94, 95% CI： 0.77-1.16, P0.05）, steroid-resistant rejection（RR： 0.77, 95% CI： 0.47-1.27, P0.05） and the number of patients with the multiple episodes of ACR（RR： 0.60, 95% CI：0.28-1.30, P0.05）. Different intervention programs（high-dose vs low-dose UDCA; early vs delayed UDCA treatment） also did not alter the outcomes.CONCLUSIONS： UDCA, as an adjuvant treatment, was not ableto prevent ACR and steroid-resistant rejection after LT. Further trials should be done to determine whether higher dose of UDCA will be beneficial.展开更多
BACKGROUND： With developments of tissue engineering and genetic engineering, we aim to culture myoblasts, which are characterized by high purity, high quality and high production, for wide application in neural regen...BACKGROUND： With developments of tissue engineering and genetic engineering, we aim to culture myoblasts, which are characterized by high purity, high quality and high production, for wide application in neural regeneration researches. OBJECTIVE： To modify traditional dissociation method in order to obtain myoblasts, which are characterized by high purity, high quality and high production, and explore the biological properties under in vitro culture. DESIGN： Observational study. SETTING： Basic Institute of Academy of Military Medical Sciences of Chinese PLA. MATERIALS： Four neonatal Wistar rats of 5 days old, both genders and mean body mass of 10 g were selected in this study. The main reagents and devices were detailed as follows： DMEM medium （Gibco Company）, fetus bovine serum （FBS, Hycolne Company）, collagenase Ⅱ （Sigma Company）, trypsin （Sigma Company）, dispase Ⅱ （Sigma Company）, desmin antibody （Fuzhou Maixin Company）, antibody Ⅱ and ABC kit （Wuhan Baster Biotechnology Company）, desk centrifuge （KUBATO, Japan）, and inverted phase contrast microscope （LEICA DMIRB, Germany）. METHODS： The experiment was carried out in the Basic Institute of Academy of Military Medical Sciences of Chinese PLA from June to October 2006. Neonatal rats were sacrificed under sterile condition to obtain skeletal muscles of limbs, which were washed with cold PBS （containing benzylpenicillin and estreptomicina）, and muscular tissue was sheared into pieces. Then, those muscular pieces were added with mixed digestive enzyme （containing 2 g/L collagenase Ⅱ ＋ 5 g/L dispase Ⅱ ＋ 0.28 g/L CaCl2） as twice volume as pieces, dealt with mechanical pipetting for 5 minutes and cultured in CO2 incubator for 10 minutes. The operation was done for three times and the muscular pieces were digested for 45 minutes in total. Moreover, cells were suspended again in order to obtain myoblasts from skeletal muscle of neonatal rats. In addition, myoblasts were purified with differential attachment technique and enzyme digestion so as to observe morphological characteristics and growth, draw growth curve, analyze surface structure under scanning electron microscope, and evaluate with Desmin immunohistochemical staining. MAIN OUTCOME MEASURES： Morphological characteristics and growth ofmyoblasts cultured in vitro. RESULTS： ①Growth of myoblasts of skeletal muscle： Primary cells had well growth, mature and differentiation. The positive rate of Desmin was 94% and purification of cells was ideal. Growth curve of cells demonstrated that myoblasts which were characterized by high purification started proliferation plentiful through transient growth lag phase （about at one or two days after inoculation）. If myoblasts were not dealt with any interventions, they might become sarcotubule gradually at 3 - 5 days after proliferative phase. During this period, myoblasts maintained a monocaryon-bipolarity state under inverted phase contrast microscope. Furthermore, the growth of cells was the strongest and reproductive activity was the most powerful. This suggested that myotube started to form; in addition, muscle fiber of contractility might form under a well culturing condition. ②Immunocytochemical stain with desmin antibody： Interzonal fiber of desmin from myoblasts showed strongly positive reaction. Positive staining existed in cytoplasm had a high nucleus-cytoplasm ratio. However, myoblasts showed negative or mildly positive reaction. CONCLUSION： It is ideal for modified multi-enzymatic digestion and double purification method to dissociate and purify myoblasts of skeletal muscle; meanwhile, these two methods are both the effective ways to provide convenient conditions to obtain seed cells for neural regeneration researches.展开更多
Background The necrosis of a large number of myocardial cells after acute myocardial infarction （AMI） results in a decrease of cardiac function and ventricle remodeling. Stem cell transplantation could improve cardi...Background The necrosis of a large number of myocardial cells after acute myocardial infarction （AMI） results in a decrease of cardiac function and ventricle remodeling. Stem cell transplantation could improve cardiac function after AMI, but the involving mechanisms have not been completely understood. The present study aimed to investigate the effects of transplantation of autologous bone marrow mononuclear cells （BM-MNC） and mesenchymal stem cells （MSCs） via the coronary artery on the ventricle remodeling after AMI as well as the mechanisms of the effects of transplantation of different stem cells on ventricle remodeling. Methods A total of 36 male pigs were enrolled in this study, which were divided into 4 groups： control group, simple infarct model group, BM-MNC transplantation group, and MSCs transplantation group. At 90 minutes when a miniature porcine model with AMI was established, transplantation of autologous BM-MNC （（4.7±1.7）×10^7） and MSCs （（6.2±1.6）×10^5） was performed in the coronary artery via a catheter. Ultrasound, electron microscope, immunohistochemical examination and real time reverse transcriptase-pelymerase chain reaction were used respectively to observe cardiac functions, counts of blood vessels of cardiac muscle, cardiac muscle nuclear factor （NF）-κB, myocardial cell apoptosis, and the expression of the mRNA of vascular endothelial growth factor （VEGF） and basic fibroblast growth factor （bFGF） in cardiac muscles. Multivariate Logistic regression was used to analyze the correlation factors of left ventricular end-diastolic diameter （EDD）. Results The number of blood vessels in the infarct zone and around its border in the BM-MNC transplantation group was more than those in the infarct model group and MSCs group （P=0.0001） and there was less myocardial cell apoptosis in the stem cell transplantation group than that in the infarct model group （all P 〈0.01）. The positive rate of NF-κB in the stem cell transplantation group was lower than that in the infarct model group （P=0.001）. The gene expression of VEGF in the infarct border zone of the BM-MNC group was higher than that in the MSCs group （P=0.0001）. The gene expression of bFGF in the infarct border zone in the MSCs transplantation group was higher than that in the infarct model group and the BM-MNC group （P=-0.0001）. Left ventricular ejection fraction was inversely proportional to the apoptotic rate of myocardial cells and cardiac muscle NF-κB but positively correlated with the number of blood vessels and the expression of VEGF and bFGF in the infarct zone and infarct border zone. The Multivariate Logistic regression analysis on the factors influencing the left ventricular end-diastolic diameter after stem cell transplantation showed that the expression of VEGF mRNA in the cardiac muscles in the infarct zone, the number of apoptotic myocardial cells and the expression of NF-κB in the infarct border zone were independent factors for predicting the inhibitory effect on the dilation of left ventricular EDD after stem cell transplantation. Conclusions Transplantation of autologous BM-MNC and MSCs in pigs can improve the condition of left ventricular remodeling and recover the cardiac functions after AMI. The improvement of cardiac functions is related to the increase of blood vessels, the increased expression of VEGF and bFGF, the reduction of myocardial cell apoptosis, and the decrease of NF-κB level in cardiac muscle tissues after stem cell transplantation.展开更多
文摘Usually it is rather difficult to obtain and retain an ideal phenotype in an animal breeding program.Inspired by the success in transplanting cellular nuclei of amphibian and fish species since the 1950s,scientists began to believe in the possibility that cloned mammals could be acquired as well in the same way.Some 20 years prior to 1986 when the British biologist S.M.Willadson succeeded in breeding the first sheep by nuclear transplant,Prof.T.C.Tung (Tong Dizhou), a famous Chinese exper-
基金supported in part by a grant from the National Nature Science Foundation of China(30972923)
文摘BACKGROUND： Acute cellular rejection（ACR） after liver transplantation（LT） is one of the most common problems faced by transplant recipients in spite of advances in immunosuppressive therapy. Recently, clinical trials reported that ursodeoxycholic acid（UDCA） reduced the incidence of ACR significantly.However, others have shown contradictory conclusion. Therefore,we performed a meta-analysis of rigorous randomized controlled trials（RCTs） to determine the efficacy of UDCA in reducing ACR after LT.DATA SOURCES： All RCTs that evaluated efficacy of UDCA as an adjuvant treatment to prevent ACR after LT were searched from PubMed/MEDLINE, EMBASE, Cochrane Central Register of Controlled Trials, ScienceDirect databases and Web of Science（from January 1981 to March 2012）. There was no language limitation in these searches. Relevant abstracts of international meetings were also searched. References of each included study were searched manually.RESULTS： A total of 234 patients from four high-quality RCTs（Jadad score 4 to 5） were included in this meta-analysis.Prophylactic use of UDCA did not decrease the incidence of ACR（RR： 0.94, 95% CI： 0.77-1.16, P0.05）, steroid-resistant rejection（RR： 0.77, 95% CI： 0.47-1.27, P0.05） and the number of patients with the multiple episodes of ACR（RR： 0.60, 95% CI：0.28-1.30, P0.05）. Different intervention programs（high-dose vs low-dose UDCA; early vs delayed UDCA treatment） also did not alter the outcomes.CONCLUSIONS： UDCA, as an adjuvant treatment, was not ableto prevent ACR and steroid-resistant rejection after LT. Further trials should be done to determine whether higher dose of UDCA will be beneficial.
基金the 38th China Postdoctoral Science Foundation, No. 2005038055 the Scientific and Technological Research Projects of Liaoning Province, No. 2005225003-14the Natural Science Foundation of Liaoning Province, No. 20052204
文摘BACKGROUND： With developments of tissue engineering and genetic engineering, we aim to culture myoblasts, which are characterized by high purity, high quality and high production, for wide application in neural regeneration researches. OBJECTIVE： To modify traditional dissociation method in order to obtain myoblasts, which are characterized by high purity, high quality and high production, and explore the biological properties under in vitro culture. DESIGN： Observational study. SETTING： Basic Institute of Academy of Military Medical Sciences of Chinese PLA. MATERIALS： Four neonatal Wistar rats of 5 days old, both genders and mean body mass of 10 g were selected in this study. The main reagents and devices were detailed as follows： DMEM medium （Gibco Company）, fetus bovine serum （FBS, Hycolne Company）, collagenase Ⅱ （Sigma Company）, trypsin （Sigma Company）, dispase Ⅱ （Sigma Company）, desmin antibody （Fuzhou Maixin Company）, antibody Ⅱ and ABC kit （Wuhan Baster Biotechnology Company）, desk centrifuge （KUBATO, Japan）, and inverted phase contrast microscope （LEICA DMIRB, Germany）. METHODS： The experiment was carried out in the Basic Institute of Academy of Military Medical Sciences of Chinese PLA from June to October 2006. Neonatal rats were sacrificed under sterile condition to obtain skeletal muscles of limbs, which were washed with cold PBS （containing benzylpenicillin and estreptomicina）, and muscular tissue was sheared into pieces. Then, those muscular pieces were added with mixed digestive enzyme （containing 2 g/L collagenase Ⅱ ＋ 5 g/L dispase Ⅱ ＋ 0.28 g/L CaCl2） as twice volume as pieces, dealt with mechanical pipetting for 5 minutes and cultured in CO2 incubator for 10 minutes. The operation was done for three times and the muscular pieces were digested for 45 minutes in total. Moreover, cells were suspended again in order to obtain myoblasts from skeletal muscle of neonatal rats. In addition, myoblasts were purified with differential attachment technique and enzyme digestion so as to observe morphological characteristics and growth, draw growth curve, analyze surface structure under scanning electron microscope, and evaluate with Desmin immunohistochemical staining. MAIN OUTCOME MEASURES： Morphological characteristics and growth ofmyoblasts cultured in vitro. RESULTS： ①Growth of myoblasts of skeletal muscle： Primary cells had well growth, mature and differentiation. The positive rate of Desmin was 94% and purification of cells was ideal. Growth curve of cells demonstrated that myoblasts which were characterized by high purification started proliferation plentiful through transient growth lag phase （about at one or two days after inoculation）. If myoblasts were not dealt with any interventions, they might become sarcotubule gradually at 3 - 5 days after proliferative phase. During this period, myoblasts maintained a monocaryon-bipolarity state under inverted phase contrast microscope. Furthermore, the growth of cells was the strongest and reproductive activity was the most powerful. This suggested that myotube started to form; in addition, muscle fiber of contractility might form under a well culturing condition. ②Immunocytochemical stain with desmin antibody： Interzonal fiber of desmin from myoblasts showed strongly positive reaction. Positive staining existed in cytoplasm had a high nucleus-cytoplasm ratio. However, myoblasts showed negative or mildly positive reaction. CONCLUSION： It is ideal for modified multi-enzymatic digestion and double purification method to dissociate and purify myoblasts of skeletal muscle; meanwhile, these two methods are both the effective ways to provide convenient conditions to obtain seed cells for neural regeneration researches.
文摘Background The necrosis of a large number of myocardial cells after acute myocardial infarction （AMI） results in a decrease of cardiac function and ventricle remodeling. Stem cell transplantation could improve cardiac function after AMI, but the involving mechanisms have not been completely understood. The present study aimed to investigate the effects of transplantation of autologous bone marrow mononuclear cells （BM-MNC） and mesenchymal stem cells （MSCs） via the coronary artery on the ventricle remodeling after AMI as well as the mechanisms of the effects of transplantation of different stem cells on ventricle remodeling. Methods A total of 36 male pigs were enrolled in this study, which were divided into 4 groups： control group, simple infarct model group, BM-MNC transplantation group, and MSCs transplantation group. At 90 minutes when a miniature porcine model with AMI was established, transplantation of autologous BM-MNC （（4.7±1.7）×10^7） and MSCs （（6.2±1.6）×10^5） was performed in the coronary artery via a catheter. Ultrasound, electron microscope, immunohistochemical examination and real time reverse transcriptase-pelymerase chain reaction were used respectively to observe cardiac functions, counts of blood vessels of cardiac muscle, cardiac muscle nuclear factor （NF）-κB, myocardial cell apoptosis, and the expression of the mRNA of vascular endothelial growth factor （VEGF） and basic fibroblast growth factor （bFGF） in cardiac muscles. Multivariate Logistic regression was used to analyze the correlation factors of left ventricular end-diastolic diameter （EDD）. Results The number of blood vessels in the infarct zone and around its border in the BM-MNC transplantation group was more than those in the infarct model group and MSCs group （P=0.0001） and there was less myocardial cell apoptosis in the stem cell transplantation group than that in the infarct model group （all P 〈0.01）. The positive rate of NF-κB in the stem cell transplantation group was lower than that in the infarct model group （P=0.001）. The gene expression of VEGF in the infarct border zone of the BM-MNC group was higher than that in the MSCs group （P=0.0001）. The gene expression of bFGF in the infarct border zone in the MSCs transplantation group was higher than that in the infarct model group and the BM-MNC group （P=-0.0001）. Left ventricular ejection fraction was inversely proportional to the apoptotic rate of myocardial cells and cardiac muscle NF-κB but positively correlated with the number of blood vessels and the expression of VEGF and bFGF in the infarct zone and infarct border zone. The Multivariate Logistic regression analysis on the factors influencing the left ventricular end-diastolic diameter after stem cell transplantation showed that the expression of VEGF mRNA in the cardiac muscles in the infarct zone, the number of apoptotic myocardial cells and the expression of NF-κB in the infarct border zone were independent factors for predicting the inhibitory effect on the dilation of left ventricular EDD after stem cell transplantation. Conclusions Transplantation of autologous BM-MNC and MSCs in pigs can improve the condition of left ventricular remodeling and recover the cardiac functions after AMI. The improvement of cardiac functions is related to the increase of blood vessels, the increased expression of VEGF and bFGF, the reduction of myocardial cell apoptosis, and the decrease of NF-κB level in cardiac muscle tissues after stem cell transplantation.
