ABSTRACT
BACKGROUND: How dose sodium nitroprusside, as a vasodilatator, affect the potential of bone marrow-derived mesenchymal stem cells (MSCs) in hematopoietic differentiation?OBJECTIVE: To observe the changes during the differentiation of MSCs into hematopoietic cells after adding sodium nitroprusside, and compare the results with those of simple MSCs transplantation.DESIGN: A randomized grouping, controlled observation.SETTINGS: Department of Microbiology and Immunology; Department of Pathophysiology, Guangdong Medical College.MATERIALS: The experiments were carried out in Guangzhou Medical College in August 2006. Twenty-seven clean-degree balb/c mice of 7-8 weeks, were used as recipients, and were randomly divided into MSCs transplantation group (n =9), sodium nitroprusside+MSCs transplantation group (n =9) and blank control group (n =9). Another 4-week-old SD rat was selected as the MSCs donor. Sodium nitroprusside for injection (50 mg/piece) was provided by Beijing Double-Crane Modern Pharmaceutical Technology Co., Ltd (National drug approval: No. H11020907).METHODS: ① Under aseptic condition, the femur of SD rat was collected. MSCs in it were isolated for culture and amplifying in vitro. MSCs of passages 6-7 were digested and centrifugated, and the density was adjusted to 1×109 L-1.Monoclonal antibody with fluorescence labeled was added into cell suspension, and the phenotype was detected with flow cytometry. ② Sodium nitroprusside (50 mg/piece) was adjusted to the terminal concentration of 200 mg/L by adding with saline. It should be used within 4 hours. ③ Before transplantation, all the mice were exposed to 5.0 Gy. X-ray for 4 hours, and the absorbed dose was 1.45 Gy per minute. After irradiation, mice in the MSCs transplantation group were directly infused via caudal vein with 0.3 mL MSCs suspension (containing 1.5×106 cells); The mice in the sodium nitroprusside+MSCstransplantation group were firstly injected with the dispensed 200 mg/L sodium nitroprusside (0.15 mL), and immediately infused with 0.3 mL MSCs suspension (containing 1.5×106 cells) after 1 minute; The mice in the blank control group were infused with isovolume serum-free culture medium. ④ At 60 days after transplantation,peripheral blood was drawn from orbits of the survived mice in each group, single cell suspensions of bone marrow and spleen were prepared after the mice were killed, the levels of rat-derived hematopoietic cells CD11a and CD45 were detected with flow cytometry.MAIN OUTCOME MEASURES: ① MSCs culture and amplification; ② Levels of rat-derived hematopoietic cells in different tissues.RESULTS: All the 27 balb/c mice as recipients survived to the end of the experiment. ① MSCs isolation and amplification: The MSCs attached with uniform shapes of spindle and proliferated rapidly after culture for 3 days, and 90% of the cells were fused without overlapping at 6 days. The cells attached completely within 24 hours after passage,extended and became spindle again, rapidly proliferated and grew, and became fused completely at 3 days. ② Levels of rat-derived hematopoietic cells in different tissues: In the MSCs transplantation group and sodium nitroprusside+MSCs transplantation group, Iow expressions of rat-derived CD11a and CD45 hematopoietic cells could be detected in peripheral blood, bone marrow and spleen, and they were obviously higher in the sodium nitroprusside+MSCs transplantation group than in the MSCs transplantation group (t=2.619, P ＜ 0.05), while negative ones in the blank control group.CONCLUSION: MSCs have the ability to differentiate into hematopoietic stem cells, which can be promoted by sodium nitroprusside.
ABSTRACT
BACKGROUND: By targeting inducing differentiation in vitro,mesenchymal.stem cells(MSCs) transform into osteoblasts,lipocytes,chondrocytes,muscular cells,neuronal cells,etc. Whether Chinese herbs act on induced differentiation of MSCs in rats or not?OBJECTIVE: To study the amplification of MSCs cultured in vitro in SD rats and efficacy of Chinese herbs on targeting inducing differentiation of neuron-like cells.DESIGN: Exploring study with repeated observation and measurement based on cells.SETTING: Department of pathophysiology in a medical college.MATERIALS: Experimental marrow collected from SD male tested-healthy rats.METHODS: By adhesion method,MSCs in rats were isolated for amplifying culture in vitro. Flow-type cell instrument was applied for the determination of its surface antigen expression. Various Chinese herbal components were used for the targeting inducing differentiation of MSCs into neuron-like cells. The cellular morphology was observed under optical microscope. and the specific antigen label of neuronal cells was determined with immunocyto-chemical method.MAIN OUTCOME MEASURES:①Results of MSCs isolation and amplification;②Results of identification of MSCs surface antigen and neuon-like cells.RESULTS: By adhesion method,MSCs in rats were isolated successfully and amplified in a large amount in vitro. It was indicated in the results determined by flow-type cell instrument that CD14,CD1 1α,CD34,CD38,CD45,CD80 and CD86 presented negative,and CD29,CD44,CD90,CD105 and CD166 presented positive. By induced with various kinds of Chinese herbs,like huangqi(Radix Astragali seu Hedysari),tianma(Rhizoma Gastrodiae),renshen (Radix Ginseng),danggui(Radix Angelicae Sinensis),naoxinshu,renshen fengwangjian for 1 to 3 hours,most MSCs transformed into neuron-like cells,presenting soma and neurite. With immunocyto-chemical staining,neuron-specific enolase(NSE) and nestin displayed positive and glial fibrillary acidic protein negative.CONCLUSION: MSCs in SD rats have the potential of multi-targeting differentiation,presenting a strong capacity of amplification and self-replacement. In a suitable inducing condition,MSCs may differentiate into neuron-like cells.
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AIM: To examine the effect of external counterpulsation(ECP) on renin-angiotensin system(RAS) and hemodynamics in dogs with myocardial ischemia and their relationship. METHODS: Acute myocardial ischemia was induced by occluding the left anterior descending of the dogs. The local renin activity, angiotensin Ⅱ(AngⅡ) level, angiotension-converting enzyme(ACE) activity were tested by biochemical methods and hemodynamics was recorded by a 8-channel physiological recorder. RESULTS: Ischemia could actiivate renin,ACE and AngⅡ in cardiovasculature and ECP reduced them except for renin in ischemic myocardium. Ischemia also could activate RAS in lung and kidney, which play an important role on circulating RAS, and ECP reduced them. Furthermore, ECP could improve hemodynamics and there existed a close relationship between local AngⅡlevel and hemodynamics. CONCLUSION: ECP can reduce local RAS and improve hemodynamics in dogs with myocardial ischemia, which might be one of mechanisms underlying the protective effect of ECP on ischemic myocardium.
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AIM: To examine the effect of bacillus calmette-guerin (BCG) on experimental asthma in guinea pigs. METHODS: Guinea pigs were sensitized with BCG and then with ovalbumin (ip). Two weeks later, guinea pigs were challenged with ovalbumin (OVA) aerosol inhalation. Thirty one Guinea pigs were divided into three groups at random control group,OVA-treated group, BCG and OVA-treated group.RESULTS: Ovalbumin inhalation caused a marked airway infiltration of eosinophils and all the animals exhibit asthmatic symptoms. Pretreatment with BCG induced typical increase in lymphocytes and monocytes in peripheral blood and in bronchoalveolar lavage fluid (BALF). BCG markedly inhibited eosinophil infiltration and attenuated the asthmatic symptoms. CONCLUSION: These data suggest that BCG exerts an inhibitory effect on asthmatic inflammation.