Objective To explore the in vitrodifferentiation of the rat mesenchymal stem cells (MSCs ) into the skeletal muscle cells induced by the myoblast differentiation factor (MyoD) and 5-azacytidine. Methods The MSCs were taken from the rat bone marrow and the suspension of MSCs was made and cultured in the homeothermia incubator which contained 5% CO2at 37℃. The cells were observed under the inverted phase contrast microscope daily. The cells spreading all the bottom of the culture bottle were defined as onepassage. The differentiation of the 3rd passage of MSCs was induced by the combination of 5-azacytidine, MyoD, transforming growth factor β1, and the insulin like growth factor 1. Nine days after the induction, the induced MSCs were collected, which were analyzed with the MTT chromatometry, theflow cytometry, and the immunohistochemistry. Results The primarily cultured MSCs grew as a colony on the walls of the culture bottle; after the culture for 5-7 days, the cells were shaped like the fibroblasts, the big flat polygonal cells, the medium sized polygonal cells, and the small triangle cells; after the culture for 12 days, the cells were found to be fused, spreadingall over the bottle bottom, but MSCs were unchanged too much in shape. After the induction by 5-azacytidine, some of the cells died, and the cells grew slowly. However, after the culture for 7 days, the cells grew remarkably, the cell volume increased gradually in a form of ellipse, fusiform or irregularity. After theculture for 14 days, the proliferated fusiform cells began to increase in a great amount. After the culture for 18-22 days, the myotubes increased in number and volume, with the nucleus increased in number, and the newly formed myotubes and the fusiform myoblst grew parallelly and separately. The immunohistochemistry for MSCs revealed that CD44 was positive in reaction, with the cytoplasm ina form of brown granules. And the nucleus had an obvious border，and CD34 was negative. The induced MSCs were found to be positive for desmin and specific myoglobulin of the skeletal muscle. The flow cytometry showed that most of the MSCs and the induced MSCs were in the stages of G0/G1，accounting for 79.4％ and 62.9％，respectively; however, the cells in the stages of G2/S accounted for 20.6％ and 36.1％. The growth curve was drawn based on MTT，which showed that MSCs weregreater in the growth speed than the induced MSCs. The two kinds of cells did not reach the platform stage，having a tendency to continuously proliferate.ConclusionIn vitro,the rat MSCs can be differentiated into the skeletal muscle cells with an induction by MyoD and 5-azacytidine, with a positive reaction for the desmin and the myoglobulin of the skeletal muscle. After the induction, the proliferation stage of MSCs can be increased, with a higher degree of the differentiation into the skeletal muscle.
ObjectiveTo evaluate the effect of pre-infusion of allogeneic lymphoyctes treated with 5-FU on the rat liver graft. MethodsRat liver transplant models from Wistar to SD were established. Four groups were designed as following: control group: only liver transplantation without any other intervention; lymphocytes group: 1 ml of untreated lymphocytes (5×106/ml) from Wistar rats were preinfused into SD rats on day 7 and 4 separately before transplantation; lymphocytes with low concentration of 5-FU group: low concentration 5-FU (7.5 μg) treated lymphocytes were preinfused as above; lymphocytes with high concentration of 5-FU group: high concentration 5-FU (15 μg) treated lymphocytes were preinfused as above. Fas-L and CD8 expression were detected by immunohistochemistry method on day 7 after transplantation. ResultsThe integral opticaldensity (IOD) of Fas-L positive lymphocytes in the lobules of liver and portal areas were higher in lymphocytes with low concentration of 5-FU group than in the other groups （Plt;0.05）. There was no difference between lymphocyte group and lymphocytes with high concentration of 5-FU group （Pgt;0.05）. The IOD of CD8+ expression in lobules of liver was not different among all the three lymphocytes treated groups （Pgt;0.05）. But in portal areas, CD8+ expression was lower in the lymphocytes with low concentration of 5-FU group than in the other groups （Plt;0.05）. ConclusionPreinfusion of lymphocytes treated with low concentration 5-FU can induce graft immune tolerance, the probable mecanism of which is the increasing Fas-L expression in graft.
A acute partial obstructive hepatocholangitis model by selective ligation and injection of E coli into left hepatic bile duct was successfully founded in rat. Using parameters including mortality, mitochondrial glutamic oxalacetic transaminase and ornithine carbamoytransferase activity, pathological observation and blood culture of bacteria, we evaluated the model. The authors emphasize that this models is superior to the wole-bile-duct-challenged cholangitis model, which is characterized by liver injury.
Objective To investigate the quantity and distribution of motor fiber of rat’s C7 nerve root. Methods Motor fiber quantity and section area in the main nerves of the upper extremity and the fascicles of C7 in 30 SD rats were analyzed.Results Fascicles and certain amount (207) of motor fibers from the anterior division of C7 were distributed to musculocutaneous nerve and median nerve, the orientation of these fibers were not clear. The ones (323) from posterior division were to the axillary, radial, and dorsal thoracic nerves, thus the orientation of these fascicles was relatively definite. Conclusion Thedistribution of the motor fibers and fascicles in the divisions of C7 in rat is similar to human beings, so rat is a relatively good model for the study of selective C7 nerve root transfer.
Objective To study the effect of low-dose cyclophosphamide (CY) on apoptosis of lung parenchyma cells in the early severe burn stage in rats. Methods Ninety clean SD male rats were randomly divided into 3 groups: the normal group (n=10), the experimental group (n=40) and the burn group (n=40). The model of degree III with 30% burn area was made in the experimental group and the burn group. CY (2 mg/kg) was injected into the abdominal cavity right after burn in the experimental group. No treatment was done in the normal group and burn group. Lung tissues were obtained at 3, 6, 12and 24 hours, respectively, after burn, and were observed by HE staining. Apoptosis of lung parenchyma cells was observed by TUNEL. Results Lung tissues were observed under the opticalmicroscopy in the normal group: the pulmonary structure was clear, and there were no inflammatory cells and exudation in the alveolar space and bronchial lumen. Besides, a few RBCs were seen. Pathological changes of lung tissues were observed under the opticalmicroscopy in the burn group: alveolar septum was obviously widened; alveolar wall was destroyed; interstitial edema and atelectasis occurred; and pathological lesion was gradually aggravated as time passed by. The pathological lesion of lung tissues mentioned above in the experimental group was better than those in the burn group. Compared with the normal group, the apoptosis ratio of lung parenchyma cells continuously increased in the burn group from the 3 hour after burn, and reached the peak at 12 hours. There were significant differences between the two groups (P lt; 0.05). However, in the experimental group, the apoptosis ratio of lung parenchyma cells increased at 3 hours after burn, cut down to normal at 6 and 12 hours, respectively, and notably decreased at 24 hours. There were significant differences between the experimental group and the normal group (P lt; 0.05). Compared with the burn group, the apoptosisrate of lung parenchyma cells in the experimental group began to decrease strikingly from the 6 hours after burn, and there were significant differences between the two groups (P lt; 0.05). Conclusion Low-dose CY can restrain the apoptosis of lung parenchyma cells in the early severe burn stage in rats and alleviate the injury of the lung.
Liver transplantation in hamster-to-rat is a good model for the research in xenograft, but it is difficult to be performed. In order to simplify its procedures, 38 cases of liver transplantation in hamster-to-rat were performed with several technique improvments on the basis of orthotopic liver transplantation in rats. In the light of hamster’s anatomy, three cuffs anastomosis method was used. Because of its high stability and high survival rate, the model can be used widely as the research for liver xenograft.
OBJECTIVE: To investigate a animal model of spinal cord injury in different degrees of impact. METHODS: A new weight-drop device was designed with the character of controlled degree of impact and time. After thirty-five rats underwent different degrees of impact, their motor function and pathological changes were observed. RESULTS: In control group, the rats could walk after reviving, and the micro-structure of spinal cord was normal. With 0.5 mm depth of impact, the rats also could walk, and the micro-structure of spinal cord did not change obviously. With 0.8 mm depth of impact, the rats could walk after several days of injury and only slight damage could be found in spinal cord. When the impact depth increased to 1.0 or 1.5 mm, the rats were paralyzed completely and could not walk after four weeks of injury. Severe injury was observed in spinal cord. CONCLUSION: This animal model of spinal cord injury is based on different degrees of impact. It has stable and repetitive characters for the research on spinal cord injury.
【摘要】 目的 用老年大鼠建立一種與人類發病年齡相符的穩定可重復的急性心肌梗死動物模型。 方法 Wistar大鼠經戊巴比妥鈉麻醉后，氣管切開插管，連通呼吸機，開胸后結扎左冠狀動脈前降支。分不同時間點測血流動力學后取出心臟行病理組織學觀察。 結果 成功建立老年大鼠心肌梗死模型并進行病理組織學驗證。 結論 建立老年大鼠心梗模型是可靠的，有助于指導臨床實踐。【Abstract】 Objective To develop a steady and repeatable acute myocardial infarction model in old rats in accordance to human age. Methods Wistar rats were anaesthetized with sodium phenobarbital. After tracheotomy intubation, respiration machine was linked. Left anterior thoracotomy was performed to ligate the left anterior descending coronary artery. After hemodynamic parameters were obtained at various time points, hearts of the rats were taken for pathologic and histological examinations. Results The model of acute myocardial infarction in old rats was established successfully and proved by pathologic and histological staining. Conclusion It is reliable to establish the myocardial infarction model in old rats, which is helpful in guiding clinical practices.
Objective To explore the effect of age and gene therapyon the differentiation of marrow mesenchymal stem cells （MSCs） of the rats. Methods MSCs from the young (1-month-old), adult (9-month-old), and the aged(24monthold) rats were expanded in culture and infected with adenovirus mediated human bone morphogenetic protein 2 gene (Ad-BMP-2). The expression of BMP-2 and osteoblastic markers such as alkaline phosphatase(ALP), collagen Ⅰ(Col Ⅰ), bone sialoprotein(BSP) and osteopontin(OPN) were assayed during the process of differentiation. Their abilities to induce ectopic bone formation in nude mice were also tested. Results There was no significant difference in the expression of BMP-2 among the 3 groups. ALP activity assay and semi-quantitative reverse transcription polymerase chain reaction(RT-PCR) demonstrated that there were no significant differences in the expression of osteoblastic markers ALP, Col-Ⅰ, OPN and BSP amongthe 3 groups. Histomorphometric analysis indicated that there were no significant differences in the volume of the newly formed ectopic bones in nude mice amongthe 3 groups. Conclusion MSCs obtained from the aged ratscan restore their osteogenic activity following human BMP-2 gene transduction, therefore provides an alternative to treating the aged bone disease.
