Objective To test the hypothesis that marrow stromal cells （MSCs）, when implanted into selfmyocardium in rabbits, can undergo milieu-dependent differentiation and express cardiomyogenic phenotypes and enhance cardiac function of ischemic hearts, through establish a clinically relevant model for autologous MSCs transplantation, Methods Thirteen New Zealand White rabbits were randomly divided into experimental group （n= 7） and control group （n= 6）. In experimental group, autotogous MSCs（3× 106 cells/30μl） labeled with Bromodeoxyuridine （BrdU） were respectively injected into superior, central and inferior sites in the periphery of the myocardial infarct region. Phosphate buffer saline （PBS） was injected into the scar of the control group hearts according to the same procedure used in the experimental group. Four weeks later, the transplanted labeled MSCs were detected by laser scanning confocal microscopy and the cardiac function were examined by echocardiogram and muhichannel physiologic recorder. Results After 4 weeks, transplanted MSCs were demonstrated myogenic differentiation with the expression of α-sarcomeric actin and connexin 43 located in intercalated disk. MSCs increased the number of vessels compared with controls in myocardial ischemia area. MSCs implantation resulted in markedly improved left ventricular contractility[left ventricular ejection fraction （LVEF）： 0. 51 ± 0.07 vs. 0. 43 ± 0.06 ,left ventricular lateral wall motion distance （LVLWMD） ：1. 75±0. 42mm vs. 1.09±0. 28mm, left ventricular systolic wall thickening ratio（LVAT） ：0. 19%±0.05% vs. 0. 11%±0.04%, left ventricular systolic pressure （LVSP）： 113. 1± 6.3mmHg vs. 99, 5 ± 5, lmmHg, left ventricular end diastolic pressure （LVEDP）： 11. 5±2. lmmHg vs, 14, 3 ±3. lmmHg, maximum rate of left ventricular pressure rise （＋dp/dtmax）：4 618. 3±365. 2 mmHg/s vs. 3 268. 1± 436.9 mmHg/s, maximum rate of left ventricular pressure fall （-dp/dtmax） ：3 008.8±346.7 mmHg/s vs. 2 536.9± 380.4 mmHg/s, P〈0.05]. Conclusion Transplanted autologous MSCs are able to undergo differentiation to form myocardial cells and improve the cardiac function of ischemia myocardium effectively. Autologous MSCs transplantation may have significant clinical potential in treatment myocardial ischemia.
ObjectiveTo study the cell apoptosis and the dynamic expression and significance of apoptosis-related genes in graft veins. MethodsA rat experimental model of autogenous graft vein was established by transplanting the right external jugular vein to infrarenal abdominal aorta in 100 Wistar rats. TUNEL and immunohistochemistry were used to detect the apoptosis, the expression of apoptosis-related genes bcl-2 and bax in vascular smooth muscle cells (VSMCs) of graft veins. ResultsWithin the 8 weeks after transplantation, the apoptotic VSMCs in the graft veins were much more than those in the control group with the apoptotic rate reaching the peak〔（28.5±16.6）%〕 on the 2nd week and dropping to （8.1±2.8）% during the 4th to 8th week. There was statistical difference compared to the control group 〔(0.5±0.2)%, P＜0.01〕. From 1 to 2 weeks, the positive rate of bcl-2 was (22.1±5.4)% which was higher than that of the control group and the 4-8 week group (P＜0.01); from 1 to 6 weeks, the expression of bax was higher than that of the control group 〔(5.5±2.3)%〕 and the postoperative 8th week group 〔(8.2±2.9)%, P＜0.01〕.Conclusionbcl-2 and bax protein may be involved in the regulation of apoptosis of VSMCs. Apoptosis of VSMCs may be an important factor in graft remodeling and graft vein stenosis.
Objective To evaluate the clinical effects of fibula flap grafts on the repair of the extremities with traumatic compound tissue defects. Methods In 12 cases, the fibula flap grafts were employed to restore the extremities with traumatic compound tissue defects. Of the 12 patients, 9 were males, 3 were females; their ages ranged from 12 to 45. There were 2 cases of tibia defect combined with fibula fracture, 2 cases of tibia defect, 2 cases of radius defect, 3 cases of ulna defect, 1 case of calcaneus defect,and 2 cases of firstmetatarsus defect. The bone defect length ranged from 4.2 to 10.6 cm, 7.8 cm in average.The skin defect area ranged from 10.0 cm×4.5 cm to 27.0 cm×15.0 cm. The free transplantation of fibular flaps were used in 9 cases, the lapse operation were used in 2 cases, retrograde shift were used in 1 case. Results Postoperational vein crisis and commonperoneal nerve traction injury were observed in category mentioned above respectively. All the 12 fibula flaps survived after proper treatments such as removalof great saphenous vein. Follow-ups were done for 6 to 24 months. Both the transferred fibula and the recipient broken end reflected bones were healed. Four patients underwent the second-phase reconstruction operation oftendon moving power. One wrist and 1 ankle underwent arthrodesis in 3 to 6 months.All the effects were satisfactory. Conclusion The fibula flap grafts provide arelatively better alternative to repair the extremities with long bone compoundtissue defects. In addition, the sensory function reconstruction of fibula flaps should be given full attention.
Abstract: Objective To evaluate if cardiac function and myocardial perfusion in acute ischemia myocardial transplanted by autologous bone mesenchymal stem cells (MSC) can be improved. Methods Sixteen New Zealand rabbits were studied.The left anterior descending coronary artery under the first diagonally branch was ligated to result in acute myocardial ischemia models,the sixteen models were divided into two groups with randomed number table. Control group(n=8): 0.6ml αminimum essential medium was injected into myocardium; transplanted group (n=8): 0.6ml medium of autologous MSC marked with 5-bromium,2-deoxy-uridine (BrdU) was injected into myocardium. Echocardiography were erformed to measure left ventricular ejection fraction(LVEF),as well as the displacement and strain of apex segment of left ventricle pre-ichemia,beforeand 4 weeks after treatment; the target myocardial tissues were harvested 4 weeks after treatment,double immunohistochemistry staining of anti-BrdU and anti-troponin T(TnT) were used to evaluate the survival and differentiation of implanted MSC; immunohistochemistry staining of anti-CD146 endothelium factor were used to evaluate the density of capillary vessels in treated myocardium. Results Double immunohistochemistry staining showed that positive cells were found in transplanted group and not found in control group. Anti-CD146 immunohistochemistry staining showed density of capillary vessels of transplanted group was significantly more than that of control group(Plt;0.05) ; LVEF,displacement and strain of cardiac apex of transplanted group improved significantly more than those of control group(Plt;0.05). Conclusion Transplanted to acute myocardium ischemia models of rabbits, MSC can differentiate into myocardium-like cells in myocardial microenvironment,and improve global and part cardiac systolic function and then improving perfusion of ischemia myocardium.
In the study of repair of massive bone defect with free vascularized fibula graft, 13 cases were reported, in which traumatic defect in 7 cases, segmental resection of bone from tumors in 5 cases and osteomylitis in 1 cases. They all were treated successfully with vascularized fibular graft. After a follow-up of 6 months to 7 year, bone healing was observed with satisfactory and rehabilitation of functions. In one case, fatigued fracture occured twice due to early walking. It was concluded that free vascularized fibular graft was very helpful in the repair of massive bone defect, but prolonged external fixation after operation might be important to prevent fractur of grafted bone.
Objective To study the feasibility of transplanting human saphanous vein endothelial cells to luminal surface of blood vessel prosthesis and to play a theoretical foundation for the clinical application of autologous endothelial cell transplantation. Methods Human saphanous vein endothelial cells were harvested with 0.1% collagenase and cultivated in vitro for 13.08±1.24 days. The cultures were confirmed as endothelial cells with the fourescent linked anti-Ⅷ antigen antibodies. The content of both 6-keto-PGF1α and Von Willebrand factor (vWF) in the supernatant were detected with ELISA and radioimmunoassay. The multiplied cells were lined in vitro onto the luminal surface of expanded polytetraflouroethylene (ePTFE) grafts precoated with fibrin glue and fibronectin, then cultivated again for 9 days. Results 11.46±2.69×１０６ of available endothelial cells could be regularly obtained, the number of endothelial cells increased 147.93±88.68 times when culture were terminated. All the cells diploid cells with a purity of 99%. The content of both 6-keto-PGF1α and vWF in the media showed no significant difference between the primary and subculture passages. The luminal surface of grafts was covered completely by a spindlelike endothelial monolayer and an even fibrin glue matrix could be seen underneath. Conclusion Endothelial cells derived from human saphanous veins might be feasible to be transplanted onto the luminal surface of ePTFE and present a potential clinical application.
PURPOSE:To investigate the approaches for transplanting retinal pigment epithelium.
METHODS,Retinal pigment epithelial eells(RPR)of pigmented rabbits' eyes prepared by rotalne preparation of our institute,were transphmted in 18 unpigmemed rabbits'eyes.Eight eyes were undergone outer approach, i.e., transplanting the RPR cells to the subretinal space of recipient eyes by way of perforating sclera and choroid;while 10 eyes were undergone internal approach by way of the routine procedure of vitrectomy with making artificial localized retinal delachment. Light and transmisskm electrone microscopy examination were done at 10th, goth, 40th and 90th day after the operation. RESULTS: In internal approach group,tbe operated eyes,revealed no difference in thickness of the neural retinal layer in transplanted and non-transplanted
area 40 days after operation tinder light microscope. Transmission electrone microscopy revealed postoperatively the transplanted RPE cells attached to the Brucb's membrane and the outer segments of photoreeeplive ceils located at a normal position at the 40th dayland the secondary lysozymes with engulfed outer segment were found in the Iransplamed cells at the 90th day. Tbe outer approached operations in eight eyes were failed owing to ehoroid hemorrhage or perforation of retina.
CONCLUSION:The internal appraach procedure is much effebtive and practical for transplantation of RPE cells.
（Chin J Ocul Fundus Dis,1997,13:160-162）
Objective To investigate the effects of ectomesenchymalstem cells on hematopoiesis after total body irradiation in rats. Methods The primary ectomesenchymal stem cells were isolated from E11.5 SD fetal mandibular processes by 25g/L trypsin and cultured with DMEM/F12. The morphology and growthrate were observed by inverted microscope. Eighty SD male rats randomly dividedinto ectomesenchymal stem cells group (n=20), fibroblast group(n=20), saline group(n=20) and control group(n=20), the first three groups were irradiated with 60Co γ rays at 6.0 Gy. The number of their bone marrow nucleated cells was counted after 4 weeks; the forming ability of colony-forming unit-granulocyte macrophage(CFU-GM) and histopathology of bone marrow were also observed. Results The cultured cells displayed monolayer growth and fibroblast-like with 2-4 processes. The ectomesenchymal stem cells could increase the number of bone marrow nucleated cells and peripheral blood white cell count, and improve the forming ability of CFU-GM. After 4 weeks of transplantation, the number of the peripheral blood white cells in group A was more than that in groups B and C(Plt;0.05), the contents of Hb in groups A and D was significantly higher than those in groups B and C(Plt;0.0). After 4 weeks, the bone morrow nucleated cells in group A were significant more than those in groups B and C(Plt;001); CFU-GM in groups A and D was higher than that in groups B and C(Plt;0.01). Conclusion Ectomesenchymal stem cells have characteristics of stem cells. It may improve hematopoiesis recovery of irradiated rats.
Objective To evaluate the biocompatibility of a new bone matrix material (NBM) composed of both organic and inorganic materials for bone tissue engineering. Methods Osteoblasts combined with NBM in vitro were cultured. The morphological characteristics was observed; cell proliferation, protein content and basic alkaline phosphatase(ALP) activity were measured. NBM combined with osteoblasts were implanted into the skeletal muscles of rabbits and the osteogenic potential of NBM was evaluated through contraat microscope, scanning electromicroscope and histological examination. In vitro osteoblasts could attach and proliferate well in the NBM, secreting lots of extracellular matrix; NBM did not cause the inhibition of proliferation and ALP activity of osteoblasts. While in vivo experiment of the NBM with osteoblasts showed that a large number of lymphacytes and phagocytes invading into the inner of the material in the rabbit skeletalmuscle were seen after 4 weeks of implantation and that no new bone formation was observed after 8 weeks. Conclusion This biocompat ibility difference between in vitro and in vivo may be due to the immunogenity of NBM which causes cellular immuno reaction so as to destroy the osteogenic environment. The immunoreaction between the host and the organic-inorganic composite materials in tissue engineering should be paid more attention to.
OBJECTIVE To explore the healing mechanism of full-thickness wound treating by the intermingled skin transplantation of large sheet allograft with autograft through studying the expression of laminin (LN). METHODS Thirty-six SD rats with 10% to 15% of total body surface area (TBSA) full-thickness were made. After 3 days, the devitalized tissue were excised and transplanted a large sheet of allograft from Wistar rats and islets of autografts were implanted 3 days later. On day 3, 5, 7, 14, 21 after allografting, the expression of LN in the grafts were detected by immunohistochemistry. RESULTS On the 7th day postallografting, LN, which played positive action of epidermal cell adhesion, still retained in the allodermis after the rejection of alloepidermis occurred. On the 14th day postallografting, there appeared scattered LN underneath the epidermal cells migrating from islets of autografts. On the 21st day postallografting, LN in the basement membrane of skin grafts had completely formed. CONCLUSION The intermingled transplantation of large sheet allograft with autograft may provide components of basement membrane for wound healing, which may help to improve the appearance and function of skin.
