Objective To supply references to tissue-engineered skin cl inical appl ications with autogenic BMSCs composited collagen membrane to repair swine full-thickness cutaneous deficiency. Methods Twenty mL bone marrow were obtained respectively from 4 swine, autogenic BMSCs were cultured and passed to the 3rd passage. The fresh bovine tendontreated by means of chemically cross-l inked was made 5 cm diameter collagen I (Col I) membrane. The 2 × 107/mL P3 swine autogenic BMSCs labeled DAPI were planted to sterile Col I membrane for 24 hours incubation, then the tissue-engineered skin was constructed. The five full-thickness skin defect of 5 cm diameter was excised to the muscle from forward to backward on the back midl ine two sides of swine. The tissue-engineered skin were implanted in the experimental group, while Col I membrane was implanted in control group. After 3 and 8 weeks of implantation, the two swine wound surface heal ing circumstance was observed and further evaluated with histology analysis and TEM. After 3 weeks of implantation, the experimental group were observed with fluorescence microscopy and staining for glycogen. Results After 3 weeks of implantation, the wound surface of control group were observed nigrescence, scab and putrescence, and after 8 weeks of implantation, also evident putrescence and scar. The wound surface of experiment group was al ive after 3 weeks implantation, appearance was leveled off and flexible without evident scar. The wound surface recovered well after 8 weeks of implantation, wound surface heal ing rate was significantly difference between the two groups (P lt; 0.01). After 3 weeks of implantation, control group were observed acestoma hyperplasia and no epidermal coverage by histology analysis. The experimental group was showed integrity epidermis and dermis structure. The basal layer was crimson and continuously positive with glycogen staining. After 8 weeks of implantation, the experimental group and control group were emerged normal skin structure. After 3 weeks of implantation in control group, a lot of neutrophil ic granulocytes and fibroblasts were noticed, but no epidermal structure was observed under TEM. In the experimental group, a lot of epidermal cells were observed, dermatome connection among epidermal cells and hemidermosome connection between basilar membrane cells and basal membrane were observed in epidermis. In the dermis experimental group, blood capillary endothel ial cells were noticed. Furthermore, considerable collagen fiber deposit was found in the surrounding tissue of fibroblasts. After 3 weeks of implantation, BMSCs labeled with DAPI were located reconstructed epidermal basement membrane and dermis by fluorescence microscopy. Conclusion Tissue-engineered skin which is composited with autogenic BMSCs as seed cells and collagen membrane were potential prospects in appl ication of repairing swine full-thickness cutaneous deficiency.
To study the method of isolating and culturing synovium-derived MSCs (SMSCs), and to investigate its multiple differentiation potential in vitro. Methods Three 2-month-old Changfeng hybrid swines weighing 8-10 kg (male and female) were used. SMSCs were harvested from the synovium of swine knee joints and cultured in vitro. When the SMSCs at passage 3 reached confluence, basic culture medium was removed, and the multi ple differentiationpotential of SMSCs was demonstrated in specific induction media (experimental group). The cells at passage 3 cultured with basic culture medium served as control group. After 21 days of chondrogenic differentiation, the cells underwent toluidine blue staining, immunohistochemistry staining and real-time fluorescence quantitative PCR detection. After 10 and 21 days of osteogenic differentiation, the cells underwent ALP staining and Al izarin red staining, respectively. After 21 days of adipogenic differentiation, the cells underwent Oil red O staining. Results SMSCs displayed long and thin or polygonal morphology 24 hours after culture. They prol iferated fast 48 hours after culture and presented large number of spindle-shaped cells with few globular cells 72 hours after culture. For the experimental group 21 days after chondrogenic induction, the cells were positive for toluidine blue staining with the formation of Aggrecan outside the cells; the immunohistochemistry staining revealed the expression of Col II; the real-time fluorescence quantitative PCR detection showed that the expressions of Col II A1, Aggrecan and SOX9 mRNA of the experimental group were greater than that of control group (P lt; 0.05). The cells were positive for ALP staining 10 days after osteogenic induction, and positive for Al izarin red staining 21 days after osteogenic induction, with the formation of calcium nodules. Oil red O staining displayed the formation of l i pid droplets inside the cells 21 days after adi pogenic induction. For the control group, the results of all the staining assays were negative except the ALP staining presenting with sl ight positive result. Conclusion SMSCs can be isolated from knee joint of swine and proliferate and differentiate into osteogenic, adi pogenic and chondrogenic cells in vitro. SMSCs may be a promising source of seed cells for tissue engineering.
OBJECTIVE: To evaluate the histological characteristics of xenogeneic graft-versus-host disease (GVHD), and to differentiate it from the allogeneic GVHD. METHODS: The original articles related to the characteristics of xenogeneic GVHD and the relationship between xenogeneic GVHD and allogeneic GVHD were widely reviewed in the past decade. RESULTS: Xenogeneic GVHD was different from allogeneic GVHD in the speed of development, position and features of pathological changes, due to the incompatibility of common simulative signals, adhesive molecules and difference of antigens. Xenogeneic spleen transplantation could induce xenogeneic GVHD. CONCLUSION: Xenogeneic GVHD can be used as a model to mimic rejection in xenotransplantation and to evaluate the compatibility of xenotransplantation.
ObjectiveTo explore the histopathological changes of the pigs′ eyes in vivo after radial optic neurotomy (RON), and provide the experimental foundation for the safety of RON.MethodsA total of 12 healthy miniature pigs were used in the experiment, in whom 8 were executed at the 1st, 3rd , 7th , and 48th day respectively after underwent RON in both eyes, and 4 were executed at the 120th day after underwent RON unilaterally (the other eye was as the control in 2 and underwent single vitrectomy in 2). All the enucleated eyes were cut in sections routinely and embeded in paraffin. The sections were stained by HE, Masson trichrome staining or Luxol fast blue staining and the different sections of optic nerve were observed by light microscope.ResultsNo damages of the major vessel wall were found and the cerebral pia mater of orbital optic nerves kept integrated. At the 1st day after the operation, the incisions came into being and local hemorrhages infiltrated into the circumambience and backside. The vacuole-like change induced by the demyelination of optic nerve fiber located at the incisions. At the 3rdday, the vacuolelike changes widened. At the 7thday, the fibroblasts aggregated at the incision, with hyperplastic neuroglia cells and dispersed pigmented granules. Lymphocytes and monocytes were the major infiltrated inflammatory cells. At the 48th day, collagen filled in the incisions and aggregated neuroglia cells of the rear optic nerves behind the incision were found, which showed weak staining with obvious boundary which was somewhat beyond the midline of optic nerves. At the 120th day, localized atrophy of optic nerve occurred under the incision. No abnormal pathological findings were found in the normal eyes and the eyes underogo vitrectomy.ConclusionsLocalized atrophy of optic nerves comes into being after the normal pig eyes in vivo underwent RON. The surgery is safe to some extent.(Chin J Ocul Fundus Dis, 2005,21:13-15)
【Abstract】ObjectiveTo compare the hemodynamic changes during operation of portal venous intubation or splenic venous intubation in extracorporeal venous bypass of swine orthotopic liver transplantation.MethodsThirty couples of healthy Duloke pigs were selected to perform orthotopic liver transplantation. According to the difference of cannula vessel of portal venous system during extracorporeal venous bypass, these pigs were divided into two groups: portal venous intubation group (n=15) and splenic venous intubation group (n=15). Hemodynamic changes were monitored continuously.ResultsTwo recipients died in portal venous intubation group, one died of unsmooth bypass in the operation, the other died of DIC. In splenic venous intubation group there was only one recipient death, who died of hemorrhagic shock. The time of anhepatic phase of splenic venous intubation group was (44.5±7.6) min, it was significantly shorter than portal venous intubation group(51.5±8.7) min(P＜0.05). Hemodynamic changes in phase Ⅲ and phase Ⅳ of portal venous intubation group were significantly different with that of splenic venous intubation group(P＜0.05). ConclusionApplication of bypass through splenic venous intubation during extracorporeal venous bypass of swine orthotopic liver transplantation can shorten the time of anhepatic phase, keep the hemodymamics relative stable in operation, and reduce the occurrence of postoperative correlative complication. It is an effective venovenous bypass pathway.