Objective Bone marrow mesenchymal stem cells (BMSCs) play an important role in repairing nerve injury, meanwhile external temperature has significant effect on BMSCs transplantation, prol iferation, and differentiation. To investigate the effect of BMSCs transplantation and mild hypothermia on repair of rat spinal cord injury (SCI). Methods Forty-five female adult SD rats (weighing 200-250 g) were made the models of hemitransection SCI and divided randomly into 3 groups according to different treatments: group A (SCI group), group B (BMSCs transplantation group), and group C [BMSCs transplantation combined with mild hypothermia (33-35 ) group]. At 1, 2, 4, 6, and 8 weeks after injury, the fuction of hind l imb was evaluated with Basso Beattie and Bresnahan (BBB) score and incl ined plane test. At 4 weeks after injury, histopathology and BrdU immunohistochemistry staining were performed. At 8 weeks after injury, horseradishperoxidase (HRP) retrograde nerve trace and transmission electron microscope (TEM) testing were performed to observe the regeneration of axon. Results After 4 weeks, the function of hind l imb obviously recovered in groups B and C, there were significant differences in BBB score between groups B, C and group A (P lt; 0.05), between group B and group C (P lt; 0.05). There was no significant difference (P gt; 0.05) in tilt angle among 3 groups after 1 and 2 weeks, and there were significant differences (P lt; 0.05) among 3 groups after 4 weeks. HE staining showed that significant cavity could be seen in group A, l ittle in group B, and no cavity in group C. BrdU immunohistochemistry staining showed that the number of positive cells was 0, 90.54 ± 6.23, and 121.22 ± 7.54 in groups A, B, and C, respectively; showing significant differences (P lt; 0.01) among 3 groups. HRP retrograde neural tracing observation showed that the number of HRP positive nerve fibers was 10.35 ± 1.72, 43.25 ± 2.65, and 84.37 ± 4.59 in groups A, B, and C, respectively, showing significant differences (P lt; 0.01) among 3 groups. TEM observation showed that a great amount of unmyel inated nerve fibers and myel inated nerve fibers were found in central transverse plane in group C. Conclusion The BMSCs transplantation play an impontant role in promotion of recovering the function of hind l imb after SCI, and mild hypothermia has synergism effects.
Objective To investigate the feasibility of imaging of bone marrow mesenchymal stem cells (BMMSCs) labeled with superparamagnetic iron oxide(SPIO) transplanted into coronary artery in vivo using magnetic resonance imaging (MRI), and the redistribution of the cells into other organs. Methods BMMSCs were isolated, cultured from bone marrow of Chinese mini swine, and double labeled with SPIO and CMDiI(Cell TrackerTM C-7001). The labeled cells were injected into left anterior descending coronary artery through a catheter. The injected cells were detected by using MRI at 1 week,3weeks after transplantation. And different organs were harvested and evaluated the redistribution of transplanted cells through pathology. Results The SPIO labeled BMMSCs injected into coronary artery could be detected through MRI and confirmed by pathology and maintained more than 3 weeks. The SPIO labeled cells could be clearly imaged as signal void lesions in the related artery. The pathology showed that the injected cells could be distributed into the area of related artery, and the cells injected into coronary artery could be found in the lung, spleen, kidney, but scarcely in the liver, the structures of these organs remained normal. Conclusion The SPIO labeled BMMSCs injected into coronary artery can be detected by using MRI, the transplanted cells can be redistributed into the non-targeted organs.
Objective To explore the middle-term outcome of autologous bone marrow mononuclear cells transplantation in the treatment of lower l imb ischemia. Methods From March 2003 to June 2005, 65 patients with lower l imb ischemia were treated by autologous bone marrow mononuclear cells transplantation. Of the patients, there were 50 males and 15 females, with a mean age of 66.5 years (range 36-89 years), including 4 cases of simple arteriosclerotic occlusion,5 cases of thromboangiitis obl iterans and 56 cases diabetic lower l imb ischemia. A total of 400 mL bone-marrow blood were extracted from the posterior superior il iac crest. And then the mononuclear cells were isolated from the bone-marrow blood in the laboratory. The amount of transplantation bone marrow mononuclear cells was (0.60-1.80) × 109 (mean 1.05 × 109). Twelve patients received cell transplantation from two to four times and the other patients one time. According to the improvement of cl inical finding, the outcome was evaluated. Results All the patients were followed up for 8-56 months (mean 21.5 months). There were 8 deaths, and the mortal ity was 12.3%; 5 were due to myocardial infarction and heart failure and 3 were due to cerebral infarction. The general effective rate was 70.8% (46/65) and the recurrent rate was 10.7% (7/65). Of them, the response to treatment lasted over 12 months in 42 cases, accounting for 91.3% (42/46); over 24 months in 24 cases, accounting for 52.2% (24/46); and over 37 months in 12 cases, accounting for 26.1% (12/46). The effective rates were 100% in 12 patients who received 2-4 times transplantation and 64.2% in 53 patients who received 1 time transplantation, showing statistically significant difference between them (P lt; 0.001). Conclusion The middle-term outcome of autologous bone marrow mononuclear cells transplantation show that it is a feasible and simple method for treatment of lower l imb ischemia.
Objective To review the status and appl ication prospect in repair of spinal cord injury by stem cells. Methods The related articles in recent years were extensively reviewed, the biological characteristics of stem cells, the experimental and cl inical studies on repair of spinal cord injury by stem cells, the mechanism of the therapy and the problem were discussed and analyzed. Results The foundational and cl inical study indicated that the great advance was made in repair of spinal cord injury, the stem cells could immigrate in the spinal cord, and differentiate into neuron and secrete neurotrophic factors. So it could promote the repair effects. Conclusion Repair of spinal cord injury by stem cells is an effective therapystrategy, but many problems remain to be resolved.
Based on the pathogenic mechanisms of age-related macular degeneration (AMD), tremendous preclinical and clinical trials have demonstrated that cell transplantation which aim to replace impaired retinal pigment epithelium (RPE) with healthy RPE cells is a promising approach to treat AMD. So far, choices of cell sources mainly are autologous RPE, iris pigment epithelium, fetal RPE, human embryonic stem cell-derived RPE and human induced pluripotent stem cell-derived RPE, and some of them are undergoing clinical researches. Grafting manners in cell-based therapies are various including RPE sheet or RPE-choroid complex transplantation, RPE cell suspension injection, and RPE sheet transplantation with scaffolds. This review is limited to cell-based therapies for RPE that damaged first in the progress of AMD and focus on recent advances in cell sources, transplantation methods, preclinical and clinical trials, and the obstacles that must be overcome.
ObjectiveTo observe the survival, migration, and effect of human amniotic epithelial cells (hAECs) on hepatic fibrosis in immune rats so as to provide the experimental theory for the clinical treatment with hAECs.
MethodsSixty-four 10-week-old male Sprague Dawley rats (weighing, 220-280 g) were randomly divided into 4 groups, sixteen rats in each group. Rat hepatic fibrosis model was induced in groups A, B, and C; hepatic fibrosis rats were injected with 4×106 hAECs in group A, and with normal saline in group B, and no treatment was given in group C; group D served as control group. After 2 weeks of transplantation, the expression of human Alu gene repeat sequence was detected by DNA-PCR method and human leucocyte antigen G (HLA-G) by immunohistochemical staining in heart, liver, spleen, kidney, lung, and brain in group A, and then the percentage of positive expression was compared between organs except spleen. Semi-quantitative analysis was done for liver fibrosis with HE staining according to Chevallier semi-quantitative histological liver fibrosis scoring system, and immunohistochemical staining for TGF-β1 was used to record immunohistochemical score (ISH), the concentrations of aspartate transaminase (AST), alanine aminotransferase (ALT), and albumin (ALB) were determined to analyze hepatic fibrosis.
ResultsAlu gene repeat sequence and HLA-G could be detected in liver, heart, brain, lung, and kidney in group A, the percentage of positive expression in the liver was significantly higher than that in the other organs (P<0.05). The histological semi-quantitative score of group A (10.47±3.20) was significantly lower than that of groups B and C[(13.84±3.46) and (13.85±3.16)](P<0.05), but no significant difference was found between groups B and C (P>0.05). The ISH scores in groups A, B, C, and D were 3.60±1.50, 5.38±2.60, 5.50±2.40, and 1.87±1.36, respectively; groups A, B, and C were significantly higher than group D, and group A was significantly lower than groups B and C (P<0.05), but there was no significant difference between groups B and C (P>0.05). The concentrations of ALT and AST in groups A, B, and C were significantly higher than those in group D, and group A was significantly lower than groups B and C (P<0.05), but there was no significant difference between groups B and C (P>0.05). The concentration of ALB in groups A, B, and C was significantly lower than that in group D, and group A was significantly higher than groups B and C (P<0.05), but there was no significant difference between groups B and C (P>0.05).
ConclusionhAECs can survive in immune rats by intrasplenic transplantation and migrate to liver, heart, brain, lung, and kidney, and the liver shows the largest migration. The transplantation of hAECs in immune rat with cirrhosis can alleviate hepatic fibrosis and improve the serum indexes of liver function.
Purpose
To investigate the characteristics of intraocular growth of mice embryonic stem cells (ESC) in nude mice.
Methods
The undifferentiated murine ESC in vitro were transplanted into the eyes of nude mice.Mophological and immunohistochemical examinations were implemented.
Results
Two to three days after transplantation,yellowish-white granules and masses were seen inside the anterior chamber and vitreous cavity and enlarged gradually.Morphological examination showed that there were undifferentiated cells and differentiated cells in anterior chamber and vitreous cavity.The morphology and alignment of some differentiated cells were similar to those of the retina of nude mice.The cells were highly positive in NSE staining.
Conclusion
The transplanted ESC could grow in the eyes of nude mice and differentiate into neurons and retina-like structure.
(Chin J Ocul Fundus Dis，2000，16：213-284)
Objective To investigate the synergistic effect of a combination of grafted olfactory ensheathing cells (OECs) from the olfactory bulbs and intrathecal injection of vascular endothel ial growth factor (VEGF) on repairing spinal cord injury, and to explore the neuroprotection on both neurons and nerve fibers. Methods OECs from neonatal rats were cultured, purified, and collected with 0.25% trypsin after 9 days. A total of 75 adult female Wistar rats (weighing 200-250 g) were randomly divided into 5 groups: group A was sham-surgery group receiving laminectomy; the spinal cord injury model was establ ished with weight-dropped apparatus in the rats of groups B, C, D, and E. Then group B was injected with 10 μL DMEM-F12 medium without serum at injury site on the 1 day and was intrathecally administrated with 10 μL sal ine solutiontwice a day during the following 1 week; group C was injected with 10 μL DMEM-F12 medium and 25 ng recombined ratVEGF165 (rrVEGF165); group D was injected with 10 μL DMEM-F12 medium containing 1 × 105 OECs and 10 μL sal ine solution; group E was injected with 10 μL DMEM-F12 medium containing 1 × 105 OECs and 25 ng rrVEGF165. The functional recovery of hindl imb was evaluated by the Basso-Beattie-Bresnahan (BBB) score at 1 day and each week from 1 to 8 weeks. The histological changes and the changes of ultrastructure were observed at 8 weeks after operation by HE and electron microscope, and the immunohistochemistry staining was used for p75 nerve growth factor receptor (p75NGFR), Caspase-3, and von Willebrand factor (vWF). Results The function of hindl imb recovered rapidly in group E; the BBB score reached the peak at 8 weeks, and it was significantly higher than those in other groups (P lt; 0.05). The histology and ultrastructure observation showed that nerve fibers and neurons were damaged seriously in group B, oderately in groups C and D, and sl ightly in group E. Numerous spared tissue between nerve stumps, fibers with regular myel ination, and neurons with l ittle vacuolar mitochondria were observed in group E. The immunohistochemistry staining revealed that Caspase-3 positive cells in groups B, C, D, and E were significantly more than that in group A (P lt; 0.05); more Caspase-3 positive cells were found in groups B and D than in groups C and E (P lt; 0.05), while no significant difference was found between groups C and E (P gt; 0.05). And more vessels per high field were examined in groups C and E than in groups A, B, and D (P lt; 0.05), while no significant difference was found between groups C and E (P gt; 0.05). The p75NGFR positive results showed the survival of OECs in groups D and E at 8 weeks after OECstransplantation. Conclusion Grafted OECs combined with intrathecal injection of VEGF has significant promotive effects on restoration of spinal cord injury in rats, can improve part function of nerve fibers, and shows neuroprotection on damaged cells and fibers, which have a synergistic effect.
Objective To introduce the cells and cell-transplantation methods for periodontal tissue engineering. Methods Recent l iterature about appl ication of cell-based therapy in periodontal tissue engineering was extensively reviewed, the cells and cell-transplantation methods were investigated. Results Mesenchymal stem cells were important cell resourcesfor periodontal tissue engineering, among which peridontal l igament stem cells were preferred. Bone marrow mesenchymal stem cells had several disadvantages in cl inical appl ication, and adipose-derived stem cells might be a promising alternative; different transplantation methods could all promote periodontal regeneration to some extent. Single-cell suspension injection could only promote a l ittle gingival regeneration, and tissue engineered scaffolds still needed some improvement to be used in periodontal regeneration, while cell sheet technique, with great cell loading abil ity and no need of scaffolds, could promote regeneration of cementum, periodontal l igament, and alveolar bone under different conditions. Conclusion Multipotent stem cells are fit to be used in periodontal tissue engineering; improvement of cell-transplantation methods will further promote periodontal regeneration.
Objective To compare single cell suspension of neural stem cells (NSCs) with neurospheres transplantation for spinal cord injury (SCI) so as to explore the therapeutic effectiveness of two NSCs transplantation methods for SCI. Methods The NSCs were isolated from the spinal cord of adult Sprague Dawley (SD) rats, purified and cultured. At passage 3, the cells were identified by Hoechst33342, Nestin staining, and gl ial fibrillary acidic protein staining for differentiated cells. Sixty adult SD rats (weighing 230-250 g) were made the SCI models at T10 level with modified Allen method and randomlydivided into 3 groups (20 rats in each). The injury sites were treated by injecting 5 μL sal ine (group A), 5 μL single cellssuspensions of NSCs at passage 3 (group B), and 5 μL neurospheres cell suspensions at passage 3 (group C). At preoperation and 3, 7, 14, 21, and 28 days after operation, the locomotor functions of each group were assessed using the Basso, Beattie, and Bresnahan (BBB) rating scale. HE staining was applied to observe the morphology of spinal cord. Subsequently immunofluorescence staining was used to observe microtubule-associated protein 2 (MAP-2). Results The cells cultured were NSCs by morphological observation and immunofluorescence staining. After 3 days of modeling surgery, BBB score significantly decreased when compared with preoperative score, and there was no significant difference among 3 groups at 3 and 7 days (P gt; 0.05). BBB score increased in different degrees with time; at 14, 21, and 28 days, BBB score of groups B and C was better than that of group A, and group C was better than group B, showing significant differences (P lt; 0.05). HE staining showed that spinal cord structure of group C was more clear than that of groups A and B, and had less scar. There was no significant difference in the number of MAP-2 positive cells among 3 groups at 3 and 7 days (P gt; 0.05). At 14, 21, and 28 days, the number of MAP-2 positive cells of groups B and C was significantly more than that of group A, and group C was more than group B, showing significant differences (P lt; 0.05). Conclusion Transplantation of neurospheres suspension compared with single cell can significantly promote NSCsto differentiate into neurons and is conducive to recover the lower extremity function after SCI.
