Objective To observe the effects of the bone marrow mesenchymal stem cells (BMSCs) on the expression of neurotrophic factor protein gene in the retinal detachment (RD) rabbits. Methods 60 healthy rabbits were randomly divided into control group (group A), retinal detachment with PBS group (group B), retinal detachment with BMSCs group (group C), 20 rabbits in each group. RD model were established for rabbits in group B and C. 10 μl PBS was injected into the subretinal space of rabbits in group B, while 10 μl CM-Dil labeled BMSC PBS was injected into subretinal space of rabbits in group C. The rabbits in the group A received no treatment. At 1, 2 and 4 weeks after modeling, the mRNA expression of basic fibroblast growth factor (bFGF), brain derived neurotrophic factor (BDNF) and ciliary neurotrophic factor (CNTF) were measured by real-time quantitative PCR. Results At 1, 2 and 4 weeks after modeling, the mRNA expression of bFGF, BDNF, CNTF on retinal tissue were increased significantly in group C as compared with group A and B (P < 0.01). At 1 week after modeling, the mRNA expression of bFGF and CNTF on retinal tissue were increased significantly in group B as compared with group A, the mRNA expression of BDNF on retinal tissue in group B was similar with group C. At 2 and 4 weeks after modeling, the mRNA expression of bFGF, BDNF, CNTF were decreased in group B as compared with group A. Conclusion Subretinal transplantation of BMSC can increase the mRNA expression of bFGF, BDNF and CNTF on retinal tissue in RD rabbits.
Objective To observe the survival of human umbilical cord derived mesenchymal stem cells (hUC-MSCs) after injection into the vitreous of rabbits,and the animal safety under those procedures.Methods Twentyseven pigmented rabbits were randomly divided into 3 groups (intravitreal injection 1 week group，2 weeks group and 4 weeks group)， each with 9 rabbits.For each animal the right eye was the experimental eye receiving hUCMSCs injection,while the left eye was the control eye receiving culture medium. The rabbit eyes were examined by slitlamp microscope, indirect ophthalmoscopy, fundus photography, fundus fluorescence angiography（FFA）and Tonopen tonometer before and after injection. hUCMSCs were labeled by CMDil in vitro, and their survival status was measured by confocal fluorescence microscopy, light microscope and transmission electron microscope at 4 weeks after injection. Results Four weeks after injection, a large number of the hUCMSCs were still alive in the vitreous cavity. The overall condition of those rabbits was good. The anterior segment and retina of experimental eyes were normal, without hyperfluorescence, hypofluorescence and leakage in the retina at 1,2 and 4 weeks after injection. There was no significant difference on IOP before and after injection at different time points (P＞0.05), and no obvious changes at cornea, anterior chamber angle,lens,retinal structure by.light microscope and transmission electron microscope examination.Conclusion hUC-MSCs can survive in the rabbit vitreous for four weeks；intravitreal injection of hUCMSCs was safe and feasible.
ObjectiveTo observe the influence of human umbilical cord mesenchymal stem cells (hUCMSC) transplantation into vitreous cavity of diabetic rats on the retinal morphology, and the expression of glial fibrillary acidic protein (GFAP) and rhodopsin (RHO). Methods78 male Sprague-Dawley rats were used. 70 rats were injected with streptozotocin by tail vein injection at a dose of 40 mg/kg to establish the diabetes mellitus model, and another 8 rats were injected with 0.1 mol/L pH 4.0 citric acid buffer at the same dose as the normal control group. After 6 weeks of modeling, 10 rats were taken as the control group of diabetic model. hUCMSC suspension was injected into the right eye vitreous cavity of the remaining 60 rats, and the same volume of Dulbecco's modified Eagle/F12 medium was injected into the left vitreous cavity as control eyes. 1, 2 and 4 weeks after transplantation, follow-up experiments were performed. The experimental eyes were labeled as U1, U2, and U4 groups, while the control eyes were recorded as D1, D2, D4, and each group consisted of 20 eyes. After paraffin section and hematoxylin-eosin staining, the structure of the retina was observed by optical microscopy and the thickness of the outer nuclear layer and the inner nuclear layer (INL) were measured. The distribution and migration of hUCMSC in rat retina were observed by frozen section-tissue immunofluorescence assay. The mRNA and protein expression of GFAP and RHO in the retina were detected by real-time quantitative polymerase chain reaction (PCR) and Western blot assays. ResultsThe results of optical microscope observation showed the normal structure of retina in normal control group. The retinal nerve fiber layer (NFL) was thinned and the number of retinal ganglion cells (RGC) in the control group of diabetic rats was decreased. The decreased number and disorder arrangement of RGC were observed as well in U1, D1 rats. The RGC number of U2, U4, D2, D4 rats was gradually decreased. Compared with D4 group, the thickness of INL in U4 group was significantly increased (P < 0.05). Tissue immunofluorescence assay showed that hUCMSC were distributed along the inner limiting membrane in the retina of the U1 group, while the number of hUCMSC in the U2 group was gradually decreased, mainly in the NFL and ganglion cell layers. Real-time PCR and Western blot data indicated that the relative expression of GFAP mRNA and protein in the diabetic retina was significantly increased, and the relative expression of RHO mRNA and protein decreased gradually in the diabetic model group and the D1, D2, D4 groups. Compared with D2 and D4 groups, the mRNA and protein expression of GFAP in U2 and U4 groups were decreased, and the relative expression of RHO mRNA and protein were all increased (P < 0.01). ConclusionhUCMSC could migrate and integrate into the retina, after the transplantation into the vitreous cavity of diabetic rats, which reduced the expression of GFAP, but enhanced the expression of RHO.
Objective To observe the effects of subretinal transplantation of rat mesenchymal stem cells (rMSCs) on Sodium Iodate (SI)induced retinal degeneration. Methods One hundred and twenty BrownNorway (BN) rats were divided into three groups including SI injection group,rMSCs transplantation group and normal control group, each with 40 rats. The retinal degeneration was induced by caudal vein injection of SI. The retinal pigment epithelium（RPE）and neural retinal were evaluated by ocular fundus photograph, fluorescein fundus angiography (FFA)，electroretinogram (ERG) and histological approach, and TUNEL（terminal deoxynucleotidyl transferasemediated dUTP nick end labeling ）. CMDiIprelabeled primary rMSCs were transplanted into the subretinal space of SIinduced rats. The survival, integration, and differentiation of rMSCs were observed between 14 day to 60 day after the transplantation.Results The rat retinal function was gradually reduced after14 days of SI injection, with a timedependent manner. After the RPE cells were damaged，the outer segments of photoreceptors became disrupted and shortened until karyopyknosis. The nuclear morphology and positive TUNEL labeling indicated that the death of photoreceptor cells was apoptosis. After rMSCs transplantation, CMDiI labeled donor cells were observed to be scattered in the subretinal space and expressed RPE cell markers. Average amplitude of b wave and Ops (oscillation potential) in ERG improved 27.80%,59.38% respectively after rMSCs transplantation.Conclusions Transplanted rMSCs can survive in subretinal space and differentiate into RPE.
ObjectiveTo observe the morphological and functional changes of retinal degeneration in mice with CLN7 neuronal ceroid-lipofuscinosis, and the therapeutic effects of glial cell derived neurotrophic factor (GDNF) and/or ciliary neurotrophic factor (CNTF) based on neural stem cells (NSC) on mouse photoreceptor cells. MethodsA total of 100 CLN7 mice aged 14 days were randomly divided into the experimental group and the control group, with 80 and 20 mice respectively. Twenty C57BL/6J mice aged 14 days were assigned as wild-type group (WT group). Mice in control group and WT group did not receive any interventions. At 2, 4, and 6 months of age, immunohistochemical staining was conducted to examine alterations in the distribution and quantity of cones, rod-bipolar cells, and cone-bipolar cells within the retinal of mice while electroretinography (ERG) examination was utilized to record scotopic a and b-waves and photopic b-wave amplitudes. At 14 days of age, the mice in the experimental group were intravitreally injected with 2 μl of CNTF-NSC, GDNF-NSC, and a 1:1 cell mixture of CNTF-NSC and GDNF-NSC (GDNF/CNTF-NSC). Those mice were then subdivided into the CNTF-NSC group, the GDNF-NSC group, and the GDNF/CNTF-NSC group accordingly. The contralateral eyes of the mice were injected with 2 μl of control NSC without neurotrophic factor (NTF) as their own control group. At 2 and 4 months of age, the rows of photoreceptor cells in mice was observed by immunohistochemical staining while ERG was performed to record amplitudes. At 4 months of age, the differentiation of grafted NSC and the expression of NTF were observed. Statistical comparisons between the groups were performed using a two-way ANOVA. ResultsCompared with WT group, the density of cones in the peripheral region of the control group at 2, 4 and 6 months of age (F=285.10), rod-bipolar cell density in central and peripheral retina (F=823.20, 346.20), cone-bipolar cell density (F=356.30, 210.60) and the scotopic amplitude of a and b waves (F=1 911.00, 387.10) in central and peripheral retina were significantly decreased, with statistical significance (P＜0.05). At the age of 4 and 6 months, the density of retinal cone cells (F=127.30) and b-wave photopic amplitude (F=51.13) in the control group were significantly decreased, and the difference was statistically significant (P＜0.05). Immunofluorescence microscopy showed that the NSC transplanted in the experimental group preferentially differentiated into astrocytes, and stably expressed CNTF and GDNF at high levels. Comparison of retinal photoreceptor nucleus lines in different treatment subgroups of the experimental group at different ages: CNTF-NSC group, at 2 months of age: the whole, central and peripheral regions were significantly different (F=31.73, 75.06, 75.06; P＜0.05); 4 months of age: The difference between the whole area and the peripheral region was statistically significant (F=12.27, 12.27; P＜0.05). GDNF/CNTF-NSC group, 2 and 4 months of age: the whole (F=27.26, 27.26) and the peripheral area (F=16.01, 13.55) were significantly different (P＜0.05). In GDNF-NSC group, there was no statistical significance at all in the whole, central and peripheral areas at different months of age (F=0.00, 0.01, 0.02; P＞0.05). ConclusionsCLN7 neuronal ceroid-lipofuscinosis mice exhibit progressively increasing degenerative alterations in photoreceptor cells and bipolar cells with age growing, aligning with both morphological and functional observations. Intravitreal administration of stem cell-based CNTF as well as GDNF/CNTF show therapeutic potential in rescuing photoreceptor cells. Nevertheless, the combined application of GDNF/CNTF-NSC do not demonstrate the anticipated synergistic protective effect. GDNF has no therapeutic effect on the retinal morphology and function in CLN7 neuronal ceroid-lipofuscinosis mice.
Objective To observe the retinal apoptosis of laser-induced retinal injury in mice after bone marrow mesenchymal stem cells transplantation. Methods Green fluorescent protein (GFP) labeled MSCs from C57BL/6 mice were cultured in vitro. A total of 135 C57BL/6 mice were divided into three groups including normal control group (15 mice), injured control group (60 mice) and MSCs treatment group (60 mice). Laser retinal injuries were induced by laser photocoagulation. One day after photocoagulation, 02 ml cell suspension, which contained 1times;106 GFP-MSCs, were injected into the mice in treatment group via tail vein, and the mice in injured control group were given equal volume of phosphate buffer solution. Animal were execute on three, seven, 14 and 21 days following laser damage. Hematoxylin and eosin (HE) staining was performed to assess the changes of injured retinas. The diameters of laser spots and areas with total loss of cells in outer nuclear layer (ONL) were analyzed by image processing software. The apoptosis of retinal cells was examined by terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL) staining. The migration of GFP-MSCs into the retina was observed by fluorescence microscope. Results HE staining showed that the retinal structures were integrated in normal control group. Retinal damages were observed both in injured control group and MSCs treatment group, but milder in the latter. Though the average diameter of area with total loss of cells in ONL of MSCs treatment group was less than the injured control group (t=5.769, P＜0.05), the diameters of laser spots show no difference (t=0.964,P＞0.05) on day three. Both the average diameter of laser spots (t=5.180, 5.417, 2.381) and area with total loss of cells in ONL (t=3.530, 3.224, 3.162) were less in the MSCs treatment group on day seven, 14 and 21 (P＜0.05). TUNEL staining shows that the apoptosis were decreased after MSCs transplantation on day three, seven, 14 and 21 (t=11.142, 7.479, 6.678, 3.953,P＜0.05). No apoptosis was observed in normal control group. Very few GFP-MSCs were observed in the retina at all time-points. They were only seen in the subretinal and choroidal neovascularization occasionally on day seven and 14. Conclusion MSCs transplantation can effectively limit the range of retinal laser damage and inhibit cell apoptosis.
ObjectiveTo investigate the behavioral recovery of spinal cord injury (SCI) rats that received transplantation of NEP1-40 gene-modified neural stem cells. MethodsNeural stem cells (NSCs) were derived from the cortex tissue of rat embryo at the age of 18 days and identified by Nestin immunofluorescence. The lentiviruses were transduced to NSCs to construct NEP1-40 gene modified NSCs. Spinal cords of 30 Sprague-Dawley rats were hemisected at the nineth thoracic vertebrae level. The rats were randomly assigned to three groups. Cell culture medium, NSCs and NEP1-40 gene-modified NSCs were transplanted into the lesion site of rats of SCI group, NSCs group and NEP1-40-NSCs group respectively 7 days after injury. Additional 10 rats served as blank control group (sham group), which only received laminectomy. Following transplantation, behavior tests including Basso, Beattie, Bresnahan (BBB) Locomotor Rating Scale and grid test were utilized to evaluate spinal cord functional recovery. ResultsBehavior tests 8 weeks after cells transplantation showed that the rats in SCI group got worst results, the BBB scores improved and the grid drop times reduced significantly in NSCs transplantation group (P<0.01) and behavioral test outcomes were best in the NEP1-40 gene-modified NSCs group (P<0.01). ConclusionNEP1-40 gene modification can significantly improve the behavioral recovery of SCI rats that received transplantation of pure neural stem cells. It can provide a new idea and reliable experimental base for the study of NSCs transplantation for spinal cord injury.
Objective To systematically review the survival outcome and safety of haploidentical hematopoietic stem cell transplantation (haplo-HSCT) for β-thalassemia. Methods The PubMed, EMbase, CNKI, WanFang Data and CBM databases were electronically searched to collect studies on haplo-HSCT for β-thalassemia from January 1, 2017 to December 31, 2021. Two reviewers independently screened the literature, extracted data and assessed the risk of bias of the included studies. Meta-analysis was then performed by using RevMan 5.4.1 software and Stata 16.0 software. Results A total of 6 case-series studies involving 286 patients were included. The results of meta-analysis indicated that overall survival (OS) and thalassemia-free survival (TFS) for β-thalassemia patients undergoing haplo-HSCT were 92.5% (95%CI 86.1% to 96.1%) and 88.5% (95%CI 74.6% to 95.3%), the incidence of Ⅲ-Ⅳ degree acute graft versus host disease (Ⅲ-Ⅳ aGvHD) and chronic graft versus host disease (cGvHD) were 11.5% (95%CI 6.5% to 20.0%) and 23.1% (95%CI 12.3% to 39.8%), and the transplantation related mortality was 6.5% (95%CI 3.8% to 10.7%). Conclusion Relevant clinical studies published in the past 5 years provide the latest information and progress of haplo-HSCT for β-thalassemia. At present, great efficacy has been shown in NF-14-TM therapeutic regimen, but the long-term efficacy remains unclear. Due to the limited quality and quantity of the included studies, more high-quality evidence from long-term comparative studies is still needed.
ObjectiveTo systematically review clinical efficacy and safety of bone marrow stem cells transplantation in treating primary dilated cardiomyopathy (DCM). MethodsSuch databases as PubMed, CENTRAL, EMbase, Web of Knowledge, VIP, CNKI, CBM and WanFang Data were searched from inception to March 2014 for the randomized controlled trials (RCTs) about bone marrow stem cells transplantation for DCM. According to the inclusion and exclusion criteria, two reviewers independently screened literature, extracted data, and assessed methodological quality of included studies. Then meta-analysis was performed using RevMan 5.2.0 software. ResultsA total of ten RCTs involving 374 patients were included. The results of meta-analysis showed that, a) for safety, after 3 months there was no significant difference in the incidence of malignant arrhythmia events between bone marrow stem cell transplantation group and routine treatment group (RR=0.81, 95%CI 0.38 to 1.72, P=0.58); and b) for efficacy, compared with the control group, left ventricular ejection fraction (LVEF) increased in the bone marrow stem cell transplantation group after 3 months (WMD=3.86, 95% CI 2.53 to 5.20, P<0.000 01) and after 6 months (WMD=5.54, 95%CI 3.02 to 8.06, P<0.000 1). The bone marrow stem cell transplantation group were better in increased 6-minute walking distance after 3 months (WMD=22.12, 95%CI 7.78 to 36.46, P=0.003), increased 6-minute walking distance after 6 months (WMD=102.79, 95%CI 50.16 to 155.41, P=0.000 1), decreased perfusion defect of myocardium percentage after 3 months (WMD=-4.00, 95%CI -5.87 to -2.13, P<0.000 1). However, there was no significant difference in left ventricular end-diastolic diameter (LVEDD) between two groups after 3 months (WMD=-0.37, 95%CI -1.67 to 0.93, P=0.57) and after 6 months (WMD=-0.70, 95%CI -2.76 to 1.36, P=0.51). ConclusionBone marrow stem cells transplantation for dilated cardiomyopathy is effective in improve patients' heart function with good safety, with significant difference. Due to limited quantity and quality of the included studies, more high quality and large-scale RCTs are needed to verify the above conclusion.