Influence of quercetin on aging of bone marrow mesenchymal stem cells induced by microgravity / 中国药理学通报

Aim To investigate the effect of quercetin on the aging model of bone marrow mesenchymal stem cells established under microgravity. Methods Using 3D gyroscope, a aging model of bone marrow mesenchymal stem cells was constructed, and after receiving quercetin and microgravity treatment, the anti-aging effect of the quercetin was evaluated by detecting related proteins and oxidation indexes. Results Compared to the control group, the expressions of age-related proteins p21, pi6, p53 and RB in the microgravity group significantly increased, while the expressions of cyclin D1 and lamin B1 significantly decreased, with statistical significance (P<0.05). In the microgravity group, mitochondrial membrane potential significantly decreased (P<0.05), ROS accumulation significantly increased (P <0.05), SOD content significantly decreased and MDA content significantly increased (P<0.05). Compared to the microgravity group, the expressions of age-related proteins p21, pi6, p53 and RB in the quercetin group significantly decreased, while the expressions of cyclin D1 and lamin B1 significantly increased, with statistical significance (P<0.05). In the quercetin group, mitochondrial membrane potential significantly increased (P<0.05), ROS accumulation significantly decreased (P<0.05), SOD content significantly increased and MDA content significantly decreased (P<0.05). Conclusions Quercetin can resist oxidation, protect mitochondrial function and normal cell cycle, thus delaying the aging of bone marrow mesenchymal stem cells induced by microgravity.

Research Advances on the Role of Bone Marrow Stromal Cell in Acute Lymphoblastic Leukemia --Review / 中国实验血液学杂志

Acute lymphoblastic leukemia (ALL) is a kind of the most common hematopoietic malignancy, its recurrence and drug resistance are closely related to the bone marrow microenvironment. Bone marrow stromal cell (BMSC) is an important part of the bone marrow microenvironment and their interaction with leukemia cells cannot be ignored. BMSC participates in and regulate signaling pathways related to proliferation or apoptosis of ALL cells by secretes cytokines or extracellular matrix proteins, thus affecting the survival of ALL cells. In this review, the research advance of several signaling pathways of the interaction between BMSC and ALL cells was summarized briefly.

Extracellular vesicle activities regulating macrophage- and tissue-mediated injury and repair responses

Macrophages are typically identified as classically activated (M1) macrophages and alternatively activated (M2) macrophages, which respectively exhibit pro- and anti-inflammatory phenotypes, and the balance between these two subtypes plays a critical role in the regulation of tissue inflammation, injury, and repair processes. Recent studies indicate that tissue cells and macrophages interact

Anti-apoptotic effect of human acute myeloid leukemia cell line HL-60 stimulated by interleukin-6 from bone marrow stromal cells HS-5 / 解剖学报

Objective To investigate the effect of interleukin( IL)-6 secreted by bone marrow stromal cells HS-5 on activity and apoptosis of human acute myeloid leukemia( AML) cells HL-60 and its possible mechanism. Methods HL-60 cells and HS-5 cells were cultured in vitro, and the co-culture system was established. Scanning electron microscope, ELISA, cell counting kit-8( CCK-8) , AnnexinV-FITC/PI, double-staining flow cytometry, Real-time PCR and Western blotting techniques were used to detect the changes of viability and apoptosis of HL-60 cells, respectively. HL-60 cells from different groups were inoculated into BALB/c nude mice to observe and record tumorigenesis. Results IL-6 secreted by bone marrow stromal cells HS-5 could enhance the viability and inhibit the apoptosis of HL-60 cells, and down-regulate the expression of pro-apoptotic gene Bax and and up-regulate the expression of anti-apoptotic gene Bcl-2 in HL-60 cells. HL-60 cells in co-culture group had the strongest tumorigenicity in BALB/c nude mice, while HL-60 cells alone had the weakest tumorigenicity. Conclusion Part of the mechanism by which bone marrow stromal cells HS-5 promote the proliferation of HL-60 cells and inhibit their apoptosis may be through the secretion of IL-6.

Research Advances in the Effect of Mechanical Regulation of Long-Chain Non-Coding RNAs on Osteogenic Differentiation of Mesenchymal Stem Cells / 医用生物力学

With the development of the 3rd-generation high-throughput sequencing technology and tissue engineering, recent studies show that many long-chain non-coding RNAs (LncRNAs) have played an important role in osteogenic differentiation of mesenchymal stem cells (MSCs). LncRNAs, which are involved in the regulation of mechanical regulation, further regulate bone-related cell functions and play a regulatory role at multiple levels, including transcription, post-transcriptional and epigenetic. LncRNAs may be involved in the osteogenic differentiation and bone remodeling of MSCs, the regulation of bone-related cell functions as a mechanical response molecule, as well as the pathological process of skeletal diseases.

Generation of Retinal Pigmented Epithelium-Like Cells from Pigmented Spheres Differentiated from Bone Marrow Stromal Cell-Derived Neurospheres

BACKGROUND: Retinal degeneration causes blindness, and cell replacement is a potential therapy. The purpose of this study is to formation of pigmented neurospheres in a simple medium, low-cost, high-performance manner over a short period of time while expressing markers of RPE cells and the activation of specific genes of the pigment cells. Also, these neurospheres have the ability to produce a monolayer of retinal pigment epithelium-like cells (RPELC) with the ability of photoreceptor outer segment phagocytosis. METHODS: BMSC were isolated from pigmented hooded male rats and were immunoreactive to BMSC markers, then converted into neurospheres, differentiated into pigmented spheres (PS), and characterized using Retinal pigment epithelium-specific 65 kDa protein (RPE65), Retinaldehyde-binding protein 1 (CRALBP) and orthodenticle homeobox 2 (OTX2) markers by immunocytochemistry, RT-PCR and RT-qPCR. The PS were harvested into RPELC. The functionality of RPELC was evaluated by phagocytosis of fluorescein-labeled photoreceptor outer segment. RESULTS: The BMSC immunophenotype was confirmed by immunostained for fibronectin, CD90, CD166 and CD44. These cells differentiated into osteogenic and lipogenic cells. The generated neurospheres were immunoreactive to nestin and stemness genes. The PS after 7–14 days were positive for RPE65 (92.76–100%), CRALBP (95.21–100%) and OTX2 (94.88–100%), and after 30 days RT-PCR, qPCR revealed increasing in gene expression. The PS formed a single layer of RPELC after cultivation and phagocyte photoreceptor outer segments. CONCLUSION: Bone marrow stromal stem cells can differentiate into functional retinal pigmented epithelium cells in a simple, low-cost, high-performancemanner over a short period of time. These cells due to expressing theRPELCgenes andmarkers can be used in cell replacement therapy for degenerative diseases including age-relatedmacular degeneration as well as retinitis pigmentosa.

Trim33 promotes lipid accumulation and expressions of adipocyte-specific genes in bone marrow stromal cells / 医学研究生学报

ObjectiveTripartite motif 33 (Trim33) is known to play a very important part in regulating osteoblast differentiation, but its role in adipocyte differentiation is rarely reported. The aim of this study was to investigate the regulatory effect of Trim33 on adipocyte differentiation.MethodsBone marrow stromal cell ST2 cells transfected with the Trim33-pcDNA3.1 plasmid were included in the experimental group and those transfected with the pcDNA3.1 plasmid taken as the control. The cells of both groups were treated with adipogenic medium to induce adipocyte differentiation, followed by determination of the expressions of the adipocyte-specific genes CCAAT enhancer binding protein alpha (C/EBPα), peroxisome proliferator-activated receptor gamma (PPARγ), adipocyte characterizing factor FABP4, and adipocytokines adipsin by oil red O staining, qRT-PCR and Western blotting.ResultsThe expression of Trim33 was significantly upregulated in the experimental group as compared with that in the control (88.51±14.31 vs 1.00±0.31, P<0.01). After 5 days of adipogenesis induction, there were dramatically more lipid droplets in the ST2 cells and the A value was markedly higher in the former than in the latter group (0.69±0.03 vs 0.34±0.03, P<0.01). Compared with the control, the cells in the experimental group exhibited remarkable increases in the relative mRNA expressions of C/EBPα, PPARγ, FABP4 and adipsin as well as the protein expressions of Trim33, PPARγ, C/EBPα and FABP4.ConclusionTrim33 promotes lipid accumulation and upregulates the expressions of adipocyte-specific genes in bone marrow stromal cells.

Osteoking Promotes the Proliferation of BMSC through Activating TGF-β/Smad Signaling Pathway in Castrate Osteoporosis Rats / 昆明医科大学学报

Objective To study the effect of OK on the differentiation of bone marrow stromal stem cells (BMSCs) through the TGFβ/Smad signaling pathway in rats. Methods We removed the bilateral ovaries of rats to replicate OP model, and isolated and cultured BMSCs. BMSCs isolated from nomal rats were cultured as control group, BMSCs isolated from OP rats were divided into 5 groups, OP model group was regularly cultured, positive control group was treated with alendronate sodium (1 μmol/L), low, medium and high OK treatment group were treated with 50 mL/L, 100 mL/L and 200 ml/L OK respectively.After 24 h incubation, all cells were collected. The proliferation rate of BMSCs was determined by MTT method, and ELISA method was employed to detect the contents of bone formation markers, RT-qPCR was used to determine the m RNA expression level of TGFβ and the protein expression levels of TGFβ, p-Smad2/3 and of Smad2/3 were detected by Western blot. Results Compared with control group, the proliferation rate of the BMSCs in OP model group was reduced, concentrations of bone formation markers (OC, PINP and BALP) were reduced, m RNA and protein expression levels of TGFβ, as well as the phosphorylated level of Smad2/3 were downregulated.The difference was statistically significant (P＜0.05). After treatment with OK, compared with model group, all the above effects were ameliorated in different degree, a dose dependent manner was observed in OK treatment group, and the treatment effects of alendronate sodium (1 μmol/L), 100 mL/L and 200 mL/L OK group were statistically significant (P ＜0.05).Conclusion OK can promote the proliferation of osteoblasts by activating TGFβ/Smad signaling pathway to achieve the effect of treating postmenopausal OP.

Experimental study of lentiviral vector-mediated bFGF gene transfection on rabbit BMSCs / 天津医药

Objective To observe the effect of lentiviral vector-mediated basic fibroblast growth factor (bFGF) gene transfection on the biological characteristics of rabbit bone marrow stromal cells(BMSCs)under in vitro culture conditions. Methods BMSCs were obtained by density gradient centrifugation and adherence screening. The bFGF gene was transfected into BMSCs by lentiviral vector and divided into bFGF transfection group,empty virus group and untransfected group according to the transfection conditions.After transfection,the morphology,expressions of bFGF mRNA and protein, cell proliferation,cell cycle and alkaline phosphatase(ALP)activity were observed in three groups of cells. Results High density BMSCs were successfully obtained by density gradient centrifugation and adherence screening.After transfection of BMSCs with bFGF gene, the cell morphology showed no significant changes, while the expressions of bFGF mRNA and protein were significantly increased, the cell proliferation curve shifted upward, the proportion of proliferating cells increased,and the activity of ALP was significantly enhanced.There were significant differences between three groups(P<0.05).Conclusion The rabbit bFGF gene is successfully introduced into the BMSCs cultured in vitro by lentiviral vector, and the target gene is stably expressed.The expression of bFGF can promote the proliferation and osteogenic differentiation of BMSCs.

Effects of canine bone marrow stromal cells after different stages of mineralization on the differentiation, proliferation of canine periodontal ligament stem cells / 口腔疾病防治

Objective @#To investigate the inductive effects of canine periodontal ligament stem cells (PDLSCs) cocultured with canine disparate differentiating-period iliac bone marrow stromal cells (I-BMSCs).@*Methods@#Purified PDLSCs were isolated by in vitro culture of cBMSCs and flow cytometry. Third-generation PDLSCs were obtained, and conditioned culture medium derived fromiliac bone marrow stromal cells (I-BMSCs-CM) was added as indicated for coculture of PDLSCs. As the control group, pure uninduced PDLSCs were routinely cultured in DMEM culture medium containing 15%FBS. The experimental groups included the I-BMSCs-CM, I-BMSCs-CM-10ds and I-BMSCs-CM-15ds groups. The I-BMSCs-CM group consisted of PDLSCs induced by I-BMSCs, the I-BMSCs-CM-10ds group consisted of PDLSCs induced by I-BMSCs after osteogenic induction for 10 days, and the I-BMSCs-CM-15ds group consisted of PDLSCs induced by I-BMSCs after osteogenic induction for 15 days. The cocultured PDLSCs were examined via the MTT assay. Total mRNA and protein were prepared at 3 and 7 days. The mRNA expression levels of runt-related transcription factor 2(Runx2), special AT-rich sequence binding protein 2(Satb2) and osteocalcin (OCN) were measured by qRT-PCR. The protein expression levels of Satb2, Runx2 and OCN were detected by Western blot.@*Results@#The PDLSCs showed a spindle-like morphology. While the BMSC-conditioned media increased PDLSCs proliferation, the media conditioned by BMSCs allowed to differentiate for 15 days (I-BMSCs-CM-15days) significantly enhanced PDLSCs proliferation (F=342.8, P=0.017). The expression levels of the analyzed genes were upregulated in the coculture groups, and the protein expression levels of Satb2, Runx2 and OCN were higher in the test groups than in the control group at 7 days. At the protein level, I-BMSCs-CM-15days upregulated the expression of Satb2 by 3.04-fold (FSatb2=24.48, P=0.014), Runx2 by 5.1-fold (FRunx2=12.25, P < 0.001), and OCN by 3.67-fold (FOCN=18.35, P=0.022).@*Conclusion @#The conditioned medium of I-BMSCs may enhance the proliferation of PDLSCs, and that of terminally differentiated bone cells probably triggered the osteogenesis of PDLSCs, suggesting important implications for periodontal engineering.

Effects of bone marrow stromal cells on the chemotherapeutic sensitivity of acute lymphoblastic leukemia cells / 中华肿瘤杂志

Objective@#To investigate the influences of bone marrow stromal cells, components of extracellular matrix and cytokine secreted by stromal cells on the chemotherapeutic sensitivity of acute lymphoblastic leukemia cells to cytosine arabinoside (Ara-C).@*Methods@#The co-culture model of acute lymphoblastic leukemia cell Sup-B15 and bone marrow stromal cell OP9 was constructed. Sup-B15 cells were cultured alone or co-cultured with OP9 cells, inactivated OP9 cells, the conditional medium (CM) of co-cultured OP9 cells and Sup-B15 cells, the CM of OP9 cells alone or Sup-B15 cells alone, respectively. The effects of different concentrations of Ara-C on the proliferation of each Sup-B1 cell group mentioned above were detected by cell counting kit-8 (CCK-8) method. The effects of different concentrations of Ara-C on the apoptosis of each group were detected by flow cytometry (FCM). The expressions of Bcl-2 protein in each group were detected by western blot.@*Results@#The results of CCK-8 test showed that the inhibitory efficiency of Ara-C was in a dose-dependent manner. With different concentrations of Ara-C treatment for 48 hours, the half maximal inhibitory concentrations (IC50) of Sup-B15 and OP9 co-cultured group, Sup-B15 and inactivated OP9 co-cultured group were 0.510 and 0.339 μg/ml, respectively, significantly higher than 0.091 μg/ml of Sup-B15 cultured alone group (P<0.05). The IC50 of CM of Sup-B15 and OP9 co-cultured group was 0.204 μg/ml, significantly higher than 0.087 μg/ml of the CM of OP9 cultured alone group (P<0.05) and 0.097 μg/ml of the CM of Sup-B15 cultured alone group (P<0.05). The results of flow cytometry showed that with 0.10 μg/ml Ara-C treatment for 24 hours, the early apoptotic cell percentages of Sup-B15 and OP9 co-cultured group, Sup-B15 and inactivated OP9 co-cultured group and Sup-B15 cultured alone group were (6.67±2.19) %, (8.95±3.04) % and (20.46±2.63) %, respectively. The early apoptotic cell percentages of Sup-B15 and OP9 co-cultured group, Sup-B15 and inactivated OP9 co-cultured group were significantly lower than that of Sup-B15 cultured alone group (P<0.05). The early apoptotic cell percentages of the CM of Sup-B15 and OP9 co-cultured group, the CM of OP9 cultured alone group and the CM of Sup-B15 cultured alone group were (11.16±2.97)%, (22.08±2.71)% and (19.25±1.57)%, respectively, the former two of which were significantly lower than the last one (P<0.05). The results of western blot showed that the relative expression levels of Bcl-2 protein of Sup-B15 cultured alone group, Sup-B15 and OP9 co-cultured group, Sup-B15 and inactivated OP9 co-cultured group, the CM of Sup-B15 and OP9 co-cultured group, the CM of OP9 cultured alone group and the CM of Sup-B15 cultured alone group were 1.00±0.00, 1.53±0.03, 1.38±0.01, 1.26±0.05, 1.03±0.01 and 0.98±0.02, respectively. The expression levels of bcl-2 protein of three combined groups were significantly higher than that of Sup-B15 cultured alone group (P<0.05). while no statistically significant difference was observed between the CM of OP9 cultured alone group and the CM of Sup-B15 cultured alone group (P>0.05).@*Conclusion@#Bone marrow stromal cell OP9, the components of bone marrow extracellular matrix and cytokine secreted by stromal cells are involved in the induction of the chemotherapeutic resistance of Sup-B15 cells to Ara-C.

Senescence induced by D-galactose and its biological mechanism in rat bone marrow stromal cells / 基础医学与临床

Objective To establish an aging model of rat bone marrow stromal cells (BMSCs) in vitro and in vivo, in order to study the senescence biology of aging BMSCs .Methods The control cell group ( in vitro):isolating, puri-fying and culturing BMSCs from healthy male SD rats .collecting the third generation ( P3) of BMSCs for analysis . The aging model group (in vitro):the P3 BMSCs were incubated with D-Galactose (D-Gal, 30 g/L) for 48 hours. The aging rat model group ( in vivo): the rats were given 120 mg D-Gal by the way of daily neck subcutaneous injection for 42 consecutive days .The control rat group ( in vivo):the rats were administrated with the same volume of saline for the same times .On the second day after the aging model was established , the BMSCs were collecting and culturing for study.1)The proliferative potency was detected by cell counting Kit-8(CCK-8);the distribution of cell cycle and apoptosis was detected by flow cytometry (FCM);2)the ratio of aging BMSCs was examined by the senescence-associated β-Galactosidase(SA-β-Gal) staining;3)malonaldehyde(MDA) content and total super-oxide dismutase(SOD) was examined activity by enzymatic assay; the level of reactive oxygen species (ROS) by DCFH-DA fluorescent staining was counted with FCM;4 ) the expression level of senescence-related signaling was proteins of P16 , P21 , P53 , CDK2 and cyclin D by Western blot .Results Compared with the matched control group, the BMSCs of aging model group displayed a significant decrease in proliferation; the BMSCs were held in G1 phase arrest as the proportion of the cells in G 1 phase increased , while that decreased in S phase ( P<0.05 );and the positive ratio of SA-β-Gal stained BMSCs also significantly increased ( P <0.05 ); BMSCs in the aging model group showed an increasing level of ROS and MDA , meanwhile a decline in total SOD activity was decreased (P<0.05);P16,P21 and P53 protein expression in aging BMSCs was obviously enhanced (P<0.05), at the same time the expression of CDK2 and cyclin D was also decreased ( P<0.05 ) .Conclusions D-Gal can be used to develope an aging model of BMSCs .It acts through up-regulation of expressions of aging-related proteins and in-hibition of oxidative stress injury and chronic inflammation .

Human leptin gene overexpression enhances osteogenesis of bone marrow stromal cells from rats / 基础医学与临床

Objective To investigate the effects of human leptin ( hLEP) gene transfection on rat bone marrow stro-mal cells ( rBMSCs) .Methods rBMSCs were cultured and transfected with adenoviruses encoding hLEP ( Ad5-hLEP-EGFP) in vitro as experimental group while rBMSCs transfected with Ad 5-EGFP and non-transfected were control groups.The proliferation was detected by MTT and the expression of collagen type Ⅰ(Col-Ⅰ) and alkaline phosphatase ( ALP) were assessed by real-time PCR.The ability of mineralized nodule forming was also examined by Alizarin red staining .The combination of transfected rBMSCs and β-tricalcium phosphate (β-TCP ) was con-structed and the osteogenic ability of the construction was evaluated in nude mice .Results hLEP could be trans-fected into rBMSCs successfully by adenovirous .After transfection , the proliferation was not affected while Col-Ⅰand ALP expressions were more pronounced in rBMSCs transfected with Ad 5-hLEP-EGFP ( P<0.05 ) .Alizarin red staining showed the ability of mineralized nodule forming was also up-regulated in Ad5-hLEP-EGFP group (P<0.05).In addition, the transfected rBMSCs adhered to β-TCP and survived well and the combination showed more new bone like tissue formation in nude mice compared to control groups .Conclusions rBMSCs transfected with hLEP might be potently used in bone or periodontal tissue regeneration .

Bone marrow stromal cells as a therapeutic treatment for cerebral ischemic / 国际生物医学工程杂志

Bone marrow stromal cells (BMSCs) are a kind of stem cells with multiple differentiation potential in bone marrow.BMSCs have been widely used in tissue engineering,cell transplantation,gene therapy and organ transplantation,due to their characteristics of wide range of sources,weak immunogenicity weak,easily transfected by exogenous gene,long survival time in the host,multi-directional differentiation,etc.Cerebral ischemia is caused by neurological impairment,which is the most common cause of death and quality-of-life impairments.The clinical manifestations of the patients with cerebral ischemia are motor function failure,sensory dysfunction and abnormal mental consciousness.A large number of studies have reported that BMSCs transplantation has the therapeutic effects of body sensory and motor function recovery,and can treat ischemic stroke.BMSCs transplantation has brought new hope for the clinical treatment of ischemic cerebrovascular disease.In this paper,the recent progress in the study of BMSCs transplantation for ischemic stroke was reviewed.The mechanism,pathways,influencing factors and clinical application of BMSCs transplantation were summarized.

Construction of miR-196b sponge lentiviral vector and its expression in bone marrow stromal cell line ST2 / 天津医药

Objective To construct miR-196b sponge lentiviral vector,and laid the foundation for studying the function of miR-196b in bone marrow stromal cells.Methods Based on the miR-196b mature sequence,a sequence consisting of 6 tandem repeats of the complementary sequence of miR-196b was designed,and which was cloned into pUC19 plasmid by using reverse PCR.Then the six-repeat sequence was cut and subcloned into pLVX-shRNA2 lentiviral vector.The lentivirus was packaged using 293T cells,and titer determination was done.The pLVX-shRNA2 lentivirus was used as the control group,and the 196b-sponge-pLVX lentivirus was the experimental group.Then ST2 cells were infected with the viruses,and the infection efficiency was calculated.The protein level of forkhead box O1 (FoxO1) was detected by Western blot assay.Results The identity of the sponge sequence was verified by sequencing.The titer of the sponge virus was 1 × 108 PFU/mL,and the infection efficiency reached 80％.Compared with the control group,the expression level of FoxO 1 protein was significantly increased (P ＜ 0.05).Conclusion The miR-196b sponge lentiviral vector is successfully constructed,and which has the capability to inhibit endogenous miR-196b.

Effect of TERT gene transfected BMSC on memory function and hippocampal CA1 region synaptic plasticity in vascular dementia rat / 重庆医学

Objective To explore the effect of telomerase reverse transcriptase(TERT) gene transfected bone marrow stem cell(BMSC)on the memory function and hippocampal CA1 region synaptic plasticity in vascular dementia rat.Methods A total of 60 rats were randomly divided into the negative control group(group A),model group(group B),conventional BMSC group(group C) and transfected BMSC group(group D).The related indicators in each group were detected by using the Morris maze test,RTPCR and Western blot respectively.Results The escape latency period in the group C and group D was significantly longer than that in the group B,which in the group D was significantly longer than that in the group C.Compared with the group A,the expressions of brain-derived neurotrophic factor(BDNF)mRNA,TERT mRNA,SYP mRNA and protein in the group B,group C and group D were significantly decreased.The synaptic cleft arrange in group A was clear with more SYN positive ceils.The synaptic cleft in the group D was clearer,and the number of SYN positive cells was close to that in group A.Conclusion TERT transfected BMSC has obvious therapeutic effect on vascular dementia rats and its mechanism may be related to the promotion of BDNF,TrkB expression and the improvement of synaptic plasticity.

Effects of Bone Marrow Stromal Cells Transplantation Combined with Ultrashort Wave Therapy on the Expression of GFAP and ED-1 after Spinal Cord Injury in Rats / 中国医科大学学报

Objective To investigate the effects of bone marrow stromal cells(BMSCs)transplantation combined with low dose ultrashort wave (USW)radiation on functional recovery and the expression of glial fibrillary acidic protein(GFAP)and ED?1 after spinal cord injury(SCI)in rats,and further discuss its action mechanism. Methods Female Sprague?Dawley rats(n=30)were randomly divided into 5 groups:sham?oper?ated,as well as control,USW,BMSCs,and USW+BMSCs that were subjected to spinal cord injury(SCI). Basso?Beattie?Bresnahan(BBB)tests were carried out before the operation and at 1 d,1 week,2 weeks,3 weeks,4 weeks after SCI. 4 weeks later,animals were sacrificed and tissues were collected to make paraffin section. Immunohistochemical staining was performed to observe the expression of GFAP and ED?1. Results 4 weeks after SCI,BBB scores were significantly higher in the USW and USW+BMSCs groups than in the control group(both P<0.001). No signifi?cant difference was observed between the BMSCs group and control group. On the expression of GFAP ,only USW+BMSCs group showed signifi?cantly decreased compared with the control group(P<0.05). All treatment groups exhibited lower ED?1 expression than the control group(all P<0.05). Conclusion Our results indicate that USW radiation alone can obviously improve neural functional recovery after SCI. The USW radi?ation and BMSCs transplantation treatment can reduce inflammation ,and USW radiation is more effective. The combination therapy did not show a synergistic action on promoting functional recovery ,but do have an effect on reducing the inflammatory response and glial scar formation.

Disc-Type Hyaline Cartilage Reconstruction Using 3D-Cell Sheet Culture of Human Bone Marrow Stromal Cells and Human Costal Chondrocytes and Maintenance of Its Shape and Phenotype after Transplantation

In this study, we developed the disc-type bio-cartilage reconstruction strategies for transplantable hyaline cartilage for reconstructive surgery using 3D-cell sheet culture of human bone marrow stromal cells and human costal chondrocytes. We compared chondrogenesis efficiency between different chondrogenic-induction methods such as micromass culture, pellet culture, and 3D-cell sheet culture. Among them, the 3D-cell sheet culture resulted in the best chondrogenesis with the disc-type bio-cartilage (>12 mm diameter in size) in vitro, but sometimes spontaneous curling and contraction of 3D-cell sheet culture resulted in the formation of bead-type cartilage, which was prevented by type I collagen coating or by culturing on amniotic membrane. Previously, it was reported that tissue-engineered cartilage reconstructed in vitro does not maintain its cartilage phenotype after transplantation but tends to transform to other tissue type such as bone or connective tissue. However, the disc-type bio-cartilage of 3D-cell sheet culture maintained its hyaline cartilage phenotype even after exposure to the osteogenic-induction condition in vitro for 3 weeks or after the transplantation for 4 weeks in mouse subcutaneous. Collectively, the disc-type bio-cartilage with 12 mm diameter can be reproducibly reconstructed by the 3D-cell sheet culture, whose hyaline cartilage phenotype and shape can be maintained under the osteogenic-induction condition as well as after the transplantation. This disc-type bio-cartilage can be proposed for the application to reconstructive surgery and repair of disc-type cartilage such as mandibular cartilage and digits.

Study on cytocompatibility and animal implantation test of domestic reticulated vitreous carbon / 中华显微外科杂志

Objective To investigate the possibility of the domestic reticulated vitreous carbon as a kind of scaffold material for bone tissue engineering,the biocompatibility of domestic reticulated vitreous carbon was first successfully tested with bone marrow stromal cells (BMSCs) in vitro and for bone tissue repair in vivo.Methods From June,2013 to August,2014,the morphology and proliferation of BMSCs co-cultured with scaffold material in vitro was measured.Differences of measurement were compared with single factor analysis of variance to detect the cytotoxicity of reticulated vitreous carbon.In vivo reticulated vitreous carbon were implanted into the bone defect site and the groin.After 12 weeks,the biocompatibility of reticulated vitreous carbon was observed.Results MTT results showed that after 7d co-culture,the survival and proliferation of BMSCs had not been significantly inhibited (P ＞ 0.05).Inverted fluorescence microscope and scanning electron microscope found that newly developed three-dimensional domestic reticulated vitreous carbon could promote adhesion,aggregation and proliferation of BMSCs in vitro.Studies in vivo demonstrate that implanted reticulated vitreous carbon with a high porosity and host bone may produce a stable connection and integration.Conclusion Non-cytotoxic domestic reticulated vitreous carbon can promote the adhesion and proliferation of bone marrow mesenchymal stem cells in vitro and has good bone induction properties in vivo.

Effect of Geraniin on Expression of Wnt3a in BMSC from Osteoporotic Rats / 昆明医科大学学报

Objective To investigate the effect of geraniin on expression of Wnt3a protein and mRNA in bone marrow stromal cell (BMSC) from osteoporotic rats. Methods The model of osteoporosis (OP) was duplicated by ovariectomy in rats. BMSCs were isolated and cultured. BMSCs from shamed rats were routinly cultured and taken as normal control, and BMSCs from OP rats were divided into model group, 1μmol/L simvastatin positive group, and geraniin group (0.01, 0.1, 1, 10, 100 μmol/L), respectively. The methods of realtime-PCR and western blot were used to assay the protein and mRNA expression of wnt3a, respectively.Results As compared with normal control group, the protein and mRNA expression of wnt3a in model group were significantly suppressed;Compared with model group, 1 μmol/L simvastatin, and 0.1, 1, 10 and 100 μmol/L geraniin significantly increased the expression of wnt3a protein and mRNA. Conclusion It is suggested that geraniin activates wnt/β-catenin pathway though increasing the expression of signaling protein wnt 3a in BMSCs from OP rats. It would be beneficial to osteogenic differentiation of BMSCs and osteogenesis.