Transfected Bone Marrow Mesenchymal Stem Research Articles

Exosomes Derived from miR‐195‐5p Transfected Bone Marrow Mesenchymal Stem Cells Alleviate Pelvic Floor Dysfunction in Rats

AbstractBone marrow‐derived mesenchymal stem cells (BMSCs) can alleviate pelvic floor dysfunction (PFD) in rats. Whether exosomes derived from miR‐195‐5p transfected BMSCs can alleviate PFD is investigated. The rat vaginal distention model is constructed to mimic PFD. miR‐195‐5p is transfected into BMSCs, and exosomes derived from miR‐195‐5p transfected BMSCs are injected into PFD rats via caudal veins. Luciferase reporter assay is performed to confirm the direct interaction between SMAD7 3′‐UTR and miR‐195‐5p mimics. Void volume, bladder void pressure, leak point pressure, and peak bladder pressure are used to evaluate urodynamic function. It is found that miR‐195‐5p directly targets Smad7 and regulates the relative Smad7 expression in rat BMSCs. Exosomes derived from miR‐195‐5p transfected BMSCs can improve urodynamic function than exosomes derived from BMSCs. On the other hand, exosomes derived from miR‐195‐5p transfected BMSCs inhibit the expressions of Smad7 but promote the expressions of elastin in PFD rats with diminished expression of MMP2 and MMP9. This study demonstrates that exosomes derived from miR‐195‐5p genetically engineered BMSCs can target the Tgf‐β/Smad7 pathway to repair connective tissues and restore damaged urodynamic function in PFD rats.

Retraction Note to: Effects of BDNF-Transfected BMSCs on Neural Functional Recovery and Synaptophysin Expression in Rats with Cerebral Infarction.

The purpose of this study was to investigate the effects of brain-derived neurotrophic factor (BDNF)-transfected bone marrow mesenchymal stem cells (BMSCs) on neural functional recovery and synaptophysin expression in rats with cerebral infarction (CI). A total of 120 healthy Sprague Dawley rats were randomly divided into sham group, control group, and model group. Craniotomy was conducted and neurological function defect scoring was used to verify the model. BDNF containing recombinant plasmid was transfected into rat BMSCs, which was verified by flow cytometry and Western Blot. After injection of the transfected BMSCs, neural functional recovery of the CI rats and synaptophysin expression were measured. After the CI rat model was established, magnetic resonance (MR) imaging, 2, 3, 5- triphenyl tetrazolium chloride (TTC) staining, and the neurological function defect scoring determined the success of the model. CD34 (-), CD45 (−), CD29 (+), and CD90 (+) cells detected showed that the obtained BMSCs have high purity. BDNF protein was highly expressed in the BMSCs successfully transfected with the recombinant plasmid. Balance beam walking score, rotating bar walking score, and screen test score were significantly lower, while synaptophysin expression was higher in the BDNF model group than those in the non-BDNF model group and sham group with time extension. BDNF can increase synaptic plasticity and neurogenesis and have a promotional role in neural functional recovery and synaptophysin expression in rats with CI. BDNF-transfected BMSCs may therefore have better treatment efficacy for CI clinically.

Vascular Endothelial Growth Factor-Transfected Bone Marrow Mesenchymal Stem Cells Improve the Recovery of Motor and Sensory Functions of Rats With Spinal Cord Injury.

Basic science. The aim of this study was to examine the effect of vascular endothelial growth factor (VEGF)-transfected bone marrow mesenchymal stem cells (BMSCs) on the recovery of motor and sensory functions of rats with spinal cord injury (SCI). There is no effective treatment to protect against SCI. BMSCs have been widely applied to the treatment of nervous system damage due to the function of prompt neurite growth and inhibition of demyelination following injury. VEGF-transfected BMSCs were injected to rats with SCI and the recovery of motor and sensory functions was observed. The Basso, Beattie, and Bresnahan, mechanical withdrawal threshold and thermal withdraw latency grading was conducted to assess the recovery status of motor and sensory functions of the SCI rats. The expression of VEGF, CD31, and NF200 was detected by immunofluorescence. The recovery of the rat motor and sensory functions in the VEGF-transfected BMSC (BMSC-VEGF) group was higher than those of the other groups with the exception of the Sham group (P < 0.05). The expression of the CD31 and NF200 proteins in the rat SCI regions was the highest in the BMSC-VEGF group, whereas the survival of BMSC in the BMSC-VEGF group was increased compared with that in the BMSC-Ad group. In addition, the injection of VEGF-transfected BMSCs can improve the angiogenesis of the injured area and retain the survival of injected cells and neurons. The injection of BMSC-VEGF improved the recovery of motor function in SCI rats. N/A.

Role of FGF-2 Transfected Bone Marrow Mesenchymal Stem Cells in Engineered Bone Tissue for Repair of Avascular Necrosis of Femoral Head in Rabbits

Background/Aims: Avascular necrosis of the femoral head (ANFH) is the focus and difficulty of orthopedic diseases. Recently, tissue engineering bone for this disease has shown a good therapeutic effect. The aim of the present study was to investigate the therapeutic effect of basic fibroblast growth factor (FGF-2) as cytokines transfected bone marrow mesenchymal stem cells (BMSCs) in constructing tissue-engineered bone for avascular necrosis of the femoral head. Methods: The FGF-2 gene overexpressed lentivirus-transfected rBMSCs with xenogeneic antigen-extracted cancellous bone (XACB) to construct tissue engineered bone, and the model of early avascular necrosis of the femoral head was established by lipopolysaccharide (LPS) combined with hormone. The models were randomly divided into five groups: A (control), B (XACB), C (XACB+rBMSCs), D (XACB+rBMSCs+Lv-GFP), and E (XACB+rBMSCs+Lv-FGF-2/GFP) groups. The therapeutic effect of the tissue engineered bone for the avascular necrosis of the femoral head was evaluated by gross anatomy, X-ray examination, immunohistochemistry and H&E staining. Results: The FGF-2 gene was transfected into rBMSCs (Multiplicity of infection [MOI] = 100) by lentivirus, and its efficiency was above 95%. The rBMSCs were successfully transfected overexpressing FGF-2 by qPCR and western blot. After tissue engineering bone implantation, X-ray examination and gross specimen observation revealed that the repair area in the E group was > 80% at six weeks, the defect was completely repaired at 12 weeks, and osteogenesis was stronger, when compared with the other groups. For the X-ray score, vascular area density and new bone formation area were higher, when compared with the other groups, and the difference was statistically significant (P< 0.05). Conclusion: FGF-2 gene overexpression lentivirus transfection BMSCs combined with XACB to construct tissue engineered bone can effectively promote vascular regeneration, and improve the repair effect of avascular necrosis of the femoral head.

In Vitro Evaluation of the Nano-Hydroxyapatite/Hydrogel Compound and Its Combination with BMP7 Transfected Bone Marrow Mesenchymal Stem Cells

<i>In Vitro</i> Evaluation of the Nano-Hydroxyapatite/Hydrogel Compound and Its Combination with BMP7 Transfected Bone Marrow Mesenchymal Stem Cells

Evaluation of a novel poly(amidoamine) with pendant aminobutyl group on the cellular properties of transfected bone marrow mesenchymal stem cells.

Stem cell-based gene therapy has been considered in the treatment of many degenerative diseases. Gene-modified stem cells should maintain its reproductive activity without losing stem cell properties, including genetic phenotype and differentiation potential. In the study, a novel poly (amidoamine) with pendant aminobutyl group (PAA-BA) designed by our group was used in the transfection of bone marrow mesenchymal stromal cells (BMSCs) and the cellular properties post-transfection were evaluated, including DNA content, colony forming capacity, genetic phenotype, and multi-directional differentiation. Two classical non-viral gene delivery vectors, polyethylenimine (PEI) and Lipofectamine 2000 (LP2000) were also used. Compared to non-transfected group, PAA-BA showed minor decreased DNA content but maintained BMSCs' phenotype, reproductive activity and multi-differentiation potential (osteogenic, chondrogenic, adipogenic, and neurogenic differentiation). Both PAA-BA and PEI transfected BMSCs demonstrated improved osteogenic differentiation ability at late stage but suppressed adipogenic as well as mature neural differentiation in vitro. LP2000 and PEI transfected BMSCs displayed significantly lower DNA content and reproductive activity. These findings suggest that PAA-BA is one of safe gene delivery vectors in BMSCs transfection and plays a role in stem cell's osteogenic and neurogenic differentiation. This study proposes the potential application of PAA-BA in BMSCs based gene therapy, in particular bone and nerve relative diseases. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 686-697, 2018.

Repairing Effect of a Nano-Artificial Bone Combined with HIF-1&lt;I&gt;α&lt;/I&gt; Transfected Bone Marrow Mesenchymal Stem Cells for Rabbit Radial Bone Defects

Repairing Effect of a Nano-Artificial Bone Combined with HIF-1&lt;I&gt;α&lt;/I&gt; Transfected Bone Marrow Mesenchymal Stem Cells for Rabbit Radial Bone Defects

Effects of BDNF-Transfected BMSCs on Neural Functional Recovery and Synaptophysin Expression in Rats with Cerebral Infarction.

The purpose of this study was to investigate the effects of brain-derived neurotrophic factor (BDNF)-transfected bone marrow mesenchymal stem cells (BMSCs) on neural functional recovery and synaptophysin expression in rats with cerebral infarction (CI). A total of 120 healthy Sprague Dawley rats were randomly divided into sham group, control group, and model group. Craniotomy was conducted and neurological function defect scoring was used to verify the model. BDNF containing recombinant plasmid was transfected into rat BMSCs, which was verified by flow cytometry and Western Blot. After injection of the transfected BMSCs, neural functional recovery of the CI rats and synaptophysin expression were measured. After the CI rat model was established, magnetic resonance (MR) imaging, 2, 3, 5- triphenyl tetrazolium chloride (TTC) staining, and the neurological function defect scoring determined the success of the model. CD34 (-), CD45 (-), CD29 (+), and CD90 (+) cells detected showed that the obtained BMSCs have high purity. BDNF protein was highly expressed in the BMSCs successfully transfected with the recombinant plasmid. Balance beam walking score, rotating bar walking score, and screen test score were significantly lower, while synaptophysin expression was higher in the BDNF model group than those in the non-BDNF model group and sham group with time extension. BDNF can increase synaptic plasticity and neurogenesis and have a promotional role in neural functional recovery and synaptophysin expression in rats with CI. BDNF-transfected BMSCs may therefore have better treatment efficacy for CI clinically.

Combining acellular nerve allografts with brain-derived neurotrophic factor transfected bone marrow mesenchymal stem cells restores sciatic nerve injury better than either intervention alone

In this study, we chemically extracted acellular nerve allografts from bilateral sciatic nerves, and repaired 10-mm sciatic nerve defects in rats using these grafts and brain-derived neurotrophic factor transfected bone marrow mesenchymal stem cells. Experiments were performed in three groups: the acellular nerve allograft bridging group, acellular nerve allograft + bone marrow mesenchymal stem cells group, and the acellular nerve allograft + brain-derived neurotrophic factor transfected bone marrow mesenchymal stem cells group. Results showed that at 8 weeks after bridging, sciatic functional index, triceps wet weight recovery rate, myelin thickness, and number of myelinated nerve fibers were significantly changed in the three groups. Variations were the largest in the acellular nerve allograft + brain-derived neurotrophic factor transfected bone marrow mesenchymal stem cells group compared with the other two groups. Experimental findings suggest that chemically extracted acellular nerve allograft combined nerve factor and mesenchymal stem cells can promote the restoration of sciatic nerve defects. The repair effect seen is better than the single application of acellular nerve allograft or acellular nerve allograft combined mesenchymal stem cell transplantation.

Transplantation KCNMA1 modified bone marrow-mesenchymal stem cell therapy for diabetes mellitus-induced erectile dysfunction

This study assessed the effect of KCNMA1 transfected bone marrow-mesenchymal stem cells (BM-MSCs) on the improvement of erectile function in diabetic rats. Sixty male Sprague-Dawley rats were injected with streptozotocin (STZ) and screened with apomorphine (APO) to establish diabetes mellitus-induced erectile dysfunction (DMED). DMED rats were randomly divided into four groups: rats in each group underwent intracavernous injection with either phosphate buffer solution (DMED+PBS), nontransfected MSCs (DMED+MSCs), empty vector transfected MSCs (DMED+null-MSCs) or KCNMA1 transfected MSCs (DMED+KCNMA1-MSCs). Before injection, high levels of KCNMA1 expression were confirmed in KCNMA1-MSCs using RT-PCR and Western blotting. The lentivirus transfected MSCs maintained their potential for multidirectional differentiation. Four weeks after injection, erectile function was ascertained by measuring intracavernous pressure (ICP). Penile tissues were collected for immunohistochemical analysis. The expression of KCNMA1 in the corpus cavernosum was increased, and the DMED+KCNMA1-MSCs group displayed a significant improvement of erectile function. We concluded that KCNMA1 was able to enhance the positive effect of MSCs in the treatment of diabetes-associated erectile dysfunction.

LIM mineralization protein 1 gene transfected bone marrow mesenchymal stem cells combined with nano-hydroxyapatite/polyamide 66 plate in the treatment of canine femoral fractures

LIM mineralization protein 1 gene transfected bone marrow mesenchymal stem cells combined with nano-hydroxyapatite/polyamide 66 plate in the treatment of canine femoral fractures

Experiment study of rabbit joint articular cartilage defects repaired by thermosensitive CS/PVA composite hydrogel engineered hTGF-β1 transfected bone marrow mesenchymal stem cells

Objective To investigate the experiment effects of rabbit joint articular cartilage defects repaired by thermosensitive CS/PVA composite hydrogel engineered hTGF-β1 transfected bone marrow mesenchymal stem cells. Methods Bone marrow mesenchymal stem cells were isolated and cultured in vitro. The positive rate of transfection was defected by cell immunohistochemistry methods after Ad-hTGF-β1 transfected for 1 week. Twenty-four adult New Zealand white rabbits with full articular cartilage defects were randomly divided into 4 groups, each group had 6 animals, both hind limbs were used in the experiment. Group A: hydrogel combined with transfected cells; Group B: hydrogel combined with untransfected cells; Group C: hydrogel group; Group D: blank control group. Specimens and histological observation were used to evaluate the repair effect after 16 weeks according to Pineda's score. Results The positive rate of hTGF-β1 expression in BMSCs was about 85.4% after transfection. After 16 weeks the defects of group A were repaired by cartilage-like tissue, the cell arrangement and densities of regenerated cartilage were similar to normal cartilage, type Ⅱ collagen immunohistochemistry were positive. There was a significant difference in Pineda's score compaired with other groups (P ＜ 0.05). Conclusion Rabbit articiular cartilage defects could be repaired by CS/PVA hydrogel engineered hTGF-β1-transfected bone marrow mesenchymal stem cells. Key words: Hydrogel; Transforming growth factor; Bone marrow mesenchymal stem cells; Joint articular cartilage defect

Recombinant hTGF-β1 adenovirus transfect bone marrow mesenchymal stem cells and the change of TAZ mRNA expression

Objective To investigate the feasibility of Ad-hTGF-β1 transfected bone marrow mesenchymal stem cell(BMSCs) into chondrocytes differentiation and the change of TAZ mRNA. Methods Rats BMSCs were obtained and cultured by whole bone marrow method, and then the third-generation cells were seeded into cell culture plate, and divided into three groups:Ad-hTGF-β1 transfected group,Ad-EGFP transfected group and the control group. The control group was added in common medium without any treatment while the other two groups were respectively added in serum-free medium containing Ad-hTGF-β1 or that containing Ad-EGFP. Seven days later, real-time fluorescent quantitation PCR and Western blot were employed for detecting the expression of TGF-β1 ,while immunohistochemical and Western blot for the expression of collagen Ⅱ , and real-time fluorescent quantitation PCR for the expression of TAZ mRNA. Results Seven days after the transfection, real-time fluorescent quantitation PCR revealed that the average relative expression of TGF-β1 was:Ad-hTGF-β1 group 0.863, Ad-EGFP group 0.183, and the control group 0.180; The average relative expression of TAZ was:Ad-hTGF-β1 group 0.810, Ad-EGFP group 0.416, and the control group 0.366.The expression difference of TGF-β1 and TAZ were statistically significant (P ＜ 0.05). Western blot and immunohistochemical proved strong collagen Ⅱ expression in Ad-hTGF-β1 group while it was detected a little in the other two groups. Conclusion BMSCs could be successfully and stably induced into chondrocytes differentiation by Ad-hTGF-β1. Meanwhile, the mRNA of TAZ is up regulate during the differentiation,so it is suppose that TGF-β1 improve BMSCs into chondrocytes differentiation by TAZ. Key words: Bone marrow mesenhymal stem cell; Chondrocytes; Gene transfection; TGF-β1

An experimental study on rat bone marrow mesenchymal stem cells transfected with dentin sialophosphoprotein gene in vitro

To evaluate the expression of dentin sialophosphoprotein (DSPP) in transfected rat bone marrow mesenchymal stem cells (BM-MSC) and the influence of the transfection. Plasmid containing mice dentin DSPP was constructed by using the cytomegalovirus (CMV) promoter and then transfected the cultured BM-MSC by lipofectamine; The expression of Pax-9 and dentin matrix protein 1 (DMP1) gene of transfected BM-MSC were detected by RT-PCR. The expression of DSPP was examined by immunocytochemical staining, and the formation ratio of mineralized nodules of transfected BM-MSC was compared with untransfected ones after mineralized induction. The constructed pcDNA3.1(+)/mDSPP could produced 3.0 kb and 5.4 kb fragments, DSPP gene and Pax9 gene were expressed 24 h and 48 h respectively, after BM-MSC were transfected Pax-9 gene was exprssed, but DMP1 gene was not; Immunohistochemical staining showed that DSPP was positive in transfected BM-MSC; The formation ratio of mineralized nodules of transfected BM-MSC was higher than that of untransfected ones after mineralized induction. The expression of mice DSPP in BM-MSC by gene transfection can induce the expression of tooth development-associated gene Pax9 and enhance the formation of mineralized nodules, which suggests that DSPP gene might induce odontogenic differentiation of BM-MSC.