scholarly journals Effect of lentivirus-mediated growth and differentiation factor-5 transfection on differentiation of rabbit nucleus pulposus mesenchymal stem cells

2020 ◽  
Author(s):  
Kun Zhu ◽  
Rui Zhao ◽  
Yuchen Ye ◽  
Gang Xu ◽  
Changchun Zhang
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Nucleus Pulposus ◽  
Lentiviral Vector ◽  
The Other ◽  
Nucleus Pulposus Cells ◽  
Disc Disease ◽  
Qrt Pcr ◽  
Differentiation Factor ◽  
Age Related

Abstract Background：Intervertebral disc degeneration (IDD) is a natural progression of age-related process. Degenerative disc disease (DDD) is a pathologic condition associated with IDD that has been one of the most common causes of chronic low back pain, which can have a severe impact on patients’ quality of life. The purpose of this study is to observe the biological and cytological characteristics of rabbit nucleus pulposus mesenchymal stem cells (NPMSCs), and to determine the effect of growth and differentiation factor-5 (GDF-5) on the differentiation of rabbit NPMSCs transducted with lentivirus vector.Methods: In vitro culture model of rabbit NPMSCs was established and NPMSCs were identified by flow cytometry (FCM) and quantitative real-time PCR (qRT-PCR). Subsequently, NPMSCs were randomly divided into three groups: the lentiviral vector carrying GDF-5 gene used to transfect NPMSCs was recorded as transfection group; the NPMSCs transfected with an ordinary lentiviral vector was recorded as control virus group; the NPMSCs alone was normal group. FCM, qRT-PCR and Western Blot (WB) were used to detect the change of NPMSCs.Results: The transfected NPMSCs by GDF-5 gene displayed elongated shape, the cell density decreased, and the positive rate of GDF-5 in the transfected group was significantly higher than that in the other two groups (P < 0.05). The mRNA expression of KRT8, KRT18, and KRT19 in the transfected group was significantly higher in comparison with the other two groups (P < 0.05), and the result of WB was consistent with that of qRT-PCR.Conclusions: GDF-5 can induce the differentiation of NPMSCs and repair degenerative intervertebral disc. Lentiviral vector carrying GDF-5 gene can be integrated into the chromosome genome of NPMSCs and promote differentiation of NPMSCs into nucleus pulposus cells (NPCs).

scholarly journals Effect of lentivirus-mediated growth and differentiation factor-5 transfection on differentiation of rabbit nucleus pulposus mesenchymal stem cells

2022 ◽  
Vol 27 (1) ◽  
Author(s):  
Kun Zhu ◽  
Rui Zhao ◽  
Yuchen Ye ◽  
Gang Xu ◽  
Changchun Zhang
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Nucleus Pulposus ◽  
Lentiviral Vector ◽  
The Other ◽  
Cell Phenotype ◽  
Mrna Levels ◽  
Qrt Pcr ◽  
Differentiation Factor ◽  
Gdf5 Gene

Abstract Background Intervertebral disc degeneration (IDD) is a natural progression of age-related processes. Associated with IDD, degenerative disc disease (DDD) is a pathologic condition implicated as a major cause of chronic lower back pain, which can have a severe impact on the quality of life of patients. As degeneration progression is associated with elevated levels of inflammatory cytokines, enhanced aggrecan and collagen degradation, and changes in the disc cell phenotype. The purpose of this study was to investigate the biological and cytological characteristics of rabbit nucleus pulposus mesenchymal stem cells (NPMSCs)—a key factor in IDD—and to determine the effect of the growth and differentiation factor-5 (GDF5) on the differentiation of rabbit NPMSCs transduced with a lentivirus vector. Methods An in vitro culture model of rabbit NPMSCs was established and NPMSCs were identified by flow cytometry (FCM) and quantitative real-time PCR (qRT-PCR). Subsequently, NPMSCs were randomly divided into three groups: a transfection group (the lentiviral vector carrying GDF5 gene used to transfect NPMSCs); a control virus group (the NPMSCs transfected with an ordinary lentiviral vector); and a normal group (the NPMSCs alone). FCM, qRT-PCR, and western blot (WB) were used to detect the changes in NPMSCs. Results The GDF5-transfected NPMSCs displayed an elongated shape, with decreased cell density, and significantly increased GDF5 positivity rate in the transfected group compared to the other two groups (P < 0.01). The mRNA levels of Krt8, Krt18, and Krt19 in the transfected group were significantly higher in comparison with the other two groups (P < 0.01), and the WB results were consistent with that of qRT-PCR. Conclusions GDF5 could induce the differentiation of NPMSCs. The lentiviral vector carrying the GDF5 gene could be integrated into the chromosome genome of NPMSCs and promoted differentiation of NPMSCs into nucleus pulposus cells. Our findings advance the development of feasible and effective therapies for IDD.

scholarly journals Effect of Lentivirus-mediated GDF5 Transfection on Differentiation of Rabbit Nucleus Pulposus Mesenchymal Stem Cells

2020 ◽  
Author(s):  
kun zhu ◽  
Rui Zhao ◽  
Yuchen Ye ◽  
Gang Xu ◽  
Changchun Zhang
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Nucleus Pulposus ◽  
Lentiviral Vector ◽  
Intervertebral Discs ◽  
Degenerative Disease ◽  
The Other ◽  
Control Group ◽  
Nucleus Pulposus Cells ◽  
Qrt Pcr

Abstract Background: Disc degenerative disease is a common senile degenerative disease, which seriously affects the quality of life of patients.The purpose of this study is to observe the biological and cytological characteristics of rabbit nucleus pulposus mesenchymal stem cells (NPMSCs), and to determine the effect of growth differentiation factor 5(GDF5) on the differentiation of rabbit NPMSCs by lentivirus transfection.Methods: In vitro culture model of rabbit NPMSCs was established and NPMSCs cells were identified by flow cytometry (FCM)and quantitative real-time PCR(qRT-PCR). Then NPMSCs were divided into three groups: lentiviral vector carrying GDF5 was used to transfect NPMSCs, to determine the transfection rate, which was recorded as transfection group, and the NPMSCs transfected with ordinary lentiviral vector was recorded as control group, NPMSCs without processing was recorded as normal group. FCM, qRT-PCR and Western Blot(WB) were used to detected the change of NPMSCs.Results: The transfected NPMSCs by GDF5 became longer and narrower, and the cell density decreased,and the positive rate of GDF5 in the transfected group was significantly higher than that in the other two groups (P<0.05). The mRNA expression of KRT8, KRT18, KRT19 in the transfected group was significantly higher than the other two groups(P<0.05),the result of WB were the same to qRT-PCR. Conclusions: GDF5 can induce the differentiation of NPMSCs and repair degenerative intervertebral discs. Lentiviral vector carrying GDF5 can be integrated into the chromosome genome of NPMSCs and promote differentiation of NPMSCs into nucleus pulposus cells(NPCs).

scholarly journals Bi-Directional Exchange of Membrane Components Occurs during Co-Culture of Mesenchymal Stem Cells and Nucleus Pulposus Cells

PLoS ONE ◽  
2012 ◽  
Vol 7 (3) ◽  
pp. e33739 ◽  
Author(s):  
Sandra Strassburg ◽  
Nigel W. Hodson ◽  
Patrick I. Hill ◽  
Stephen M. Richardson ◽  
Judith A. Hoyland
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Nucleus Pulposus ◽  
Nucleus Pulposus Cells ◽  
Membrane Components

Transplantation of mesenchymal stem cells and nucleus pulposus cells in a degenerative disc model in rabbits: a comparison of 2 cell types as potential candidates for disc regeneration

2011 ◽  
Vol 14 (3) ◽  
pp. 322-329 ◽  
Author(s):  
Ganjun Feng ◽  
Xianfeng Zhao ◽  
Hao Liu ◽  
Huina Zhang ◽  
Xiangjun Chen ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Nucleus Pulposus ◽  
Disc Degeneration ◽  
Signal Intensity ◽  
Disc Height ◽  
Nucleus Pulposus Cells ◽  
Disc Regeneration ◽  
Disc Model

Object The aim of this study was to compare transplanted mesenchymal stem cells (MSCs) with nucleus pulposus cells (NPCs) in a degenerative disc model in rabbits to determine the better candidate for disc cell therapy. Methods Mesenchymal stem cells and NPCs were transplanted in a rabbit model of disc degeneration. Changes in disc height, according to plain radiography, T2-weighted signal intensity on MR imaging, histology, sulfated glycosaminoglycan (sGAG)/DNA, and associated gene expression levels, were evaluated among healthy controls without surgery, sham-operated animals in which only disc degeneration was induced, MSC-transplanted animals, and NPC-transplanted animals for a 16-week period. Results Sixteen weeks after cell transplantation, in the MSC- and NPC-transplanted groups, the decline in the disc height index was reduced and T2-weighted signal intensity increased compared with the sham-operated group. Safranin O staining showed a high GAG content, which was also supported by sGAG/DNA assessment. Disc regeneration was also confirmed at the gene expression level using real-time polymerase chain reaction. However, no significant differences in expression were found between the NPC- and MSC-transplanted groups. Conclusions Study data showed that MSC transplantation is effective for the treatment of disc degeneration and seems to be an ideal substitute for NPCs.

scholarly journals Age-Related Changes in Nucleus Pulposus Mesenchymal Stem Cells: An In Vitro Study in Rats

2017 ◽  
Vol 2017 ◽  
pp. 1-13 ◽  
Author(s):  
Yachao Zhao ◽  
Zhiwei Jia ◽  
Shanshan Huang ◽  
Yaohong Wu ◽  
Longgang Liu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Nucleus Pulposus ◽  
In Vitro Study ◽  
Multilineage Differentiation ◽  
Sprague Dawley ◽  
Age Related ◽  
Pathway Activation ◽  
Age Related Changes

The functions of mesenchymal stem cells (MSCs) appear to decline with age due to cellular senescence, which could reduce the efficacy of MSCs-based therapies. Recently, MSCs have been identified in the nucleus pulposus, which offers great potential for intervertebral disc (IVD) repair. However, this potential might be affected by the senescence of nucleus pulposus MSCs (NPMSCs), but whether or not this exists remains unknown. The aim of this study was to investigate the age-related changes in NPMSCs. NPMSCs isolated from young (3-month-old) and old (14-month-old) Sprague-Dawley rats were cultured in vitro. Differences in morphology, proliferation, colony formation, multilineage differentiation, cell cycle, and expression ofβ-galactosidase (SA-β-gal) and senescent markers (p53, p21, and p16) were compared between groups. Both young and old NPMSCs fulfilled the criteria for definition as MSCs. Moreover, young NPMSCs presented better proliferation, colony-forming, and multilineage differentiation capacities than old NPMSCs. Old NPMSCs displayed senescent features, including significantly increased G0/G1 phase arrest, increased SA-β-gal expression, decreased S phase entry, and significant p53-p21-pRB pathway activation. Therefore, this is the first study demonstrating that senescent NPMSCs accumulate in IVD with age. The efficacy of NPMSCs is compromised by donor age, which should be taken into consideration prior to clinical application.

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