Lentivirus-mediated TGF-β3, CTGF and TIMP1 gene transduction as a gene therapy for intervertebral disc degeneration in an in vivo rabbit model

Liu, Yong; Yu, Tao; Ma, Xuexiao; Xiang, Hongfei; Hu, Yang; Chen, Bohua

doi:10.3892/etm.2016.3063

Cited by 31 publications

(33 citation statements)

References 22 publications

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“…Cell therapy, such as mesenchymal stem cells (MSCs) in matrix constructs, has been identified as a promising therapeutic strategy to promote regeneration; however, limitations remain regarding cell retention and viability in the degenerate IVD . Pro‐anabolic gene therapy‐based approaches have been proposed including viral transfection of SOX9, which resulted in increases in matrix synthesis (collagen II) in degenerate rabbit NP . However, the use of viral vectors for gene delivery poses concerns of insertional mutagenesis and unwarranted immune responses .…”

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confidence: 99%

“…[16][17][18][19][20] Proanabolic gene therapy-based approaches have been proposed including viral transfection of SOX9, which resulted in increases in matrix synthesis (collagen II) in degenerate rabbit NP. [21][22][23] However, the use of viral vectors for gene delivery poses concerns of insertional mutagenesis and unwarranted immune responses. [24][25][26] Nonviral plasmid-based vector systems eliminate much of the risk associated with viral vectors and have been used successfully in conjunction with novel tissuenanotransfection for rescue of mice skin tissue under ischemic conditions.…”

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confidence: 99%

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Nonviral Transfection With Brachyury Reprograms Human Intervertebral Disc Cells to a Pro‐Anabolic Anti‐Catabolic/Inflammatory Phenotype: A Proof of Concept Study

Tang

Richards

Khan

et al. 2019

Journal Orthopaedic Research

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Intervertebral disc (IVD) degeneration is a major contributor to chronic low back pain and is characterized by decreases in cellularity and proteoglycan synthesis, upregulation of matrix degradation, and increases in pro‐inflammatory factors with neurovascular invasion. Current treatments fail to target the underlying pathology or promote tissue repair and approaches such as viral transfection raise safety concerns due to mutagenesis and unwarranted immune responses. To avoid such concerns, nonviral transfection is a viable method of gene delivery into the host cell while bypassing the caveats of viral delivery. Brachyury is expressed in the developing notochord and is associated with an immature healthy nucleus pulposus (NP). We hypothesize that Brachyury can reprogram degenerate NP cells to a healthy pro‐anabolic phenotype with increased proteoglycan content and decreased expression of catabolic, inflammatory, and neurovascular markers. NP cells obtained from human autopsy and surgical tissues were transfected with plasmids encoding for Brachyury or an empty vector control via bulk electroporation. Post transfection, cells were seeded in three‐dimensional agarose constructs cultured over 4 weeks and analyzed for viability, gene expression, and proteoglycan. Results demonstrated successful transfection of both autopsy and surgical NP cells. We observed long‐term Brachyury expression, significant increased expression of NP phenotypic markers FOXF1, KRT19, and chondrogenic marker SOX9 with decreases in inflammatory cytokines IL1‐β/IL6, NGF, and MMPs and significant increases in glycosaminoglycan accumulation. These results highlight nonviral transfection with developmental transcription factors, such as Brachyury, as a promising method to reprogram degenerate human disc cells toward a healthy NP phenotype. Clinical significance: This project proposes a novel translational approach for the treatment of intervertebral disc degeneration via direct reprogramming of diseased human patient‐derived IVD cells to a healthy phenotype. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:2389–2400, 2019

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confidence: 99%

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confidence: 99%

Nonviral Transfection With Brachyury Reprograms Human Intervertebral Disc Cells to a Pro‐Anabolic Anti‐Catabolic/Inflammatory Phenotype: A Proof of Concept Study

Tang

Richards

Khan

et al. 2019

Journal Orthopaedic Research

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“…Additionally, the advancements in the field of molecular biology and techniques have led to discovery of many new therapies, such as gene therapy, which has been widely applied to LDD clinical treatment. [ 19 , 20 ] The study found that compared with the control group, TSLP-positive expression was lower than that in the case group. The case group showed a higher TSLPR protein expression.…”

Section: Discussionmentioning

confidence: 99%

Role of thymic stromal lymphopoietin in the pathogenesis of lumbar disc degeneration

Wang

Yi²,

Li³

2017

Medicine

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Background:This study aims to investigate the role of thymic stromal lymphopoietin (TSLP) in the pathogenesis of lumbar disc degeneration (LDD).Methods:Nucleus pulposus tissues were collected from 77 LDD patients (the case group), in addition, normal tissues were extracted from 21 patients suffering from lumbar fractures (the control group). Immunohistochemistry was applied in order to detect TSLP positive expression. In accordance with varying transfection, the cells were divided into TSLP-siRNA, TSLP-siRNA + TSLPR-siRNA, control, blank, anti-TSLPR, and IgG groups. Western blotting was used in order to detect TSLP expression in tissues, and TSLP and type II collagen (COL2AL) in cell culture media were detected using enzyme linked immunosorbent assay (ELISA). Cell viability was measured using a MTT assay. Aggrecan levels were detected using antonopulos, and cell apoptosis was determined using flow cytometry.Results:TSLP-positive expression was found to be significantly higher in the case group compared with the control group. LDD patients’ Pfirrmann grades and preoperative visual analogue scale (VAS) scores were associated with TSLP-positive rate. Cells transfected with TSLP-siRNA and TSLPR-siRNA plasmids exhibited lower TSLP and thymic stromal lymphopoietin receptor (TSLPR) protein expression compared with the control and blank groups. Compared with the control and blank groups, there was significantly higher cell viability, lower cell apoptosis, and higher COL2AL and Aggrecan levels in the TSLP-siRNA, anti-TSLPR, and TSLP-siRNA+TSLPR-siRNA groups; there were significant differences between the TSLP-siRNA, anti-TSLPR, and TSLP-siRNA+TSLPR-siRNA groups and IgG group (all P < .05)Conclusion:Our study provides evidence for the hypothesis that TSLP could reflect the histological severity of LDD, and TSLP-siRNA and, TSLPR-siRNA could inhibit apoptosis of nucleus pulposus cells. The evident information obtained from the investigation could lead the way for new therapeutic approaches regarding LDD treatment.

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“…Although CTGF could not directly stimulate the proportion of DNA in fibroblasts, it could raise the growth factor of fibroblasts, which further strengthens the effects of CTGF and other growth factors . The study of in vitro cell culture shows that TGF‐β could promote the rapid, significant, and lasting increase in CTGF …”

Section: Discussionmentioning

confidence: 99%

The research of transgenic human nucleus pulposus cell transplantation in the treatment of lumbar disc degeneration

Wang

Jin

Kong

et al. 2019

The Kaohsiung J of Med Scie

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The present study determines whether the in vivo injection of TGFβ1 and CTGF mediated by AAV2 to transfect nucleus pulposus cells in degenerative lumbar discs can reverse the biological effects of rhesus lumbar disc degeneration. A total of 42 lumbar discs obtained from six rhesus monkeys were classified into three groups: experimental group, control group, and blank group. Degenerative lumbar discs were respectively injected with double gene-transfected human nucleus pulposus cells using minimally invasive techniques. Immumohistochemical staining, RT-PCR, and western blot were performed to observe the biological effects of double genetransfected human nucleus pulposus cells in degenerative lumbar discs on rhesus lumbar disc degeneration. At 4, 8, and 12 weeks after the transplantation of nucleus pulposus cells, the expression levels of TGF-ß1, CTGF, proteoglycan mRNA, and type-II collagen were detected by RT-PCR. The values of immumohistochemical staining and RT-PCR in the experimental group increased at 8 weeks, decreased with time at 12 weeks, and remained greater than the values in the control group, and the differences were statistically significant (P < .05). The western blot revealed that the values in the experimental group decreased with time, but remained greater than those in the PBS control group and blank control group, and the differences were statistically significant (P < .05). The double gene-transfection of human nucleus pulposus cells in degenerative lumbar discs mediated by rAAV2 can be continuously expressed in vivo after transplantation in lumbar discs of rhesus monkeys, and promotes the synthesis of proteoglycan and type II collagen, achieving the treatment purpose. K E Y W O R D S

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Lentivirus-mediated TGF-β3, CTGF and TIMP1 gene transduction as a gene therapy for intervertebral disc degeneration in an in vivo rabbit model

Cited by 31 publications

References 22 publications

Nonviral Transfection With Brachyury Reprograms Human Intervertebral Disc Cells to a Pro‐Anabolic Anti‐Catabolic/Inflammatory Phenotype: A Proof of Concept Study

Nonviral Transfection With Brachyury Reprograms Human Intervertebral Disc Cells to a Pro‐Anabolic Anti‐Catabolic/Inflammatory Phenotype: A Proof of Concept Study

Role of thymic stromal lymphopoietin in the pathogenesis of lumbar disc degeneration

The research of transgenic human nucleus pulposus cell transplantation in the treatment of lumbar disc degeneration

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