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Treating human meniscal fibrochondrocytes with hIGF-1 gene by liposome.

ABSTRACT: The menisci are intraarticular fibrocartilaginous structures essential to the normal function of the knee that lack the ability to self-repair. Human meniscal fibrochondrocytes may respond to beneficial genes like human insulin growth factor-1 (hIGF-1) and the meniscal cell may be a feasible donor for gene therapy. To explore this possibility, we amplified the hIGF-1 gene sequence in full length and cloned it into a bicistronic plasmid. This gene was then transfected into cultured human meniscal fibrochondrocytes by the liposome FuGene 6. Green fluorescence was expressed in part of the cells 6 hours after transfection and increased gradually, with a peak concentration of the hIGF-1 in the supernatants to 22.68 ng/mL 56 hours after transfection. Phenotypes of some cells changed and the proliferation accelerated after transfection. Flow cytometry analysis demonstrated upregulation of cell numbers in the G2 and S stages after hIGF-1 gene introduction. We conclude the hIGF-1 gene can be transfected into the human meniscal cell efficiently by liposome and it causes accelerated proliferation and differentiation. Within 10 days after transfection, the cytokine appears to be secreted into supernatants with the bioactivity and promotes the proliferation of the NIH 3T3 cell line.

SUBMITTER: Zhang HN

PROVIDER: S-EPMC2772929 | biostudies-literature | 2009 Dec

REPOSITORIES: biostudies-literature

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Treating human meniscal fibrochondrocytes with hIGF-1 gene by liposome.

Zhang Hai-ning HN Leng Ping P Wang Ying-zhen YZ Zhang Jie J

Clinical orthopaedics and related research 20090508 12

The menisci are intraarticular fibrocartilaginous structures essential to the normal function of the knee that lack the ability to self-repair. Human meniscal fibrochondrocytes may respond to beneficial genes like human insulin growth factor-1 (hIGF-1) and the meniscal cell may be a feasible donor for gene therapy. To explore this possibility, we amplified the hIGF-1 gene sequence in full length and cloned it into a bicistronic plasmid. This gene was then transfected into cultured human meniscal ...[more]

PMID: 19424673

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Project description:Menisci play a vital role in load transmission, shock absorption and joint stability. The current dogma is that the menisci simply protects the cartilage and play no role in osteoarthritis (OA) unless they are injured. However, there is increasing evidence suggesting that OA menisci may not merely be bystanders in the disease process of OA. This study sought: 1) to determine the prevalence of meniscal degeneration in OA patients, 2) to examine gene expression in OA meniscal cells compared to normal control meniscal cells, and 3) to test the hypothesis that OA meniscal cells are different from normal meniscal cells. The grades of meniscal degeneration correlated in a positive fashion with the grades of articular cartilage degeneration (r = 0.672; P < 0.0001). Many genes classified in the biological processes of immune response, inflammatory response, biomineral formation and cell proliferation, including major histocompatibility complex, class II, DP alpha 1 (HLA-DPA1), integrin, beta 2 (ITGB2), ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1), ankylosis, progressive homolog (ANKH) and fibroblast growth factor 7 (FGF7), were expressed at higher levels compared to normal control meniscal cells. In addition, many genes that were previously implicated in OA were also expressed at higher levels in OA meniscal cells, including ADAM metallopeptidase with thrombospondin type 1 motif, 5 (ADAMTS5). Our findings suggest that OA is a whole joint disease. Meniscal cells may play an active role in the development of OA. Investigation of the gene expression profiles of OA meniscal cells may reveal new therapeutic targets for OA therapy and also may uncover novel disease markers for early diagnosis of OA. Studies were approved by our human subjects Institutional Review Board. Menisci and articular cartilage were collected during joint replacement surgery for OA patients (OA specimens) and lower limb amputation surgery for osteosarcoma patients (normal control specimens), and graded. Meniscal cells were prepared from the meniscal tissues and expanded in monolayer culture. Differential gene expression in OA meniscal cells and normal control meniscal cells was examined using Affymetrix microarray and real time RT-PCR.

Project description:BackgroundMenisci play a vital role in load transmission, shock absorption and joint stability. There is increasing evidence suggesting that OA menisci may not merely be bystanders in the disease process of OA. This study sought: 1) to determine the prevalence of meniscal degeneration in OA patients, and 2) to examine gene expression in OA meniscal cells compared to normal meniscal cells.MethodsStudies were approved by our human subjects Institutional Review Board. Menisci and articular cartilage were collected during joint replacement surgery for OA patients and lower limb amputation surgery for osteosarcoma patients (normal control specimens), and graded. Meniscal cells were prepared from these meniscal tissues and expanded in monolayer culture. Differential gene expression in OA meniscal cells and normal meniscal cells was examined using Affymetrix microarray and real time RT-PCR.ResultsThe grades of meniscal degeneration correlated with the grades of articular cartilage degeneration (r = 0.672; P < 0.0001). Many of the genes classified in the biological processes of immune response, inflammatory response, biomineral formation and cell proliferation, including major histocompatibility complex, class II, DP alpha 1 (HLA-DPA1), integrin, beta 2 (ITGB2), ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1), ankylosis, progressive homolog (ANKH) and fibroblast growth factor 7 (FGF7), were expressed at significantly higher levels in OA meniscal cells compared to normal meniscal cells. Importantly, many of the genes that have been shown to be differentially expressed in other OA cell types/tissues, including ADAM metallopeptidase with thrombospondin type 1 motif 5 (ADAMTS5) and prostaglandin E synthase (PTGES), were found to be expressed at significantly higher levels in OA meniscal cells. This consistency suggests that many of the genes detected in our study are disease-specific.ConclusionOur findings suggest that OA is a whole joint disease. Meniscal cells may play an active role in the development of OA. Investigation of the gene expression profiles of OA meniscal cells may reveal new therapeutic targets for OA therapy and also may uncover novel disease markers for early diagnosis of OA.

Project description:BACKGROUND:Meniscal tears are traditionally classified into traumatic versus degenerative tears. Although this classification plays a major role in clinical decision making, no consensus exists on the exact definition of a traumatic or degenerative tear, and the histopathological basis for this classification is unclear. PURPOSE:To assess the histological degree of meniscal degeneration in patients with a traumatic meniscal tear, as compared with intact meniscal tissue and osteoarthritic meniscal tissue. STUDY DESIGN:Descriptive laboratory study. METHODS:Traumatically torn meniscal tissue was collected during arthroscopic partial meniscectomy. As a control group, intact meniscal tissue was used from transfemoral amputations or direct postmortem dissections. Meniscal tissue from osteoarthritic knees was obtained during total knee replacement surgery. Meniscal tissue was processed, stained, and histologically analyzed with the Pauli scoring system (range, 0-18), comprising the subdomains surface integrity, cellularity, collagen organization, and matrix staining. Scoring was performed by 2 independent observers, blinded to condition, region, and patient data of the meniscus. RESULTS:The traumatic meniscal tear group contained 43 patients (34 men; median age, 29 years; median body mass index [BMI], 24 kg/m2); the intact meniscal tissue group, 8 patients (3 men; median age, 58 years; median BMI, 30 kg/m2); and the osteoarthritic group, 14 patients (4 men; median age, 66 years; median BMI, 28 kg/m2). After adjustment for sex, age, and BMI, patients with a traumatic meniscal tear had a significantly higher histological score than patients with intact meniscal tissue (2.7-point difference; P = .035). Histological score between the traumatic and osteoarthritic groups was not different. CONCLUSION:Traumatically torn menisci possess a higher degree of degeneration than intact menisci. Our results suggest that patients with a traumatic meniscal tear may already have had a certain degree of meniscal degeneration. These findings potentially challenge the classic view of traumatic versus degenerative meniscal tears. CLINICAL RELEVANCE:Our findings provide a better understanding of the tissue condition of a torn meniscus. This knowledge may help clinicians decide on choice of treatment and may lead to new perspectives to prevent knee osteoarthritis in patients with a torn meniscus.

Dataset Information

Treating human meniscal fibrochondrocytes with hIGF-1 gene by liposome.

Publications

Treating human meniscal fibrochondrocytes with hIGF-1 gene by liposome.

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