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Fibroblast growth factor 18 increases the trophic effects of bone marrow mesenchymal stem cells on chondrocytes isolated from late stage osteoarthritic patients.

Zhang Z, Wang Y, Li M, Li J, Wu J - Stem Cells Int (2014)

Bottom Line: Finally, coimplantation of MSCs with OA chondrocytes produces more matrix than chondrocytes only.In conclusion, FGF18 can restore the responsiveness of OA chondrocytes to the trophic effects of MSCs.Coimplantation of MSCs and OA chondrocytes treated with FGF18 may be a good alternative cell source for regenerating cartilage tissue that is degraded during OA pathological changes.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthopedic Surgery, The First Affiliated Hospital of Harbin Medical University, 23 You Zheng Street, Nangang District, Harbin 150001, China.

ABSTRACT
Coculture of mesenchymal stem cells with chondrocytes increases production of cartilaginous matrix. Chondrocytes isolated from late stage osteoarthritic patients usually lost their phenotype of producing cartilaginous matrix. Fibroblast growth factor 18 is believed to redifferentiate OA chondrocyte into functionally active chondrocytes. The aim of this study is to investigate the supportive effects of MSCs on OA chondrocytes and test if FGF18 could enhance the responsiveness of OA chondrocytes to the support of MSCs in a coculture system. Both pellet and transwell co-cultures were used. GAG quantification, hydroxyproline assay, and qPCR were performed. An ectopic models of cartilage formation was also applied. Our data indicated that, in pellets coculture of MSCs and OA chondrocytes, matrix production was increased in the presence of FGF18, comparing to the monoculture of chondrocytes. Results from transwell coculture study showed that expression of matrix producing genes in OA chondrocytes increased when cocultured with MSCs with FGF18 in culture medium, while hypertrophic genes were not changed by coculture. Finally, coimplantation of MSCs with OA chondrocytes produces more matrix than chondrocytes only. In conclusion, FGF18 can restore the responsiveness of OA chondrocytes to the trophic effects of MSCs. Coimplantation of MSCs and OA chondrocytes treated with FGF18 may be a good alternative cell source for regenerating cartilage tissue that is degraded during OA pathological changes.

No MeSH data available.


Related in: MedlinePlus

Coculture of MSCs and OA chondrocytes increases GAG formation and collagen biosynthesis with the presence of FGF18. (a) Toluidine blue staining was performed at week 4 after aggregation that was cultured in medium without FGF18 to detect GAGs. Scale bar = 100 μm. (b) Toluidine blue staining was performed at week 4 after aggregation that was cultured in medium containing FGF18 to detect GAGs. Scale bar = 100 μm. (c) GAG assay was performed to quantify GAG deposited in cell pellets (n = 3) at week 4 after aggregation. Total GAGs (μg) were normalized to total DNA (μg). Error bar reflects standard deviation. P values were calculated with Student's t-test. (d) Hydroxyproline assay was carried out to measure total collagen contents (n = 3). Total collagen (μg) was normalized to total DNA (μg). Error bar reflects standard deviation. P values were calculated with Student's t-test. (e) Deposition of COL II in pellets was measured by ELISA (n = 3). COL II contents were normalized to total DNA. P values were calculated with Student's t-test.
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fig1: Coculture of MSCs and OA chondrocytes increases GAG formation and collagen biosynthesis with the presence of FGF18. (a) Toluidine blue staining was performed at week 4 after aggregation that was cultured in medium without FGF18 to detect GAGs. Scale bar = 100 μm. (b) Toluidine blue staining was performed at week 4 after aggregation that was cultured in medium containing FGF18 to detect GAGs. Scale bar = 100 μm. (c) GAG assay was performed to quantify GAG deposited in cell pellets (n = 3) at week 4 after aggregation. Total GAGs (μg) were normalized to total DNA (μg). Error bar reflects standard deviation. P values were calculated with Student's t-test. (d) Hydroxyproline assay was carried out to measure total collagen contents (n = 3). Total collagen (μg) was normalized to total DNA (μg). Error bar reflects standard deviation. P values were calculated with Student's t-test. (e) Deposition of COL II in pellets was measured by ELISA (n = 3). COL II contents were normalized to total DNA. P values were calculated with Student's t-test.

Mentions: Four weeks after aggregation, cell pellets of monoculture and coculture were harvested for histological examination, GAG assay, and collagen quantification. As shown in Figure 1(a), not much GAG was deposited into extracellular matrix in pellets cultured in serum-free medium without FGF18. This is not surprising because chondrocytes isolated from end stage OA patients already lost their ability to produce cartilaginous matrix. With or without the supports from MSCs, they cannot lay down matrix rich in GAG and collagen. Upon stimulation of FGF18, these OA chondrocytes quickly regain their machinery for matrix synthesis and deposit abundant GAG in the pellets (Figure 1(b)). Building on this redifferentiation process, MSCs could further promote GAG synthesis of chondrocytes due to their trophic effects. Quantitative measurements for GAG and total collagen confirmed our impression on histological staining (Figures 1(c) and 1(d)). Without FGF18 in the medium, 1 μg of DNA is roughly corresponding to 2 μg of GAG and 1 μg hydroxyproline. And there is no difference between monoculture and coculture pellets. Once FGF18 is added to the medium, three- to fourfold increases were seen in GAG and collagen quantification. More importantly, GAG produced in coculture pellets was significantly more than that in monoculture pellets. The same trend was observed in collagen quantification. Since hydroxyproline assay cannot distinguish cartilage specific collagen type II (COL II) from other collagens, ELISA was performed to quantify the amount of COL II in cell pellets. Upon stimulation of FGF18, monoculture and coculture pellets deposit 3-4 times more COL II in matrix. As expected, coculture pellets contain significantly more COL II than monoculture pellets (Figure 1(e)). This indicated that trophic effects of MSCs may increase matrix formation of OA chondrocytes with the redifferentiation of chondrocytes stimulated by FGF1.


Fibroblast growth factor 18 increases the trophic effects of bone marrow mesenchymal stem cells on chondrocytes isolated from late stage osteoarthritic patients.

Zhang Z, Wang Y, Li M, Li J, Wu J - Stem Cells Int (2014)

Coculture of MSCs and OA chondrocytes increases GAG formation and collagen biosynthesis with the presence of FGF18. (a) Toluidine blue staining was performed at week 4 after aggregation that was cultured in medium without FGF18 to detect GAGs. Scale bar = 100 μm. (b) Toluidine blue staining was performed at week 4 after aggregation that was cultured in medium containing FGF18 to detect GAGs. Scale bar = 100 μm. (c) GAG assay was performed to quantify GAG deposited in cell pellets (n = 3) at week 4 after aggregation. Total GAGs (μg) were normalized to total DNA (μg). Error bar reflects standard deviation. P values were calculated with Student's t-test. (d) Hydroxyproline assay was carried out to measure total collagen contents (n = 3). Total collagen (μg) was normalized to total DNA (μg). Error bar reflects standard deviation. P values were calculated with Student's t-test. (e) Deposition of COL II in pellets was measured by ELISA (n = 3). COL II contents were normalized to total DNA. P values were calculated with Student's t-test.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig1: Coculture of MSCs and OA chondrocytes increases GAG formation and collagen biosynthesis with the presence of FGF18. (a) Toluidine blue staining was performed at week 4 after aggregation that was cultured in medium without FGF18 to detect GAGs. Scale bar = 100 μm. (b) Toluidine blue staining was performed at week 4 after aggregation that was cultured in medium containing FGF18 to detect GAGs. Scale bar = 100 μm. (c) GAG assay was performed to quantify GAG deposited in cell pellets (n = 3) at week 4 after aggregation. Total GAGs (μg) were normalized to total DNA (μg). Error bar reflects standard deviation. P values were calculated with Student's t-test. (d) Hydroxyproline assay was carried out to measure total collagen contents (n = 3). Total collagen (μg) was normalized to total DNA (μg). Error bar reflects standard deviation. P values were calculated with Student's t-test. (e) Deposition of COL II in pellets was measured by ELISA (n = 3). COL II contents were normalized to total DNA. P values were calculated with Student's t-test.
Mentions: Four weeks after aggregation, cell pellets of monoculture and coculture were harvested for histological examination, GAG assay, and collagen quantification. As shown in Figure 1(a), not much GAG was deposited into extracellular matrix in pellets cultured in serum-free medium without FGF18. This is not surprising because chondrocytes isolated from end stage OA patients already lost their ability to produce cartilaginous matrix. With or without the supports from MSCs, they cannot lay down matrix rich in GAG and collagen. Upon stimulation of FGF18, these OA chondrocytes quickly regain their machinery for matrix synthesis and deposit abundant GAG in the pellets (Figure 1(b)). Building on this redifferentiation process, MSCs could further promote GAG synthesis of chondrocytes due to their trophic effects. Quantitative measurements for GAG and total collagen confirmed our impression on histological staining (Figures 1(c) and 1(d)). Without FGF18 in the medium, 1 μg of DNA is roughly corresponding to 2 μg of GAG and 1 μg hydroxyproline. And there is no difference between monoculture and coculture pellets. Once FGF18 is added to the medium, three- to fourfold increases were seen in GAG and collagen quantification. More importantly, GAG produced in coculture pellets was significantly more than that in monoculture pellets. The same trend was observed in collagen quantification. Since hydroxyproline assay cannot distinguish cartilage specific collagen type II (COL II) from other collagens, ELISA was performed to quantify the amount of COL II in cell pellets. Upon stimulation of FGF18, monoculture and coculture pellets deposit 3-4 times more COL II in matrix. As expected, coculture pellets contain significantly more COL II than monoculture pellets (Figure 1(e)). This indicated that trophic effects of MSCs may increase matrix formation of OA chondrocytes with the redifferentiation of chondrocytes stimulated by FGF1.

Bottom Line: Finally, coimplantation of MSCs with OA chondrocytes produces more matrix than chondrocytes only.In conclusion, FGF18 can restore the responsiveness of OA chondrocytes to the trophic effects of MSCs.Coimplantation of MSCs and OA chondrocytes treated with FGF18 may be a good alternative cell source for regenerating cartilage tissue that is degraded during OA pathological changes.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthopedic Surgery, The First Affiliated Hospital of Harbin Medical University, 23 You Zheng Street, Nangang District, Harbin 150001, China.

ABSTRACT
Coculture of mesenchymal stem cells with chondrocytes increases production of cartilaginous matrix. Chondrocytes isolated from late stage osteoarthritic patients usually lost their phenotype of producing cartilaginous matrix. Fibroblast growth factor 18 is believed to redifferentiate OA chondrocyte into functionally active chondrocytes. The aim of this study is to investigate the supportive effects of MSCs on OA chondrocytes and test if FGF18 could enhance the responsiveness of OA chondrocytes to the support of MSCs in a coculture system. Both pellet and transwell co-cultures were used. GAG quantification, hydroxyproline assay, and qPCR were performed. An ectopic models of cartilage formation was also applied. Our data indicated that, in pellets coculture of MSCs and OA chondrocytes, matrix production was increased in the presence of FGF18, comparing to the monoculture of chondrocytes. Results from transwell coculture study showed that expression of matrix producing genes in OA chondrocytes increased when cocultured with MSCs with FGF18 in culture medium, while hypertrophic genes were not changed by coculture. Finally, coimplantation of MSCs with OA chondrocytes produces more matrix than chondrocytes only. In conclusion, FGF18 can restore the responsiveness of OA chondrocytes to the trophic effects of MSCs. Coimplantation of MSCs and OA chondrocytes treated with FGF18 may be a good alternative cell source for regenerating cartilage tissue that is degraded during OA pathological changes.

No MeSH data available.


Related in: MedlinePlus