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Abnormal osteogenic and chondrogenic differentiation of human mesenchymal stem cells from patients with adolescent idiopathic scoliosis in response to melatonin.

Chen C, Xu C, Zhou T, Gao B, Zhou H, Chen C, Zhang C, Huang D, Su P - Mol Med Rep (2016)

Bottom Line: Alkaline phosphatase (ALP) activity assays, quantitative glycosaminoglycan (GAG) analysis, and reverse transcription‑quantitative polymerase chain reaction analysis were performed.Melatonin increased ALP activity, GAG synthesis and upregulated the expression of genes involved in osteogenic and chondrogenic differentiation including, ALP, osteopontin, osteocalcin, runt-related transcription factor 2, collagen type II, collagen type X, aggrecan and sex‑determining region Y-box 9 in the normal control hMSCs, but did not affect the AIS groups.These results indicate a potential modulating role of melatonin via the MT2 receptor on abnormal osteogenic and chondrogenic differentiaation in patients with AIS.

View Article: PubMed Central - PubMed

Affiliation: Department of Spine Surgery, The First Affiliated Hospital of Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.

ABSTRACT
Abnormalities of membranous and endochondral ossification in patients with adolescent idiopathic scoliosis (AIS) remain incompletely understood. To investigate abnormalities in the melatonin signaling pathway and cellular response to melatonin in AIS, a case‑control study of osteogenic and chondrogenic differentiation was performed using human mesenchymal stem cells (hMSCs). AIS was diagnosed by physical and radiographic examination. hMSCs were isolated from the bone marrow of patients with AIS and control subjects (n=12 each), and purified by density gradient centrifugation. The expression levels of melatonin receptors (MTs) 1 and 2 were detected by western blotting. Osteogenic and chondrogenic differentiation was induced by culturing hMSCs in osteogenic and chondrogenic media containing vehicle or 50 nM melatonin. Alkaline phosphatase (ALP) activity assays, quantitative glycosaminoglycan (GAG) analysis, and reverse transcription‑quantitative polymerase chain reaction analysis were performed. Compared with controls, MT2 demonstrated low expression in the AIS group. Melatonin increased ALP activity, GAG synthesis and upregulated the expression of genes involved in osteogenic and chondrogenic differentiation including, ALP, osteopontin, osteocalcin, runt-related transcription factor 2, collagen type II, collagen type X, aggrecan and sex‑determining region Y-box 9 in the normal control hMSCs, but did not affect the AIS groups. Thus, AIS hMSCs exhibit abnormal cellular responses to melatonin during osteogenic and chondrogenic differentiation, which may be associated with abnormal membranous and endochondral ossification, and skeletal growth. These results indicate a potential modulating role of melatonin via the MT2 receptor on abnormal osteogenic and chondrogenic differentiaation in patients with AIS.

No MeSH data available.


Related in: MedlinePlus

Effect of melatonin on quantitative analysis of GAG in human mesenchymal stem cells. GAG content was analyzed quantitatively and normalized by DNA content after 21 days of differentiation. Melatonin treatment increased GAG synthesis significantly in the normal control groups, but not in the AIS groups. *P<0.05 vs. control-CHO group. GAG, glycoaminoglycan; CHO, chondrogenesis; AIS, adolescent idiopathic scoliosis; Mel, melatonin.
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f4-mmr-14-02-1201: Effect of melatonin on quantitative analysis of GAG in human mesenchymal stem cells. GAG content was analyzed quantitatively and normalized by DNA content after 21 days of differentiation. Melatonin treatment increased GAG synthesis significantly in the normal control groups, but not in the AIS groups. *P<0.05 vs. control-CHO group. GAG, glycoaminoglycan; CHO, chondrogenesis; AIS, adolescent idiopathic scoliosis; Mel, melatonin.

Mentions: During chondrogenic differentiation, melatonin treatment increased GAG synthesis in normal control MSCs, which was elevated significantly on day 21 compared with untreated normal control cells (P=0.0318), however, melatonin exhibited no effect in the AIS groups (Fig. 4). Consistent with the results of quantitative GAG analysis, during chondrogenic differentiation, melatonin significantly upregulated the mRNA expression levels of COL2A1, COL10A1, aggrecan and SOX9 in the normal control group compared with untreated cells (P=0.00415, P=0.01942, P=0.02773 and P=0.0069, respectively), which are critical for chondrogenic differentiation, however, no effect was observed in the AIS groups (Fig. 5). The SOX9 level was marginally downregulated in the AIS-CHO/Mel group compared with the AIS-CHO group, although this difference was not significant.


Abnormal osteogenic and chondrogenic differentiation of human mesenchymal stem cells from patients with adolescent idiopathic scoliosis in response to melatonin.

Chen C, Xu C, Zhou T, Gao B, Zhou H, Chen C, Zhang C, Huang D, Su P - Mol Med Rep (2016)

Effect of melatonin on quantitative analysis of GAG in human mesenchymal stem cells. GAG content was analyzed quantitatively and normalized by DNA content after 21 days of differentiation. Melatonin treatment increased GAG synthesis significantly in the normal control groups, but not in the AIS groups. *P<0.05 vs. control-CHO group. GAG, glycoaminoglycan; CHO, chondrogenesis; AIS, adolescent idiopathic scoliosis; Mel, melatonin.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4940077&req=5

f4-mmr-14-02-1201: Effect of melatonin on quantitative analysis of GAG in human mesenchymal stem cells. GAG content was analyzed quantitatively and normalized by DNA content after 21 days of differentiation. Melatonin treatment increased GAG synthesis significantly in the normal control groups, but not in the AIS groups. *P<0.05 vs. control-CHO group. GAG, glycoaminoglycan; CHO, chondrogenesis; AIS, adolescent idiopathic scoliosis; Mel, melatonin.
Mentions: During chondrogenic differentiation, melatonin treatment increased GAG synthesis in normal control MSCs, which was elevated significantly on day 21 compared with untreated normal control cells (P=0.0318), however, melatonin exhibited no effect in the AIS groups (Fig. 4). Consistent with the results of quantitative GAG analysis, during chondrogenic differentiation, melatonin significantly upregulated the mRNA expression levels of COL2A1, COL10A1, aggrecan and SOX9 in the normal control group compared with untreated cells (P=0.00415, P=0.01942, P=0.02773 and P=0.0069, respectively), which are critical for chondrogenic differentiation, however, no effect was observed in the AIS groups (Fig. 5). The SOX9 level was marginally downregulated in the AIS-CHO/Mel group compared with the AIS-CHO group, although this difference was not significant.

Bottom Line: Alkaline phosphatase (ALP) activity assays, quantitative glycosaminoglycan (GAG) analysis, and reverse transcription‑quantitative polymerase chain reaction analysis were performed.Melatonin increased ALP activity, GAG synthesis and upregulated the expression of genes involved in osteogenic and chondrogenic differentiation including, ALP, osteopontin, osteocalcin, runt-related transcription factor 2, collagen type II, collagen type X, aggrecan and sex‑determining region Y-box 9 in the normal control hMSCs, but did not affect the AIS groups.These results indicate a potential modulating role of melatonin via the MT2 receptor on abnormal osteogenic and chondrogenic differentiaation in patients with AIS.

View Article: PubMed Central - PubMed

Affiliation: Department of Spine Surgery, The First Affiliated Hospital of Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.

ABSTRACT
Abnormalities of membranous and endochondral ossification in patients with adolescent idiopathic scoliosis (AIS) remain incompletely understood. To investigate abnormalities in the melatonin signaling pathway and cellular response to melatonin in AIS, a case‑control study of osteogenic and chondrogenic differentiation was performed using human mesenchymal stem cells (hMSCs). AIS was diagnosed by physical and radiographic examination. hMSCs were isolated from the bone marrow of patients with AIS and control subjects (n=12 each), and purified by density gradient centrifugation. The expression levels of melatonin receptors (MTs) 1 and 2 were detected by western blotting. Osteogenic and chondrogenic differentiation was induced by culturing hMSCs in osteogenic and chondrogenic media containing vehicle or 50 nM melatonin. Alkaline phosphatase (ALP) activity assays, quantitative glycosaminoglycan (GAG) analysis, and reverse transcription‑quantitative polymerase chain reaction analysis were performed. Compared with controls, MT2 demonstrated low expression in the AIS group. Melatonin increased ALP activity, GAG synthesis and upregulated the expression of genes involved in osteogenic and chondrogenic differentiation including, ALP, osteopontin, osteocalcin, runt-related transcription factor 2, collagen type II, collagen type X, aggrecan and sex‑determining region Y-box 9 in the normal control hMSCs, but did not affect the AIS groups. Thus, AIS hMSCs exhibit abnormal cellular responses to melatonin during osteogenic and chondrogenic differentiation, which may be associated with abnormal membranous and endochondral ossification, and skeletal growth. These results indicate a potential modulating role of melatonin via the MT2 receptor on abnormal osteogenic and chondrogenic differentiaation in patients with AIS.

No MeSH data available.


Related in: MedlinePlus