scholarly journals JKAMP Inhibits the Osteogenic Capacity of Adipose-derived Stem Cells in Diabetic Osteoporosis by Modulating the Wnt Signaling Pathway Through Intragenic DNA Methylation

2020 ◽  
Author(s):  
Shuanglin Peng ◽  
Sirong Shi ◽  
Gang Tao ◽  
Yanjing Li ◽  
Dexuan Xiao ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Stem Cells ◽  
Wnt Signaling ◽  
Signaling Pathway ◽  
Wnt Signaling Pathway ◽  
Underlying Mechanism ◽  
Osteogenic Potential ◽  
Adipose Derived Stem Cells ◽  
Alizarin Red ◽  
Diabetic Osteoporosis

Abstract Background: Diabetic osteoporosis (DOP) is a systemic metabolic bone disease caused by diabetes mellitus (DM). Adipose-derived stem cells (ASCs) play an important role in bone regeneration. Our previous study confirmed that ASCs from DOP mice (DOP-ASCs) have a lower osteogenesis potential compared with control ASCs (CON-ASCs). However, the cause of this poor osteogenesis has not been elucidated. Therefore, this study investigated the underlying mechanism of the decline in the osteogenic potential of DOP-ASCs from the perspective of epigenetics and explored methods to enhance their osteogenic capacity. Methods: The expression level of JNK1-associated membrane protein (JKAMP) and degree of DNA methylation in CON-ASCs and DOP-ASCs were measured by mRNA expression profiling and MeDIP sequencing, respectively. JKAMP small interfering RNA (siRNA) and a Jkamp overexpression plasmid were used to assess the role of JKAMP in osteogenic differentiation of CON-ASCs and DOP-ASCs. Immunofluorescence, qPCR, and western blotting were used to measure changes in expression of Wnt signaling pathway-related genes and osteogenesis-related molecules after osteogenesis induction. Alizarin red and ALP staining was used to confirm the osteogenic potential of stem cells. Bisulfite-specific PCR (BSP) was used to detect JKAMP methylation degree. Results: Expression of JKAMP and osteogenesis-related molecules (RUNX2 and OPN) in DOP-ASCs was decreased significantly in comparison with CON-ASCs. JKAMP silencing inhibited the Wnt signaling pathway and reduced the osteogenic ability of CON-ASCs. Overexpression of JKAMP in DOP-ASCs rescued the impaired osteogenic capacity caused by DOP. Moreover, JKAMP in DOP-ASCs contained intragenic DNA hypermethylated regions related to the downregulation of JKAMP expression. Conclusions: Intragenic DNA methylation inhibits the Wnt signaling pathway by suppressing expression of JKAMP and the osteogenic ability of DOP-ASCs. This study shows an epigenetic explanation for the reduced osteogenic ability of DOP-ASCs, and provides a potential therapeutic target to prevent and treat osteoporosis.

scholarly journals JKAMP inhibits the osteogenic capacity of adipose-derived stem cells in diabetic osteoporosis by modulating the Wnt signaling pathway through intragenic DNA methylation

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Shuanglin Peng ◽  
Sirong Shi ◽  
Gang Tao ◽  
Yanjing Li ◽  
Dexuan Xiao ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Stem Cells ◽  
Wnt Signaling ◽  
Signaling Pathway ◽  
Wnt Signaling Pathway ◽  
Underlying Mechanism ◽  
Osteogenic Potential ◽  
Adipose Derived Stem Cells ◽  
Alizarin Red ◽  
Diabetic Osteoporosis

Abstract Background Diabetic osteoporosis (DOP) is a systemic metabolic bone disease caused by diabetes mellitus (DM). Adipose-derived stem cells (ASCs) play an important role in bone regeneration. Our previous study confirmed that ASCs from DOP mice (DOP-ASCs) have a lower osteogenesis potential compared with control ASCs (CON-ASCs). However, the cause of this poor osteogenesis has not been elucidated. Therefore, this study investigated the underlying mechanism of the decline in the osteogenic potential of DOP-ASCs from the perspective of epigenetics and explored methods to enhance their osteogenic capacity. Methods The expression level of JNK1-associated membrane protein (JKAMP) and degree of DNA methylation in CON-ASCs and DOP-ASCs were measured by mRNA expression profiling and MeDIP sequencing, respectively. JKAMP small interfering RNA (siRNA) and a Jkamp overexpression plasmid were used to assess the role of JKAMP in osteogenic differentiation of CON-ASCs and DOP-ASCs. Immunofluorescence, qPCR, and western blotting were used to measure changes in expression of Wnt signaling pathway-related genes and osteogenesis-related molecules after osteogenesis induction. Alizarin red and ALP staining was used to confirm the osteogenic potential of stem cells. Bisulfite-specific PCR (BSP) was used to detect JKAMP methylation degree. Results Expression of JKAMP and osteogenesis-related molecules (RUNX2 and OPN) in DOP-ASCs was decreased significantly in comparison with CON-ASCs. JKAMP silencing inhibited the Wnt signaling pathway and reduced the osteogenic ability of CON-ASCs. Overexpression of JKAMP in DOP-ASCs rescued the impaired osteogenic capacity caused by DOP. Moreover, JKAMP in DOP-ASCs contained intragenic DNA hypermethylated regions related to the downregulation of JKAMP expression. Conclusions Intragenic DNA methylation inhibits the osteogenic ability of DOP-ASCs by suppressing expression of JKAMP and the Wnt signaling pathway. This study shows an epigenetic explanation for the reduced osteogenic ability of DOP-ASCs and provides a potential therapeutic target to prevent and treat osteoporosis.

scholarly journals Role of Fzd6 in Regulating the Osteogenic Differentiation of Adipose-derived Stem Cells in Osteoporosis

2021 ◽  
Author(s):  
Tianli Wu ◽  
Zhihao Yao ◽  
Gang Tao ◽  
Fangzhi Lou ◽  
Hui Tang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Osteogenic Differentiation ◽  
Wnt Signaling Pathway ◽  
Osteogenic Potential ◽  
Adipose Derived Stem Cells ◽  
Alizarin Red ◽  
Differentiation Potential

Abstract Objective: Although it has been demonstrated that adipose-derived stem cells (ASCs) from osteoporosis mice (OP-ASCs) exhibit impaired osteogenic differentiation potential, the molecular mechanism has not yet been elucidated. We found that Fzd6 was decreased in OP-ASCs compared with ASCs. This study investigates the effects and underlying mechanisms of Fzd6 in the osteogenic potential of OP-ASCs. Methods: Fzd6 expression in ASCs and OP-ASCs was measured by PCR gene chip. Fzd6 overexpression and silencing lentiviruses were used to evaluate the role of Fzd6 in the osteogenic differentiation of OP-ASCs. Real-time PCR (qPCR) and western blotting (WB) was performed to detect the expression of Fzd6 and bone-related molecules, including runt-related transcription factor 2 (Runx2) and osteopontin (Opn). Alizarin red staining and Alkaline phosphatase (ALP) staining was performed following osteogenic induction. Microscopic CT (Micro-CT), hematoxylin and eosin staining (H&E) staining, and Masson staining were used to assess the role of Fzd6 in osteogenic differentiation of osteoporosis (OP) mice in vivo.Results: Expression of Fzd6 was decreased significantly in OP-ASCs. Fzd6 silencing down-regulated the osteogenic ability of OP-ASCs in vitro. Overexpression of Fzd6 rescued the impaired osteogenic capacity in OP-ASCs in vitro. We obtained similar results in vivo.Conclusions: Fzd6 plays an important role in regulating the osteogenic ability of OP-ASCs both in vivo and in vitro. Overexpression of Fzd6 associated with the Wnt signaling pathway promotes the osteogenic ability of OP-ASCs, which provides new insights for the prevention and treatment of OP.

scholarly journals Mechanism of Wnt Pathway on Cartilage Differentiation of Adipose-Derived Stem cells

2020 ◽  
Author(s):  
Wanwu Dai ◽  
Bo Zhang ◽  
Zuquan Huang ◽  
Guili Sun ◽  
Yejing Yang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Wnt Signaling ◽  
Signaling Pathway ◽  
Chondrogenic Differentiation ◽  
Wnt Pathway ◽  
Wnt Signaling Pathway ◽  
Adipose Derived Stem Cells ◽  
Expression Levels ◽  
Cartilage Differentiation ◽  
Late Stages

Abstract Background: Adipose-derived stem cells ( ADSCs ) therapy is considered as a promising alternative to treat osteoarthritis (OA).Considerable evidence has shown that the Wnt signaling pathway is involved in chondrogenic differentiation in various stem cells.In the present study, we explored in detail the mechanism of of Wnt signaling pathway in the regulation of adipose-derived stem cell chondrogenic differentiation.Methohs: Adipose-derived stem cells were extracted from Sprague-Dawley groin area, isolated, cultured and verified. Then LiCl and DKK-1 used to induce ADSCs cartilage differentiation to detect indicators of cartilage at different stages, such as collagen type II (Collagen2a), aggrecan (Aggrecan) and Wnt pathway key proteins Sox9, β-catenin, Glycogen synthase kinase-3β (GSK-3β) expression.Subsequently, lentivirus was used as the vector, the full-length Sox9 gene was transfected into ADSCs and the expression levels of relevant indicators were detected again after stable expression.Results: In the process of inducing differentiation, the Wnt pathway promoted the rapid proliferation of ADSCs in the early, middle and late stages, and up-regulates Sox9 expression.Cartilage indicators Collagen 2a and Aggrecan expression levels were not significantly increased in the early stage, but significantly increased in the middle and late stages (P <0.05). At the later stage, however, the Wnt pathway down-regulated Sox9 expression, weakened the mature cartilage phenotype, promoted cartilage hypertrophy and early cartilage ossification. Finally, the overexpression of Sox9 feedback inhibits the Wnt pathway activity, down-regulates β-catenin expression, maintains cartilage phenotype, delays cartilage hypertrophy and early osteogenesis.Conclusion:The Wnt pathway regulates chondrogenic differentiation of ADSCs by regulating Sox9, but its regulatory mechanism is significantly different at different stages of induced differentiation.

scholarly journals DNA methylation mediated RSPO2 to promote follicular development in mammals

2021 ◽  
Vol 12 (7) ◽  
Author(s):  
Xiaofeng Zhou ◽  
Yingting He ◽  
Nian Li ◽  
Guofeng Bai ◽  
Xiangchun Pan ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Wnt Signaling ◽  
Signaling Pathway ◽  
Methylation Level ◽  
Molecular Mechanisms ◽  
Cpg Island ◽  
Wnt Signaling Pathway ◽  
Follicular Development ◽  
Expression Level

AbstractIn female mammals, the proliferation, apoptosis, and estradiol-17β (E2) secretion of granulosa cells (GCs) have come to decide the fate of follicles. DNA methylation and RSPO2 gene of Wnt signaling pathway have been reported to involve in the survival of GCs and follicular development. However, the molecular mechanisms for how DNA methylation regulates the expression of RSPO2 and participates in the follicular development are not clear. In this study, we found that the mRNA and protein levels of RSPO2 significantly increased during follicular development, but the DNA methylation level of RSPO2 promoter decreased gradually. Inhibition of DNA methylation or DNMT1 knockdown could decrease the methylation level of CpG island (CGI) in RSPO2 promoter and upregulate the expression level of RSPO2 in porcine GCs. The hypomethylation of −758/−749 and −563/−553 regions in RSPO2 promoter facilitated the occupancy of transcription factor E2F1 and promoted the transcriptional activity of RSPO2. Moreover, RSPO2 promoted the proliferation of GCs with increasing the expression level of PCNA, CDK1, and CCND1 and promoted the E2 secretion of GCs with increasing the expression level of CYP19A1 and HSD17B1 and inhibited the apoptosis of GCs with decreasing the expression level of Caspase3, cleaved Caspase3, cleaved Caspase8, cleaved Caspase9, cleaved PARP, and BAX. In addition, RSPO2 knockdown promoted the apoptosis of GCs, blocked the development of follicles, and delayed the onset of puberty with decreasing the expression level of Wnt signaling pathway-related genes (LGR4 and CTNNB1) in vivo. Taken together, the hypomethylation of −758/−749 and −563/−553 regions in RSPO2 promoter facilitated the occupancy of E2F1 and enhanced the transcription of RSPO2, which further promoted the proliferation and E2 secretion of GCs, inhibited the apoptosis of GCs, and ultimately ameliorated the development of follicles through Wnt signaling pathway. This study will provide useful information for further exploration on DNA-methylation-mediated RSPO2 pathway during follicular development.

scholarly journals Dickkopf Wnt signaling pathway inhibitor 1 regulates the differentiation of mouse embryonic stem cells in vitro and in vivo

2015 ◽  
Vol 13 (1) ◽  
pp. 720-730 ◽  
Author(s):  
LIPING OU ◽  
LIAOQIONG FANG ◽  
HEJING TANG ◽  
HAI QIAO ◽  
XIAOMEI ZHANG ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Wnt Signaling ◽  
Signaling Pathway ◽  
Wnt Signaling Pathway ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cells
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