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Rffect of steroid on mRNA expression of neuropeptide receptors in human bone marrow mesenchymal stem cells

Yisheng Wang, Zhanfeng Zhang, Shao-yang Zhu

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Abstract

Objective To investigate the effect of steroid on the mRNA expression of calcitonin gene-related peptide receptor (CGRPR),substance P receptor (SPR),peroxisome proliferators-activated receptor-γ (PPARγ),and Runx2 in human bone marrow mesenchymal stem cells (hBMSCs).Methods hBMSCs were isolated and purified by density gradient centrifugation combined with plastic-adhering culture,and identified by flow cytometry.At the third generation,hBMSCs were randomly divided into two groups:experimental group and control group.The hBMSCs in experimental group were treated with 1 ×10 -7 mol/L dexamethasone,and those in control group were routinely cultured.The morphological changes and growth trend wer observed,and adipocytes were stained with oil red O.The expression changes of mRNA were analyzed by using real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR).Results Adherent cells were fiber-shaped and had a swirl-like growth.The positive rate of CD29 and CD44 was ( 91.4 ± 0.7 ) ％ and ( 93.8 ± 0.5 ) ％,while that of CD34 was ( 2.7 ± 0.8 ) ％,respectively.The cells in experimental group grew slower than in control group,while those in control group proliferated greatly.The number of adipocytes in control group was increased significantly.The mRNA expression levels of CGRPR,SPR and Runx2 in experimental group was significantlly lower than those in control group,with the 2△△Ct being 0.10 ±0.03,0.16 ±0.04,and 0.23 ±0.05 respectively ( all P＜0.01),while the 2△△Ct of PPARγwas 5.08 ±3.37 (P＜0.01).Conclusion Steroid could induce hBMSCs adipogenic differentiation,inhibit the mRNA expression of CGRPR,SPR,Runx2,and then restrain cells growth,proliferative ability and osteogenic differentiation of the hBMSCs,which might be related to the pathogenesis of steroid-induced osteonecrosis. Key words: Bone marrow mesenchymal stem cells; Steroid; Neuropeptide; Calcitonin generelated peptide receptor; Substance P receptor

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Objective To investigate the effect of steroid on the mRNA expression of calcitonin gene-related peptide receptor (CGRPR),substance P receptor (SPR),peroxisome proliferators-activated receptor-γ (PPARγ),and Runx2 in human bone marrow mesenchymal stem cells (hBMSCs).Methods hBMSCs were isolated and purified by density gradient centrifugation combined with plastic-adhering culture,and identified by flow cytometry.At the third generation,hBMSCs were randomly divided into two groups:experimental group and control group.The hBMSCs in experimental group were treated with 1 ×10 -7 mol/L dexamethasone,and those in control group were routinely cultured.The morphological changes and growth trend wer observed,and adipocytes were stained with oil red O.The expression changes of mRNA were analyzed by using real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR).Results Adherent cells were fiber-shaped and had a swirl-like growth.The positive rate of CD29 and CD44 was ( 91.4 ± 0.7 ) ％ and ( 93.8 ± 0.5 ) ％,while that of CD34 was ( 2.7 ± 0.8 ) ％,respectively.The cells in experimental group grew slower than in control group,while those in control group proliferated greatly.The number of adipocytes in control group was increased significantly.The mRNA expression levels of CGRPR,SPR and Runx2 in experimental group was significantlly lower than those in control group,with the 2△△Ct being 0.10 ±0.03,0.16 ±0.04,and 0.23 ±0.05 respectively ( all P＜0.01),while the 2△△Ct of PPARγwas 5.08 ±3.37 (P＜0.01).Conclusion Steroid could induce hBMSCs adipogenic differentiation,inhibit the mRNA expression of CGRPR,SPR,Runx2,and then restrain cells growth,proliferative ability and osteogenic differentiation of the hBMSCs,which might be related to the pathogenesis of steroid-induced osteonecrosis. Key words: Bone marrow mesenchymal stem cells; Steroid; Neuropeptide; Calcitonin generelated peptide receptor; Substance P receptor

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Available abstract

Objective To investigate the effect of steroid on the mRNA expression of calcitonin gene-related peptide receptor (CGRPR),substance P receptor (SPR),peroxisome proliferators-activated receptor-γ (PPARγ),and Runx2 in human bone marrow mesenchymal stem cells (hBMSCs).Methods hBMSCs were isolated and purified by density gradient centrifugation combined with plastic-adhering culture,and identified by flow cytometry.At the third generation,hBMSCs were randomly divided into two groups:experimental group and control group.The hBMSCs in experimental group were treated with 1 ×10 -7 mol/L dexamethasone,and those in control group were routinely cultured.The morphological changes and growth trend wer observed,and adipocytes were stained with oil red O.The expression changes of mRNA were analyzed by using real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR).Results Adherent cells were fiber-shaped and had a swirl-like growth.The positive rate of CD29 and CD44 was ( 91.4 ± 0.7 ) ％ and ( 93.8 ± 0.5 ) ％,while that of CD34 was ( 2.7 ± 0.8 ) ％,respectively.The cells in experimental group grew slower than in control group,while those in control group proliferated greatly.The number of adipocytes in control group was increased significantly.The mRNA expression levels of CGRPR,SPR and Runx2 in experimental group was significantlly lower than those in control group,with the 2△△Ct being 0.10 ±0.03,0.16 ±0.04,and 0.23 ±0.05 respectively ( all P＜0.01),while the 2△△Ct of PPARγwas 5.08 ±3.37 (P＜0.01).Conclusion Steroid could induce hBMSCs adipogenic differentiation,inhibit the mRNA expression of CGRPR,SPR,Runx2,and then restrain cells growth,proliferative ability and osteogenic differentiation of the hBMSCs,which might be related to the pathogenesis of steroid-induced osteonecrosis. Key words: Bone marrow mesenchymal stem cells; Steroid; Neuropeptide; Calcitonin generelated peptide receptor; Substance P receptor

Key concepts: Mesenchymal stem cell, CD44, RUNX2, Flow cytometry, CD34, Receptor, Messenger RNA, Molecular biology

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Rffect of steroid on mRNA expression of neuropeptide receptors in human bone marrow mesenchymal stem cells — Research Paper | ScholarLens