{"title":"[aGVHD小鼠血清损伤骨髓间充质干细胞成脂分化体外细胞模型的建立及应用]。","authors":"Run-Xiang Xu, Pei-Lin Li, Jia-Yi Tian, Jie Tang, Bo-Feng Yin, Fu-Hao Yu, Fei-Yan Wang, Xiao-Tong Li, Xiao-Yu Zhang, Wen-Rong Xia, Heng Zhu, Li Ding","doi":"10.19746/j.cnki.issn.1009-2137.2025.01.038","DOIUrl":null,"url":null,"abstract":"<p><strong>Objective: </strong>To establish an <i>in vitro</i> cell model simulating acute graft-versus-host disease (aGVHD) bone marrow microenvironment injury with the advantage of mouse serum of aGVHD model and explore the effect of serum of aGVHD mouse on the adipogenic differentiation ability of mesenchymal stem cells (MSCs).</p><p><strong>Methods: </strong>The 6-8-week-old C57BL/6N female mice and BALB/c female mice were used as the donor and recipient mice of the aGVHD model, respectively. Bone marrow transplantation (BMT) mouse model (<i>n</i>=20) was established by being injected with bone marrow cells (1×10<sup>7</sup> per mouse) from donor mice within 4-6 hours after receiving a lethal dose (8.0 Gy, 72.76 cGy/min) of γ ray general irradiation. A mouse model of aGVHD (<i>n</i>=20) was established by infusing a total of 0.4 ml of a mixture of donor mouse-derived bone marrow cells (1×10<sup>7</sup> per mouse) and spleen lymphocytes (2×10<sup>6</sup> per mouse). The blood was removed from the eyeballs and the mouse serum was aspirated on the 7<sup>th</sup> day after modeling. Bone marrow-derived MSCs were isolated from 1-week-old C57BL/6N male mice and incubated with 2%, 5% and 10% BMT mouse serum and aGVHD mouse serum in the medium, respectively. The effect of serum in the two groups on the <i>in vitro</i> adipogenic differentiation ability of mouse MSCs was detected by Oil Red O staining. The expression levels of related proteins PPARγ and CEBPα were detected by Western blot. The expression differences of key adipogenic transcription factors including <i>PPARγ, CEBPα, FABP4</i> and <i>LPL</i> were determined by real-time quantitative PCR (RT-qPCR).</p><p><strong>Results: </strong>An <i>in vitro</i> cell model simulating the damage of bone marrow microenvironment in mice with aGVHD was successfully established. Oil Red O staining showed that the number of orange-red fatty droplets was significantly reduced and the adipogenic differentiation ability of MSC was impaired at aGVHD serum concentration of 10% compared with BMT serum. Western blot experiments showed that adipogenesis-related proteins PPARγ and CEBPα expressed in MSCs were down-regulated. Further RT-qPCR assay showed that the production of <i>PPARγ, CEBPα, FABP4</i> and <i>LPL</i>, the key transcription factors for adipogenic differentiation of MSC, were significantly reduced.</p><p><strong>Conclusion: </strong>The adipogenic differentiation capacity of MSCs is inhibited by aGVHD mouse serum.</p>","PeriodicalId":35777,"journal":{"name":"中国实验血液学杂志","volume":"33 1","pages":"255-261"},"PeriodicalIF":0.0000,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"[Establishment and Application of an in Vitro Cellular Model of Adipogenic Differentiation of Bone Marrow Mesenchymal Stem Cells with Serum Injury in aGVHD Mouse].\",\"authors\":\"Run-Xiang Xu, Pei-Lin Li, Jia-Yi Tian, Jie Tang, Bo-Feng Yin, Fu-Hao Yu, Fei-Yan Wang, Xiao-Tong Li, Xiao-Yu Zhang, Wen-Rong Xia, Heng Zhu, Li Ding\",\"doi\":\"10.19746/j.cnki.issn.1009-2137.2025.01.038\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Objective: </strong>To establish an <i>in vitro</i> cell model simulating acute graft-versus-host disease (aGVHD) bone marrow microenvironment injury with the advantage of mouse serum of aGVHD model and explore the effect of serum of aGVHD mouse on the adipogenic differentiation ability of mesenchymal stem cells (MSCs).</p><p><strong>Methods: </strong>The 6-8-week-old C57BL/6N female mice and BALB/c female mice were used as the donor and recipient mice of the aGVHD model, respectively. Bone marrow transplantation (BMT) mouse model (<i>n</i>=20) was established by being injected with bone marrow cells (1×10<sup>7</sup> per mouse) from donor mice within 4-6 hours after receiving a lethal dose (8.0 Gy, 72.76 cGy/min) of γ ray general irradiation. A mouse model of aGVHD (<i>n</i>=20) was established by infusing a total of 0.4 ml of a mixture of donor mouse-derived bone marrow cells (1×10<sup>7</sup> per mouse) and spleen lymphocytes (2×10<sup>6</sup> per mouse). The blood was removed from the eyeballs and the mouse serum was aspirated on the 7<sup>th</sup> day after modeling. Bone marrow-derived MSCs were isolated from 1-week-old C57BL/6N male mice and incubated with 2%, 5% and 10% BMT mouse serum and aGVHD mouse serum in the medium, respectively. The effect of serum in the two groups on the <i>in vitro</i> adipogenic differentiation ability of mouse MSCs was detected by Oil Red O staining. The expression levels of related proteins PPARγ and CEBPα were detected by Western blot. The expression differences of key adipogenic transcription factors including <i>PPARγ, CEBPα, FABP4</i> and <i>LPL</i> were determined by real-time quantitative PCR (RT-qPCR).</p><p><strong>Results: </strong>An <i>in vitro</i> cell model simulating the damage of bone marrow microenvironment in mice with aGVHD was successfully established. Oil Red O staining showed that the number of orange-red fatty droplets was significantly reduced and the adipogenic differentiation ability of MSC was impaired at aGVHD serum concentration of 10% compared with BMT serum. Western blot experiments showed that adipogenesis-related proteins PPARγ and CEBPα expressed in MSCs were down-regulated. Further RT-qPCR assay showed that the production of <i>PPARγ, CEBPα, FABP4</i> and <i>LPL</i>, the key transcription factors for adipogenic differentiation of MSC, were significantly reduced.</p><p><strong>Conclusion: </strong>The adipogenic differentiation capacity of MSCs is inhibited by aGVHD mouse serum.</p>\",\"PeriodicalId\":35777,\"journal\":{\"name\":\"中国实验血液学杂志\",\"volume\":\"33 1\",\"pages\":\"255-261\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2025-02-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"中国实验血液学杂志\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.19746/j.cnki.issn.1009-2137.2025.01.038\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q4\",\"JCRName\":\"Medicine\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"中国实验血液学杂志","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.19746/j.cnki.issn.1009-2137.2025.01.038","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"Medicine","Score":null,"Total":0}
[Establishment and Application of an in Vitro Cellular Model of Adipogenic Differentiation of Bone Marrow Mesenchymal Stem Cells with Serum Injury in aGVHD Mouse].
Objective: To establish an in vitro cell model simulating acute graft-versus-host disease (aGVHD) bone marrow microenvironment injury with the advantage of mouse serum of aGVHD model and explore the effect of serum of aGVHD mouse on the adipogenic differentiation ability of mesenchymal stem cells (MSCs).
Methods: The 6-8-week-old C57BL/6N female mice and BALB/c female mice were used as the donor and recipient mice of the aGVHD model, respectively. Bone marrow transplantation (BMT) mouse model (n=20) was established by being injected with bone marrow cells (1×107 per mouse) from donor mice within 4-6 hours after receiving a lethal dose (8.0 Gy, 72.76 cGy/min) of γ ray general irradiation. A mouse model of aGVHD (n=20) was established by infusing a total of 0.4 ml of a mixture of donor mouse-derived bone marrow cells (1×107 per mouse) and spleen lymphocytes (2×106 per mouse). The blood was removed from the eyeballs and the mouse serum was aspirated on the 7th day after modeling. Bone marrow-derived MSCs were isolated from 1-week-old C57BL/6N male mice and incubated with 2%, 5% and 10% BMT mouse serum and aGVHD mouse serum in the medium, respectively. The effect of serum in the two groups on the in vitro adipogenic differentiation ability of mouse MSCs was detected by Oil Red O staining. The expression levels of related proteins PPARγ and CEBPα were detected by Western blot. The expression differences of key adipogenic transcription factors including PPARγ, CEBPα, FABP4 and LPL were determined by real-time quantitative PCR (RT-qPCR).
Results: An in vitro cell model simulating the damage of bone marrow microenvironment in mice with aGVHD was successfully established. Oil Red O staining showed that the number of orange-red fatty droplets was significantly reduced and the adipogenic differentiation ability of MSC was impaired at aGVHD serum concentration of 10% compared with BMT serum. Western blot experiments showed that adipogenesis-related proteins PPARγ and CEBPα expressed in MSCs were down-regulated. Further RT-qPCR assay showed that the production of PPARγ, CEBPα, FABP4 and LPL, the key transcription factors for adipogenic differentiation of MSC, were significantly reduced.
Conclusion: The adipogenic differentiation capacity of MSCs is inhibited by aGVHD mouse serum.
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