{"title":"CREB1 抑制剂 666-15 可维持软骨的稳态并缓解骨关节炎的进展。","authors":"Ying Wang, Zhimin Wu, Guoqiang Yan, Shan Li, Yanzhuo Zhang, Guangping Li, Chengai Wu","doi":"10.1302/2046-3758.131.BJR-2023-0016.R2","DOIUrl":null,"url":null,"abstract":"<p><strong>Aims: </strong>cAMP response element binding protein (CREB1) is involved in the progression of osteoarthritis (OA). However, available findings about the role of CREB1 in OA are inconsistent. 666-15 is a potent and selective CREB1 inhibitor, but its role in OA is unclear. This study aimed to investigate the precise role of CREB1 in OA, and whether 666-15 exerts an anti-OA effect.</p><p><strong>Methods: </strong>CREB1 activity and expression of a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4) in cells and tissues were measured by immunoblotting and immunohistochemical (IHC) staining. The effect of 666-15 on chondrocyte viability and apoptosis was examined by cell counting kit-8 (CCK-8) assay, JC-10, and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) staining. The effect of 666-15 on the microstructure of subchondral bone, and the synthesis and catabolism of cartilage, in anterior cruciate ligament transection mice were detected by micro-CT, safranin O and fast green (S/F), immunohistochemical staining, and enzyme-linked immunosorbent assay (ELISA).</p><p><strong>Results: </strong>CREB1 was hyperactive in osteoarthritic articular cartilage, interleukin (IL)-1β-treated cartilage explants, and IL-1β- or carbonyl cyanide 3-chlorophenylhydrazone (CCCP)-treated chondrocytes. 666-15 enhanced cell viability of OA-like chondrocytes and alleviated IL-1β- or CCCP-induced chondrocyte injury through inhibition of mitochondrial dysfunction-associated apoptosis. Moreover, inhibition of CREB1 by 666-15 suppressed expression of ADAMTS4. Additionally, 666-15 alleviated joint degeneration in an ACLT mouse model.</p><p><strong>Conclusion: </strong>Hyperactive CREB1 played a critical role in OA development, and 666-15 exerted anti-IL-1β or anti-CCCP effects in vitro as well as joint-protective effects in vivo. 666-15 may therefore be used as a promising anti-OA drug.</p>","PeriodicalId":9074,"journal":{"name":"Bone & Joint Research","volume":"13 1","pages":"4-18"},"PeriodicalIF":4.7000,"publicationDate":"2024-01-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10758301/pdf/","citationCount":"0","resultStr":"{\"title\":\"The CREB1 inhibitor 666-15 maintains cartilage homeostasis and mitigates osteoarthritis progression.\",\"authors\":\"Ying Wang, Zhimin Wu, Guoqiang Yan, Shan Li, Yanzhuo Zhang, Guangping Li, Chengai Wu\",\"doi\":\"10.1302/2046-3758.131.BJR-2023-0016.R2\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Aims: </strong>cAMP response element binding protein (CREB1) is involved in the progression of osteoarthritis (OA). However, available findings about the role of CREB1 in OA are inconsistent. 666-15 is a potent and selective CREB1 inhibitor, but its role in OA is unclear. This study aimed to investigate the precise role of CREB1 in OA, and whether 666-15 exerts an anti-OA effect.</p><p><strong>Methods: </strong>CREB1 activity and expression of a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4) in cells and tissues were measured by immunoblotting and immunohistochemical (IHC) staining. The effect of 666-15 on chondrocyte viability and apoptosis was examined by cell counting kit-8 (CCK-8) assay, JC-10, and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) staining. The effect of 666-15 on the microstructure of subchondral bone, and the synthesis and catabolism of cartilage, in anterior cruciate ligament transection mice were detected by micro-CT, safranin O and fast green (S/F), immunohistochemical staining, and enzyme-linked immunosorbent assay (ELISA).</p><p><strong>Results: </strong>CREB1 was hyperactive in osteoarthritic articular cartilage, interleukin (IL)-1β-treated cartilage explants, and IL-1β- or carbonyl cyanide 3-chlorophenylhydrazone (CCCP)-treated chondrocytes. 666-15 enhanced cell viability of OA-like chondrocytes and alleviated IL-1β- or CCCP-induced chondrocyte injury through inhibition of mitochondrial dysfunction-associated apoptosis. Moreover, inhibition of CREB1 by 666-15 suppressed expression of ADAMTS4. Additionally, 666-15 alleviated joint degeneration in an ACLT mouse model.</p><p><strong>Conclusion: </strong>Hyperactive CREB1 played a critical role in OA development, and 666-15 exerted anti-IL-1β or anti-CCCP effects in vitro as well as joint-protective effects in vivo. 666-15 may therefore be used as a promising anti-OA drug.</p>\",\"PeriodicalId\":9074,\"journal\":{\"name\":\"Bone & Joint Research\",\"volume\":\"13 1\",\"pages\":\"4-18\"},\"PeriodicalIF\":4.7000,\"publicationDate\":\"2024-01-02\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10758301/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Bone & Joint Research\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1302/2046-3758.131.BJR-2023-0016.R2\",\"RegionNum\":2,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"CELL & TISSUE ENGINEERING\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Bone & Joint Research","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1302/2046-3758.131.BJR-2023-0016.R2","RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"CELL & TISSUE ENGINEERING","Score":null,"Total":0}
引用次数: 0
摘要
目的:cAMP 反应元件结合蛋白(CREB1)与骨关节炎(OA)的进展有关。然而,有关 CREB1 在 OA 中作用的现有研究结果并不一致。666-15 是一种强效的选择性 CREB1 抑制剂,但它在 OA 中的作用尚不清楚。本研究旨在探讨 CREB1 在 OA 中的确切作用,以及 666-15 是否具有抗 OA 作用:方法:通过免疫印迹和免疫组织化学(IHC)染色检测细胞和组织中 CREB1 的活性和具有血栓软骨素基序 4 的崩解素和金属蛋白酶(ADAMTS4)的表达。细胞计数试剂盒-8(CCK-8)测定法、JC-10 和末端脱氧核苷酸转移酶介导的 dUTP 缺口末端标记(TUNEL)染色法检测了 666-15 对软骨细胞活力和凋亡的影响。通过显微 CT、黄蓍素 O 和快绿素(S/F)、免疫组化染色和酶联免疫吸附试验(ELISA)检测了 666-15 对前十字韧带横断小鼠软骨下骨的微观结构以及软骨的合成和分解的影响:结果:CREB1在骨关节炎关节软骨、白细胞介素(IL)-1β处理过的软骨外植体、IL-1β或羰基氰化3-氯苯腙(CCCP)处理过的软骨细胞中活性亢进。666-15 通过抑制线粒体功能障碍相关的细胞凋亡,增强了 OA 类软骨细胞的细胞活力,减轻了 IL-1β 或 CCCP 诱导的软骨细胞损伤。此外,666-15 对 CREB1 的抑制还能抑制 ADAMTS4 的表达。此外,666-15 还能缓解 ACLT 小鼠模型中的关节退化:结论:亢进的 CREB1 在 OA 发生过程中起着关键作用,666-15 在体外具有抗 IL-1β 或抗CCCP 的作用,在体内具有保护关节的作用。因此,666-15 可以作为一种很有前景的抗 OA 药物。
The CREB1 inhibitor 666-15 maintains cartilage homeostasis and mitigates osteoarthritis progression.
Aims: cAMP response element binding protein (CREB1) is involved in the progression of osteoarthritis (OA). However, available findings about the role of CREB1 in OA are inconsistent. 666-15 is a potent and selective CREB1 inhibitor, but its role in OA is unclear. This study aimed to investigate the precise role of CREB1 in OA, and whether 666-15 exerts an anti-OA effect.
Methods: CREB1 activity and expression of a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4) in cells and tissues were measured by immunoblotting and immunohistochemical (IHC) staining. The effect of 666-15 on chondrocyte viability and apoptosis was examined by cell counting kit-8 (CCK-8) assay, JC-10, and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) staining. The effect of 666-15 on the microstructure of subchondral bone, and the synthesis and catabolism of cartilage, in anterior cruciate ligament transection mice were detected by micro-CT, safranin O and fast green (S/F), immunohistochemical staining, and enzyme-linked immunosorbent assay (ELISA).
Results: CREB1 was hyperactive in osteoarthritic articular cartilage, interleukin (IL)-1β-treated cartilage explants, and IL-1β- or carbonyl cyanide 3-chlorophenylhydrazone (CCCP)-treated chondrocytes. 666-15 enhanced cell viability of OA-like chondrocytes and alleviated IL-1β- or CCCP-induced chondrocyte injury through inhibition of mitochondrial dysfunction-associated apoptosis. Moreover, inhibition of CREB1 by 666-15 suppressed expression of ADAMTS4. Additionally, 666-15 alleviated joint degeneration in an ACLT mouse model.
Conclusion: Hyperactive CREB1 played a critical role in OA development, and 666-15 exerted anti-IL-1β or anti-CCCP effects in vitro as well as joint-protective effects in vivo. 666-15 may therefore be used as a promising anti-OA drug.