Institute of Geriatrics (Shanghai University), Affiliated Nantong Hospital of Shanghai University (The Sixth People's Hospital of Nantong), School of Medicine, Shanghai University, Nantong 226011, China; Cardiac Regeneration and Ageing Lab, Institute of Cardiovascular Sciences, Shanghai Engineering Research Center of Organ Repair, School of Life Science, Shanghai University, Shanghai 200444, China.
Research Center for Translational Medicine at East Hospital, Tongji University School of Life Science and Technology, Shanghai, 200092, China.
EBioMedicine. 2022 Nov;85:104274. doi: 10.1016/j.ebiom.2022.104274. Epub 2022 Sep 28.
Homeodomain-Interacting Protein Kinase 2 (HIPK2) has been reported to maintain basal cardiac function, however, its role in pathological cardiac remodeling remains unclear.
HIPK2 inhibitors (tBID and PKI1H) treated mice and two lines of HIPK2 mice were subjected to transverse aortic constriction (TAC). HIPK2 knockdown were performed in neonatal rat cardiomyocytes (NRCMs), neonatal rat cardiac fibroblasts (NRCFs), and human embryonic stem cell-derived cardiomyocytes (hESC-CMs). Microarray analysis was used to screen HIPK2 targets. Overexpression of early growth response 3 (EGR3) and C-type lectin receptor 4D (CLEC4D) were performed in NRCMs, while an activator of Smad3 was used in NRCFs, to rescue the effects of HIPK2 knockdown. Finally, the effects of EGR3 and CLEC4D knockdown by AAV9 in TAC were determined.
HIPK2 was elevated in TAC mice model, as well as cardiomyocyte hypertrophy and NRCFs fibrosis model. Pharmacological and genetic inhibition of HIPK2 improved cardiac function and suppressed cardiac hypertrophy and fibrosis induced by TAC. In vitro, HIPK2 inhibition prevented cardiomyocyte hypertrophic growth and NRCFs proliferation and differentiation. At the mechanistic level, we identified EGR3 and CLEC4D as new targets of HIPK2, which were regulated by ERK1/2-CREB and mediated the protective function of HIPK2 inhibition in cardiomyocytes. Meanwhile, inhibition of phosphorylation of Smad3 was responsible for the suppression of cardiac fibroblasts proliferation and differentiation by HIPK2 inhibition. Finally, we found that inhibition of EGR3 or CLEC4D protected against TAC.
HIPK2 inhibition protects against pathological cardiac remodeling by reducing EGR3 and CLEC4D with ERK1/2-CREB inhibition in cardiomyocytes, and by suppressing the phosphorylation of Smad3 in cardiac fibroblasts.
This work was supported by the grants from National Key Research and Development Project (2018YFE0113500 to J.X.), National Natural Science Foundation of China (82020108002 and 81911540486 to J.X., 81400647 to MJ Xu), the grant from Science and Technology Commission of Shanghai Municipality (21XD1421300 and 20DZ2255400 to J.X.), the "Dawn" Program of Shanghai Education Commission (19SG34 to J.X.), and Shanghai Sailing Program (21YF1413200 to Q.Z.).
同源结构域相互作用蛋白激酶 2（HIPK2）已被报道维持基础心脏功能，但其在病理性心脏重构中的作用尚不清楚。
用 HIPK2 抑制剂（tBID 和 PKI1H）处理小鼠和两种 HIPK2 敲除小鼠，并进行横主动脉缩窄（TAC）。在乳鼠心肌细胞（NRCMs）、乳鼠心肌成纤维细胞（NRCFs）和人胚胎干细胞衍生的心肌细胞（hESC-CMs）中进行 HIPK2 敲低。采用微阵列分析筛选 HIPK2 靶标。在 NRCMs 中过表达早期生长反应基因 3（EGR3）和 C 型凝集素受体 4D（CLEC4D），在 NRCFs 中使用 Smad3 激活剂来挽救 HIPK2 敲低的作用。最后，通过 TAC 中的 AAV9 确定 EGR3 和 CLEC4D 敲低的效果。
在 TAC 小鼠模型以及心肌肥厚和 NRCFs 纤维化模型中，HIPK2 升高。药理学和遗传学抑制 HIPK2 可改善心脏功能，并抑制 TAC 引起的心肌肥厚和纤维化。在体外，HIPK2 抑制可防止心肌细胞肥大生长和 NRCFs 增殖和分化。在机制水平上，我们确定 EGR3 和 CLEC4D 是 HIPK2 的新靶标，它们受 ERK1/2-CREB 调节，并介导 HIPK2 抑制在心肌细胞中的保护作用。同时，抑制 Smad3 的磷酸化是 HIPK2 抑制抑制心肌成纤维细胞增殖和分化的原因。最后，我们发现抑制 EGR3 或 CLEC4D 可防止 TAC。
HIPK2 抑制通过抑制 ERK1/2-CREB 抑制心肌细胞中的 EGR3 和 CLEC4D，以及抑制心脏成纤维细胞中 Smad3 的磷酸化，从而防止病理性心脏重构。
本工作得到国家重点研发计划（2018YFE0113500 给 J.X.）、国家自然科学基金（82020108002 和 81911540486 给 J.X.，81400647 给 MJ Xu）、上海市科委（21XD1421300 和 20DZ2255400 给 J.X.）、上海市教委“曙光计划”（19SG34 给 J.X.）和上海市扬帆计划（21YF1413200 给 Q.Z.）的资助。
