Department of Biochemistry, University of Madras, Guindy Campus,Chennai 600025, India.
Department of Biochemistry, University of Madras, Guindy Campus,Chennai 600025, India.
Eur J Pharmacol. 2017 Oct 15;813:94-104. doi: 10.1016/j.ejphar.2017.07.042. Epub 2017 Jul 22.
The present study was aimed to determine whether stimulating Npr1 gene activity using Valporic acid (VA), a small short chain fatty acid molecule can enhance ANP mediated anti-hypertrophic activity in isoproterenol (ISO) - treated H9c2 cells in vitro. H9c2 cells were treated with ISO (10 M) and co-treated with VA (10 M) in the presence and absence of ANP (10M), for 48h. ATRA (10 M) was used as a positive inducer of Npr1 gene transcription. The mRNA expression of Npr1 and PKG-I genes, proto-oncogenes (c-fos, c-jun and c-myc) and hypertrophic markers (ANP, BNP, α-sk and β-MyHC), genes were determined by quantitative PCR (qPCR). The protein profiling of NPR-A, PKG-I and cGMP were evaluated by Western blot, immunofluorescence and ELISA respectively. A marked reduction in the level of expression of Npr1 (3- fold) and PKG-I (2.5-fold) genes and increased expression of proto-oncogenes (p< 0.001, respectively) and hypertrophic marker genes (p<0.001, respectively) were noticed in the ISO-treated H9c2 cells as compared with control cells. In contrast, the VA treated cells showed maximal Npr1 gene expression (3.5-fold) as compared with ATRA treated cells (2 fold), which is well correlated with the intracellular cGMP levels (80% vs 60%) and reduced (2.5-fold) HDAC -1&-2 mRNA expression. Furthermore, VA or ATRA treatment effectively reversed the ISO-induced altered expression of Npr1 and PKG-I genes, proto-oncogenes, and hypertrophic markers genes. Interestingly, the results of the present study suggest that ANP mediated anti-hypertrophic activity was enhanced with either VA (p<0.001) or ATRA (p<0.01) co-treatment. Together, we conclude that VA in combination with ANP can be a novel therapeutical approach for the treatment and management of left ventricular cardiac hypertrophy.
本研究旨在确定使用丙戊酸（VA）刺激 Npr1 基因活性是否可以增强 ANP 介导的异丙肾上腺素（ISO）处理的 H9c2 细胞体外抗肥大活性。将 H9c2 细胞用 ISO（10μM）处理，并在存在和不存在 ANP（10μM）的情况下用 VA（10μM）共同处理 48 小时。ATRA（10μM）用作 Npr1 基因转录的阳性诱导剂。通过定量 PCR（qPCR）测定 Npr1 和 PKG-I 基因、原癌基因（c-fos、c-jun 和 c-myc）和肥大标志物（ANP、BNP、α-sk 和 β-MyHC）的 mRNA 表达。通过 Western blot、免疫荧光和 ELISA 分别评估 NPR-A、PKG-I 和 cGMP 的蛋白谱。与对照细胞相比，ISO 处理的 H9c2 细胞中 Npr1（3 倍）和 PKG-I（2.5 倍）基因的表达水平明显降低，原癌基因（p<0.001，分别）和肥大标志物基因（p<0.001，分别）的表达增加。相比之下，与 ATRA 处理的细胞相比，VA 处理的细胞表现出最大的 Npr1 基因表达（3.5 倍），这与细胞内 cGMP 水平（80%对 60%）和减少的 HDAC-1&-2 mRNA 表达（2.5 倍）相关。此外，VA 或 ATRA 处理可有效逆转 ISO 诱导的 Npr1 和 PKG-I 基因、原癌基因和肥大标志物基因表达的改变。有趣的是，本研究的结果表明，无论是 VA（p<0.001）还是 ATRA（p<0.01）联合治疗，都可以增强 ANP 介导的抗肥大活性。总之，我们得出结论，VA 联合 ANP 可以成为治疗和管理左心室心肌肥厚的新治疗方法。
