Rat Primary Cardiac Fibroblasts

ang II Induced the Upregulation of Rat Cardiac Fibroblasts' Expression of TIMP1 and Activated Fibrotic Signaling

AF is often related to atrial fibrosis, the primary cellular cause of which is cardiac fibroblast-mediated production of ECM. TIMP1, an ECM modulator, has been identified as a critical player in cardiovascular fibrosis; however, its role in AF remains unknown. Zhou's team examined the function of TIMP1 in the angiotensin II (ang II)-induced activation of rat cardiac fibroblasts.

The expression of TIMP1 in rat cardiac fibroblasts treated with different concentrations (1 nM, 10 nM, 100 nM, and 1 µM) and time points (12, 24, and 48 hours) of ang II was analyzed by qRT-PCR assay. The mRNA expression of TIMP1 was found to be maximum on 48 hours of treatment with 1 µM of ang II, which increased in a dose- and time-dependent manner (Fig. 1A, B). Western blotting analysis revealed a marked upregulation of TGF-β1, α-SMA, p-Smad3, and p-Smad2 in cells treated with 1 µM of ang II for 48 hours (Fig. 1C, D). However, non-phosphorylated forms of Smad2 and Smad3 were not significantly altered upon ang II treatment. These results suggest the possible involvement of TIMP1-mediated mechanisms linking ang II stimulation and activation of fibrotic signaling in AF.

Fmod Increased in DCM Rat Heart and RPCFs Treated with High Glucose of Fibrosis

Diabetic cardiomyopathy (DCM) is marked by excessive extracellular matrix protein deposition, leading to myocardial fibrosis. Fibromodulin (Fmod) is key in fibrotic diseases, but its role in DCM is unclear. Dai's team explored Fmod's role in DCM-related myocardial fibrosis using DCM rat models and high-glucose-exposed rat primary cardiac fibroblasts (RPCFs). They assessed mRNA and protein expression levels of Col1a1, Col3a1, α-SMA and Fmod in both models.

HE staining showed normal arrangement of myocardial fibers and cardiomyocytes in control rat hearts. In contrast, DCM group hearts had irregular, thickened, and fragmented fibers with increased interstitial spacing. Masson trichrome staining revealed more collagen fibers in the DCM group compared to controls. Immunohistochemistry also showed higher levels of Col1a1, Col3a1, and α-SMA in DCM hearts. Primary cardiac fibroblasts had a purity of over 90%. RT-qPCR and Western blot results showed elevated Fmod, Col1a1, Col3a1, and α-SMA levels in DCM hearts and high glucose-treated RPCFs. These findings indicate significant collagen production in DCM rats and enhanced collagen synthesis in RPCFs under high glucose conditions (Fig. 2).