P19.CL6

The Gene Expression Levels of Cd38, Ryr2, and Fkbp12.6 in Cardiomyocytes Were Decreased by IH

Sleep apnea syndrome (SAS) manifests as repeated oxygen desaturation and reoxygenation patterns known as intermittent hypoxia and poses risks for cardiovascular disease and insulin resistance/type 2 diabetes.

Takasawa et al. investigated the connections between IH stress and CVD. They treated rat H9c2 and mouse P19.CL6 cardiomyocytes with experimental IH or normoxia for 24 hours and examined mRNA expression levels of components in the Cd38-cyclic ADP-ribose (cADPR) signaling pathway. Using real-time reverse transcriptase-polymerase chain reaction (RT-PCR), they analyze mRNA levels of Cd38 (which encodes ADP-ribosyl cyclase/cyclic ADP-ribose [cADPR] hydrolase, EC 3.2.2.6), Ryr2 and Fkbp12.6 (a cADPR receptor [Fkbp1b]) in mouse P19.CL6 and rat H9c2 cells. IH led to significant reductions in mRNA levels of Cd38, Ryr2, and Fkbp12.6 within mouse P19.CL6 cardiomyocytes and rat H9c2 cardiomyocytes as demonstrated by Figure 1 and Figure 2. The researchers also evaluated the protein levels of Cd38, Ryr2, and Fkbp12.6 in H9c2 cells through Western blot assays and discovered significant reductions caused by IH with statistical significance values of p = 0.0205, p = 0.0241, and p = 0.0078, respectively (Fig. 3).

Fig. 1. The mRNA levels of mouse (A) Cd38, (B) Ryr2, and (C) Fkbp12.6 in mouse P19.CL6 cardiomyocytes (Takasawa S, Makino M, et al., 2022).

Fig. 2. The mRNA levels of mouse (A) Cd38, (B) Ryr2, and (C) Fkbp12.6 in rat H9c2 cardiomyocytes (Takasawa S, Makino M, et al., 2022).

Fig. 3. Relative protein expression levels of (A) Cd38, (B) Ryr2, and (C) Fkbp in rat H9c2 cardiomyocytes subjected to IH (Takasawa S, Makino M, et al., 2022).

Gene Expressions of Reg IV and Hgf Were Increased by IH in Cardiomyocytes

The study focuses on sleep apnea syndrome (SAS), which causes intermittent hypoxia (IH) and consequently is a significant risk factor for cardiovascular diseases (CVD) and metabolic disorders like insulin resistance and Type 2 diabetes. SAS affects oxygen levels during sleep, resulting in numerous health complications, including CVD. Takasawa et al. aims to explore the mechanisms underlying how IH influences cardiomyocyte gene expression.

They treated rat H9c2 cardiomyocytes and mouse P19.CL6 cells under normoxia, intermittent hypoxia (IH), or sustained hypoxia (SH) for 24 hours. They measured the mRNA levels related to inflammation, growth, and function in these cells, including genes like Il-6, Tgfβ1, Ccl2, Tnfα, Vegf-A, Cd38, Reg genes, and Hgf using real-time RT-PCR. In rat H9c2 cells, Tgfβ1, Ccl2, Tnfα, Flt-1, Reg IV, and Hgf increased significantly with IH, as shown in Figure 4. In contrast, only Reg IV and Hgf mRNAs were significantly increased by IH in mouse P19.CL6 cells (Fig. 4). ELISA revealed that IH significantly raised Reg IV and Hgf protein levels in P19.CL6 cell media (Reg IV: 30.16 pg/mL vs. 91.83 pg/mL, p = 0.0025, and Hgf: 101.9 pg/mL vs. 106.3 pg/mL, p = 0.0046) (Fig. 5).

Fig. 4. The mRNA levels of rat Il-6, Il-33, Il-17A, Il18, Tgfβ1, Cxcl12, Tnfα, Ccl2, Vegf-A, Flt-1, Flk-1, Cd38, Reg I, PAP I, PAP II, PAP III, Reg IV, Extl3, Hgf, and c-Met in rat H9c2 cardiomyocytes (Takasawa S, Itaya-Hironaka A, et al., 2022).

Fig. 5. The mRNA levels of mouse Il-6, Il-8, Il-17A, Il-18, Tgfβ1, Ccl2, Cxcl12, Tnfα, Vegf-A, Flt-1, Flk-1, Cd38, Reg I, Reg II, Reg IIIα, Reg IIIβ, Reg IIIγ, Reg IIIδ, Reg IV, Extl3, Hgf, and c-Met in mouse P19.CL6 cardiomyocytes (Takasawa S, Itaya-Hironaka A, et al., 2022).

Fig. 6. Concentrations of (A) Reg IV and (B) Hgf in a mouse P19.CL6 cardiomyocyte culture medium (Takasawa S, Itaya-Hironaka A, et al., 2022).