Cyp2j5-Gene Deletion Affects on Acetylcholine and Adenosine-Induced Relaxation in Mice: Role of Angiotensin-II and CYP-Epoxygenase Inhibitor

Abstract

Previously, we showed vascular endothelial overexpression of human-CYP2J2 enhances coronary reactive hyperemia in Tie2-CYP2J2 Tr mice, and eNOS −/− mice had overexpression of CYP2J-epoxygenase with adenosine A 2A receptor-induced enhance relaxation, but we did not see the response in CYP2J-epoxygenase knockout mice. Therefore, we hypothesized that Cyp2j5-gene deletion affects acetylcholine-and 5'-Nethylcarboxamidoadenosine (NECA) (adenosine)-induced relaxation and their response is partially inhibited by angiotensin-II (Ang-II) in mice. Acetylcholine (Ach)-induced response was tested with N-(methylsulfonyl)-2-(2-propynyloxy)-benzenehexanamide (MS-PPOH, CYP-epoxygenase inhibitor; 10 −5 M) and Ang-II (10 −6 M). In Cyp2j5 −/− mice, ACh-induced relaxation was different from C57Bl/6 mice, at 10 −5 M (76.1 ± 3.3 vs. 58.3 ± 5.2, P < 0.05). However, ACh-induced relaxation was not blocked by MS-PPOH in Cyp2j5 −/− : 58.5 ± 5.0%, P > 0.05, but blocked in C57Bl/6: 52.3 ± 7.5%, P < 0.05, and Ang-II reduces AChinduced relaxation in both Cyp2j5 −/− and C57Bl/6 mice (38.8 ± 3.9% and 45.9 ± 7.8, P <0.05). In addition, NECA-induced response was tested with Ang-II. In Cyp2j5 −/− mice, NECA-induced response was not different from C57Bl/6 mice at 10 −5 M (23.1 ± 2.1 vs. 21.1 ± 3.8, P > 0.05). However, NECA-induced response was reduced by Ang-II in both Cyp2j5 −/− and C57Bl/6 mice (−10.8 ± 2.3% and 3.2 ± 2.7, P < 0.05). Data suggest that ACh-induced relaxation in Cyp2j5 −/− mice depends on nitric oxide (NO) but not CYPepoxygenases, and the NECA-induced different response in male vs. female Cyp2j5 −/− mice when Ang-II treated.

Adult mouse fibroblasts retain organ-specific transcriptomic identity

“…Among the top 10 upregulated genes in transplanted fibroblasts, 6 were shared by the ectopically transplanted HFs and TFs, including the serum amyloid A Saa3, secreted during the acute phase of inflammation ( Forte et al, 2020 ); two metabolic enzymes, Slc27a2, primarily expressed in kidney and liver involved in lipid biosynthesis and fatty acid degradation, and Pck1 key regulator of gluconeogenesis; the cytochrome gene Cy2j5 involved in vasorelaxation ( Agba et al, 2020 ); the kidney abundant protein Kap, androgen-regulated, proximal tubule-specific not expressed at detectable levels in tissue other than the kidney ( Toole et al, 1979 ) , Fut9 a fucosyltransferase with the highest expression in adult pancreas, placenta, kidney ( Figure 7k ). In summary, while transplanted fibroblasts maintained their core identity, they responded to the kidney microenvironment by expressing a subset of kidney-specific genes, modulating positional code genes and activating common and cell-specific pathways in the attempt to adapt to the new, more hypoxic condition.…”

“…The differential regulation of Hoxd10 and Hoxd3 may suggest the acquisition of a cortex-like phenotype by transplanted KFs. KEGG analysis of the genes modulated in response to the transplant showed upregulation of pathways related to fibrosis and damage response (TNF signaling, ECM-receptor interaction, protein digestion and absorption) and downregulation of oxidative phosphorylation, steroid biosynthesis, and p53 signaling among the common regulated genes (Figure 7l Among the top 10 upregulated genes in transplanted fibroblasts, 6 were shared by the ectopically transplanted HFs and TFs, including the serum amyloid A Saa3, secreted during the acute phase of inflammation [22]; two metabolic enzymes, Slc27a2, primarily expressed in kidney and liver involved in lipid biosynthesis and fatty acid degradation, and Pck1 key regulator of gluconeogenesis; the cytochrome gene Cy2j5 involved in vasorelaxation [61]; the kidney abundant protein Kap, androgen-regulated, proximal tubule-specific not expressed at detectable levels in tissue other than the kidney [62],…”

Adenosine A2A receptor and vascular response: role of soluble epoxide hydrolase, adenosine A1 receptor and angiotensin-II

“…Mice were sacked with pentobarbital sodium (100 mg/kg ip) and extracted their aortas to study protein expression as per our previously published protocol [2,3,12,14,30,[38][39][40][41]. In brief, aortas extracted from both A 2A AR −/− and C57Bl/6 mice were treated with 1 ml of lysis buffer for protein extraction [14].…”

“…In brief, aortas extracted from both A 2A AR −/− and C57Bl/6 mice were treated with 1 ml of lysis buffer for protein extraction [14]. Gel electrophoresis and Western blot analysis were done according to the protocol described by us [2,3,12,14,30,38]. Following the blocking with nonfat dry milk, the nitrocellulose membranes were incubated with monoclonal and polyclonal primary antibodies raised against for A 1 AR, sEH, CYP4A, CYP2C and β-actin [14].…”

Adult fibroblasts retain organ-specific transcriptomic identity

“…Interestingly, Cardiac Hypertrophy Signaling (which include many pro-fibrotic signals like AngII, TGFb, IGF1), and HIF1a signaling were predicted to be downregulated in KFs and upregulated in HFs and TFs, possibly inferring a better resilience of KFs to the kidney capsule environment. Among the top 10 upregulated genes in transplanted fibroblasts, 6 were shared by the ectopically transplanted HFs and TFs, including the serum amyloid A Saa3, secreted during the acute phase of inflammation [22]; two metabolic enzymes, Slc27a2, primarily expressed in kidney and liver involved in lipid biosynthesis and fatty acid degradation, and Pck1 key regulator of gluconeogenesis; the cytochrome gene Cy2j5 involved in vasorelaxation [61]; the kidney abundant protein Kap, androgen-regulated, proximal tubule-specific not expressed at detectable levels in tissue other than the kidney [62], Fut9 a fucosyltransferase with the highest expression in adult pancreas, placenta, kidney (Fig. 7m).…”