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. 2019 Mar 27:2019:5410108.
doi: 10.1155/2019/5410108. eCollection 2019.

The Role of Epoxyeicosatrienoic Acids in Diabetes Mellitus-Induced Impaired Vascular Relaxation of Aortic Rings in Ovariectomized Sprague-Dawley Rats

Dragan Manojlović  1 , Ana Stupin  2   3 , Anita Matić  2 , Zrinka Mihaljević  2 , Sanja Novak  2 , Ines Drenjančević  2
Affiliations

The Role of Epoxyeicosatrienoic Acids in Diabetes Mellitus-Induced Impaired Vascular Relaxation of Aortic Rings in Ovariectomized Sprague-Dawley Rats

Dragan Manojlović et al. Int J Endocrinol. .

Abstract

Aim: The present study was aimed at determining if type 1 diabetes mellitus (DM) affects vascular function and at elucidating the mechanisms mediating vasorelaxation in both nonovariectomized and ovariectomized Sprague-Dawley (SD) rats.

Materials and methods: Eighty female SD rats were divided into four groups: nonovariectomized healthy (non-OVX-CTR) and diabetic (non-OVX-DM) rats and ovariectomized healthy (OVX-CTR) and diabetic (OVX-DM) rats. Bilateral ovariectomy was performed at the age of 5 weeks, and type 1 DM was induced by streptozotocin at the age of 6 weeks. At the age of 12 weeks, acetylcholine-induced relaxation (AChIR) was assessed in aortic rings in the absence/presence of L-NAME, Indomethacin, and MS-PPOH. Aortic tissue mRNA expression of eNOS, iNOS, COX-1, COX-2, thromboxane synthase 1 (TBXAS1), CYP4A1, CYP4A3, and CYP2J3, as well as plasma oxidative stress, was measured.

Results: AChIR did not differ in non-OVX-DM rats compared to non-OVX-CTR ones. AChIR was significantly reduced in the OVX-DM group compared to the OVX-CTR group. MS-PPOH did not reduce AChIR in OVX-DM rats as it did in OVX-CTR ones. CYP4a3 mRNA expression in OVX-DM rats was significantly lower compared to that in the OVX-CTR group.

Conclusions: Female sex hormones may protect vasorelaxation in type 1 diabetic rats. Type 1 diabetes impairs vasorelaxation in response to ACh in ovariectomized rats (but not in nonovariectomized rats) by affecting vasorelaxation pathways mediated by EETs.

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Figures

Figure 1
Figure 1
ACh-induced relaxation (AChIR) of isolated rat aorta rings in non-OVX-CTR and non-OVX-DM rats and OVX-CTR and OVX-DM rats. AChIR was significantly impaired in the OVX-DM group when compared to the non-OVX-DM group of rats at 10−7-10−5 M ACh concentration. Both non-OVX-DM and OVX-DM rats exhibited lower sensitivity to ACh compared to their corresponding controls (table within the panel). AChIR was significantly impaired in OVX-CTR compared to non-OVX-CTR (at 10−7-10−5 M ACh concentration) and OVX-DM compared to non-OVX-DM groups of rats (at 10−7-10−5 M ACh concentration). Both OVX-CTR and OVX-DM rats exhibited lower sensitivity to ACh compared to non-OVX groups, respectively (table within the panel). LogEC50 values (shown in the corresponding table) were compared by Student's t-test. ∗P < 0.05: OVX-CTR vs. OVX-DM, OVX-DM vs. non-OVX-DM. †P < 0.05: OVX-CTR vs. non-OVX-CTR. ‡P < 0.05: non-OVX-CTR vs. non-OVX-DM, OVX-DM vs. non-OVX-DM. §P < 0.05: OVX-CTR vs. OVX-DM, OVX-CTR vs. non-OVX-CTR. Ach concentration: 10−9 to 10−5 mol L−1. N: number of aortic rings. EC50 (mol L−1): half maximal effective concentration presents the concentration of ACh (mol L−1) which induces a response halfway between the baseline and maximum.
Figure 2
Figure 2
Mechanisms of acetylcholine-induced relaxation (AChIR) response of isolated rat aorta rings in non-OVX-CTR and non-OVX-DM rats. (a) and (b) present the relaxation of isolated aortic rings in response to ACh (presented as log [ACh] of ACh concentration (10−9 to 10−5 M)) in the non-OVX-CTR (a) and non-OVX-DM (b) groups of rats. The presence of L-NAME and MS-PPOH significantly reduced the AChIR of isolated rat aortic rings in the non-OVX-CTR group (a) and non-OVX-DM group (b) of rats. Indomethacin administration did not have any significant effects on AChIR in both the non-OVX-CTR (a) and non-OVX-DM (b) groups of rats. Data were compared by two-way ANOVA and Bonferroni post hoc tests. Sensitivity to ACh in the presence of L-NAME and MS-PPOH was significantly decreased compared to the basic response or response to ACh in the presence of Indomethacin in both the non-OVX-CTR and non-OVX-DM groups of rats. LogEC50 values were compared by one-way ANOVA followed by Holm-Sidak pairwise multiple comparison. Statistically significant (P < 0.05) AChIR in the presence of L-NAME (∗) or MS-PPOH (†) compared to the baseline ACh response. P < 0.05: ‡L-NAME vs. baseline, Indomethacin, and MS-PPOH; §MS-PPOH vs. baseline and Indomethacin. Concentrations—ACh: 10−9 to 10−5 mmol L−1, L-NAME: 3 × 10−4 mmol L−1, Indomethacin: 10−5 mmol L−1, and MS-PPOH: 10−5 mmol L−1. N: number of aortic rings. EC50 (mol L−1): half maximal effective concentration presents the concentration of ACh (mol L−1) which induces a response halfway between the baseline and maximum.
Figure 3
Figure 3
Mechanisms of the acetylcholine-induced relaxation (AChIR) response of isolated rat aorta rings in OVX-CTR and OVX-DM rats. (a) and (b) present the relaxation of isolated aortic rings in response to ACh (presented as log [ACh] of ACh concentration (10−9 to 10−5 M)) in the OVX-CTR (a) and OVX-DM (b) groups of rats. The presence of L-NAME and MS-PPOH significantly reduced the AChIR of isolated rat aortic rings in the OVX-CTR group (a), and only the presence of L-NAME significantly reduced the AChIR in the OVX-DM group (b) of rats. Indomethacin administration did not have any significant effects on AChIR in both the OVX-CTR (a) and OVX-DM (b) groups of rats. Data were compared by two-way ANOVA and Bonferroni post hoc tests. Sensitivity to ACh in the presence of L-NAME and MS-PPOH was significantly decreased compared to the basic response or response to ACh in the presence of Indomethacin in OVX-CTR, while in OVX-DM, sensitivity to ACh in the presence of L-NAME was significantly decreased compared to the basic response or response to ACh in the presence of Indomethacin or MS-PPOH. LogEC50 values were compared by one-way ANOVA followed by Holm-Sidak pairwise multiple comparison. Statistically significant (P < 0.05) AChIR in the presence of L-NAME (∗) or MS-PPOH (†) compared to the baseline ACh response. P < 0.05: ‡L-NAME vs. baseline, Indomethacin, and MS-PPOH; §MS-PPOH vs. baseline and Indomethacin. Concentrations—ACh: 10−9 to 10−5 mmol L−1, L-NAME: 3 × 10−4 mmol L−1, Indomethacin: 10−5 mmol L−1, and MS-PPOH: 10−5 mmol L−1. N: number of aortic rings. EC50 (mol L−1): half maximal effective concentration presents the concentration of ACh (mol L−1) which induces a response halfway between the baseline and maximum.
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