Angiotensin type 1 receptor mediates renal production and conversion of prostaglandins E 2 to F 2 α in conscious diabetic rats

Introduction

Pathogenesis of the renal complications in diabetes is multifaceted and not completely understood.^1–3 Evidence supports a critical role for renin–angiotensin system (RAS) activation in the diabetes-related disturbances in renal microcirculation, inflammation and salt balance.^4–10 Inappropriate and uncontrolled activation of the local paracrine RAS sets in motion a series of turbulences in several critical cellular processes which can contribute to the pro-inflammatory changes seen in diabetes. ⁹ One of these cellular processes affected by hyperglycemia and RAS activation is the COX-derived prostaglandins generation.^3,11–13 Among prostaglandins, prostaglandin E₂ (PGE₂) in particular has a powerful pro-inflammatory effect and has been tightly linked to RAS activation.^5,14–25

Previous studies demonstrated that under normal conditions (normal salt and no diabetes), the activity of RAS is minimal, with no effects for angiotensin receptor blockers on basal prostaglandins generation.^22,23,25 However, under low salt condition, we demonstrated that AT₁R mediates PGE₂ generation^22,26 and, in turn, PGE₂ stimulates renin release and angiotensin II formation.^27–29 This crosstalk between AT₁R and PGE₂ is buffered by the enzyme 9-keto reductase, which increases the conversion of PGE₂ to prostaglandins F_2α (PGF_2α), a prostaglandin metabolite that has an inhibitory effect on RAS.^{22–26,30,31} The effects of chronic hyperglycemia on the interaction between RAS and prostaglandins E₂ and F_2α have not been described. We hypothesized that in diabetes, an aberrant AT₁R activity enhances the biosynthesis of PGE₂ and impairs the activity of the enzyme PGE₂ 9-ketoreductase, leading to reduction in the inactivation of PGE₂ to PGF_2α.

Materials and methods

Results

Blood glucose at baseline was 86±4.5mg/dl and increased to 362±38 mg/dl five days after receiving STZ (p<0.001). After the induction of diabetes, RIF Ang II recovery levels increased significantly from 0.3±0.02 pg/min to 0.4 ± 0.02 pg/min (p=0.013 vs. baseline) at the third week and to 0.5±0.01 pg/min (p< 0.001 vs. baseline) at the sixth week (Figure 1). There were significant increases in the levels of RIF PGE₂, from 17.5±1.4 pg/min to 76.3±14.5 pg/min at three weeks (p< 0.01) and to 133.2±13.1 pg/min (p<0.001 vs. baseline, p<0.01 vs. three weeks of diabetes) at six weeks after the onset of diabetes (Figure 2). Similarly, PGF_2α increased from 5.7±0.9 pg/min under normoglycemia to 51.7±3.8 pg/min at three weeks (p< 0.001) and to 60.5±12.4 pg/min at six weeks (p< 0.001 vs. baseline) after the onset of diabetes (Figure 3). RIF PGE₂ and PGF_2α increased 3- and 9-fold respectively by the third week, and 6- and 11-fold by the sixth week respectively. The ratio of PGF_2α/PGE₂ ratio (Figure 4) initially increased from 0.3 at baseline to 0.7 by the third week of diabetes. At six weeks of diabetes, the PGF_2α/PGE₂ ratio was reduced to 0.4, suggesting a decline in activity of the enzyme PGE 9-ketoreductase.

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Figure 1.

Renal interstitial fluid angiotensin II in conscious rats at baseline and with the development of diabetes (DM), in response to valsartan (n=8; solid line) or placebo treatment (n=8; dashed line).

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Figure 2.

Renal interstitial fluid prostaglandins E₂ (PGE₂) in conscious rats at baseline and with the development of diabetes (DM), in response to valsartan (n=8; solid line) or placebo treatment (n=8; dashed line).

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Figure 3.

Renal interstitial fluid prostaglandins F_2α (PGF_2α) in conscious rats at baseline and with the development of diabetes, in response to valsartan (n=8; solid line) or placebo treatment (n=8; dashed line).

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Figure 4.

Renal interstitial fluid prostaglandins E₂:F₂ (PGE₂:PGF_2α) ratio in conscious rats at baseline and with the development of diabetes (DM), in response to valsartan (n=8; solid line) or placebo treatment (n=8; dashed line).

Under normal glycemic conditions, treatment with valsartan did not significantly affect the RIF Ang II, PGE₂ and PGF_2α levels. The administration of valsartan to the diabetic animals led to a significant increase in the level of Ang II, to 0.5 ± 0.03 pg/min (p< 0.001 vs. untreated three weeks of diabetes) at the third week, and to 0.7±0.05 pg/min (p< 0.01 vs. untreated six weeks of diabetes) at the sixth week. Valsartan treatment also led to a significant decrease in the levels of RIF PGE₂, to 35.4±6.8 pg/min at the third week (p< 0.05 vs. untreated three weeks of diabetes) and to 32.6±8.4 pg/min at the sixth week (p< 0.001 vs. untreated six weeks of diabetes) of diabetes (53%, 75% respectively). Similarly, valsartan led to a significant decrease in the level of RIF PGF_2α, to 21.1 + 4.1 pg/min at the third week (p< 0.05 vs. untreated three weeks of diabetes) and to 23.2±5.0 at the sixth week (p< 0.01 vs. untreated six weeks of diabetes) of diabetes (59.3%, 61.6% respectively). Valsartan increased the PGF_2α/PGE₂ ratio from 0.4 to 0.7 at week 6 of diabetes.

Discussion

Accumulating evidence supports a close relationship between the prostaglandin system and the RAS in the kidney. Both systems are also strongly linked to the development of inflammation and the pathogenesis of diabetes complications. However, the interface between prostaglandin and RAS in diabetes remained unclear and mechanistically vague. Prior studies have been hindered by the difficulty in continuous monitoring of early and late paracrine changes in tissues of diabetic animals. The use of the microdialysis technique allowed us to sample in real time and longitudinally in the same, live, and conscious animals the interaction between renal cortical angiotensin II, PGE₂ and PGF_2α before and after induction of diabetes and in response to AT₁R blockade. Here, we demonstrate a role for AT₁R in the increased biosynthesis of PGE₂ and PGF_2α in diabetes and on the conversion of PGE₂ to PGF_2α.

In alignment with previous reports^22,26 under normal conditions, RAS activity is minimal with no effects of AT₁R blockade on Ang II, PGE₂ or PGF_2α levels. Also analogous to our previous report under low salt conditions, we have seen activation of the RAS in early diabetes with a rise in Ang II generation and signaling through AT₁R that mediated the increase in prostaglandins generation. Our current study further clarified the effects of chronic activation of RAS on prostaglandins release and the impact of this on the regulation of prostaglandins–RAS crosstalk. The observed increased generation of PGE₂ and reduction in conversion rate of PGE₂ to PGF_2α in diabetes provides a mechanistic explanation of previous reports that suggested an interaction between renin and prostaglandins^36–38 and reveals a pathological positive feedback loop with the RAS (increased biosynthesis of prostaglandins and reduced conversion of the RAS-enhancer PGE₂ to the RAS inhibitor PGF_2α) in diabetes. The increase in the level of PGE₂ by the third week of diabetes was associated with a concordant increase in the level of PGF_2α in the kidney. Interestingly, while the conversion of PGE₂ to PGF_2α was initially enhanced by hyperglycemia, this rate of conversion declined by the sixth week of diabetes despite increasing levels of PGE₂. This paradoxical early increase in the conversion rate may result from either the increase in PGE₂ synthesis or alteration in PGE₂ metabolism by 9-ketoreductase or both. A reduction in PGE₂ 9-ketoreductase activity, however, could only explain the later decline in the rate of conversion of the powerful pro-inflammatory, renin enhancer PGE₂ to the less active PGF_2α in diabetes, despite increased generation of PGE_2. The restoration of this conversion rate with valsartan treatment supports the notion that in diabetes, AT₁R has an inhibitory effect on PGE₂ 9-ketoreductase activity.

In the present study we observed a counter regulatory upregulation of RIF Ang II with valsartan treatment in diabetes. Previous studies suggested that treatment with AT₁R antagonists are associated with a rise in plasma Ang II concentration as the result of inhibition of the AT₁R-mediated negative feedback on renin release.^39–42 Previous studies have also suggested that the resultant increased Ang II stimulates AT₂R. ⁴³ Our current study suggests a role for AT₁R in reducing the activity of PGE₂ 9-ketoreductase with the progression of diabetes. We have observed an initial increase in PGE₂ 9-ketoreductase activity that was followed by decline as diabetes progressed; a potential role for the anti-inflammatory AT₂R cannot be excluded and may explain the initial increase in the rate of conversion of PGE₂ to PGF_2α at three weeks of diabetes (through an intact AT₂R). With the progression of diabetes, increase in Ang II and AT₁R activity, a failure in AT₂R may propagate the reduced activity of PGE₂ 9-ketoreductase. It is tempting also to speculate that the increased rate of conversion of PGE₂ to PGF_2α observed with valsartan treatment during the third week of diabetes is mediated through enhancing the activity of the unopposed and functional AT₂R. We previously reported a direct role for AT₂R in modulating the activity of PGE 9-ketoreductase.^22,26 It is possible that the use of AT₁R blockade enhances the AT₂R function in diabetes. ⁴⁴ In addition, the observed increase in Ang II levels in diabetes could result from the less opposed inhibitory action of PGF2α on renin release.^30,31

Conclusion

The current study suggests that in diabetes, Ang II, through AT₁R, plays a critical role in generation and conversion of PGE₂ to PGF_2α. With progression of diabetes, PGE₂ to PGF_2α conversion is impaired, leading to further accumulation of the potent pro-inflammatory, renin enhancer PGE₂ and the establishment of a vicious cycle between RAS and prostaglandins systems. Given the strong link between RAS, prostaglandins and their downstream signaling pathways mediating angiogenesis, cell adhesion, motility, invasion, morphology, vascular permeability and fibrosis, elucidating the mechanisms involved in the interaction between the arachidonic acid and angiotensin systems may help fine tune future strategies for management of diabetes complications.