Abstract
In various models of chronic kidney disease, the amount and localization of Cx43 in the nephron is known to increase, but the intracellular pathways that regulate these changes have not been identified. Therefore, we proposed that: “In the model of renal damage induced by infusion of angiotensin II (AngII), a RhoA/ROCK-dependent pathway, is activated and regulates the abundance of renal Cx43”. In rats, we evaluated: 1) the time-point where the renal damage induced by AngII is no longer reversible; and 2) the involvement of a RhoA/ROCK-dependent pathway and its relationship with the amount of Cx43 in this irreversible stage. Systolic blood pressure (SBP) and renal function (urinary protein/urinary creatinine: Uprot/UCrea) were evaluated as systemic and organ outcomes, respectively. In kidney tissue, we also evaluated: 1) oxidative stress (amount of thiobarbituric acid reactive species), 2) inflammation (immunoperoxidase detection of the inflammatory markers ED-1 and IL-1β), 3) fibrosis (immune detection of type III collagen; Col III) and 4) activity of RhoA/ROCK (amount of phosphorylated MYPT1; p-MYPT1). The ratio Uprot/UCrea, SBP, oxidative stress, inflammation, amount of Cx43 and p-MYPT1 remained high 2 weeks after suspending AngII treatment in rats treated for 4 weeks with AngII. These responses were not observed in rats treated with AngII for less than 4 weeks, in which all measurements returned spontaneously close to the control values after suspending AngII treatment. Rats treated with AngII for 6 weeks and co-treated for the last 4 weeks with Fasudil, an inhibitor of ROCK, showed high SBP but did not present renal damage or increased amount of renal Cx43. Therefore, renal damage induced by AngII correlates with the activation of RhoA/ROCK and the increase in Cx43 amounts and can be prevented by inhibitors of this pathway.
Highlights
In the last years Chronic kidney disease (CKD) has been increasing steadily worldwide turning into a public health problem
We first studied the time course of renal damage induced by angiotensin II (AngII), as well as the spontaneous recovery of the kidney from the damage induced by AngII
Consistent with previous results, the amount of Thiobarbituric Acid Reactive Substances (TBARS) (Figure 7C and D), the distribution of ED-1 (Figure 7E) and the amount of IL-1β (Figure 7F), collagen type III (Col III) immunoreactivity (Fig. 7G), amounts of p-myosin phosphatase target subunit 1 (MYPT-1) (Figure 7H) and Cx43 (Figure 7I) in the group treated with AngII for 6 weeks were high, but in kidneys from rats treated with AngII + Fasudil, the values for these parameters were similar to that detected in the kidneys of control rats
Summary
In the last years Chronic kidney disease (CKD) has been increasing steadily worldwide turning into a public health problem. Fasudil has been shown to inhibit the cascade of events that leads to the increase in expression of pro-inflammatory cytokines, extracellular matrix genes, OS, and macrophage infiltration Both RhoA and ROCK have been considered as therapeutic targets to prevent kidney damage and hypertension [19,22,24,25]. This opposed regulation of GJs and Cx43 HCs by pro-inflammatory factors occur in cultures of proximal tubule and mesangial cells treated with metabolic inhibitors or pro-inflammatory cytokines, where an increase in the activity of Cx HCs has been demonstrated [30,31] In pathological conditions such as hypertension, the amount of renal Cxs is altered. Based on the evidence described above, in this study we evaluated whether the RhoA/ROCK pathway is activated and regulates the abundance of Cx43, which might contribute to renal tissue damage in the model of renal damage induced by infusion of AngII
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