In this study we suggested that mercury exposure accelerated the hypertension development in young SHR animals. This behavior seen in the mercury treated SHR appears to be caused by a reduced bioavailability of NO, which plays a crucial role in the blood pressure regulation decreasing its negative modulation by oxidative stress. These findings were reinforced by the plasma nitrite/nitrate reduction and the reduction of SBP produced by tempol in SHR after mercury exposure. Losartan reduced the SBP of all groups. In addition, after mercury exposure the mercury treated SHR group showed reduced ACE activity (Figure 7) and in a lesser extent the increment of SBP. These findings should reduce the accelerated hypertension development of mercury treated SHR group instead of increasing it. Therefore, the increase in SBP of SHR exposed group should be occuring by another mechanism involving oxidative stress and reduction of nitric oxide bioavailability.
Previous findings have shown that mercury exposure in normotensive rats, both acute and for 30 days, affects endothelial function without changing arterial blood pressure even though reducing NO bioavailability 24, 25, 31 increasing the production of free radicals.11, 22, 32, 33 However, in normotensive animals previous report showed that chronic Hg exposure for long periods (60 days) increases blood pressure.23, 34 Based on these findings we argued whether the association of mercury with predisposing hypertensive effects could accelerate the occurrence of hazardous consequences as increase of arterial blood pressure, oxidative stress and reduced NO modulation.
Acute exposure to small doses of mercury chloride causes elevation of SBP35,36 but exposure for 30 days does not change SBP in Wistar animals.22, 23, 25, 34, 36 In addition, even using high doses of chronically administered mercury chloride there is no interference with the systolic or diastolic blood pressure of adult Wistar rats.23, 34 These findings suggest that acute mercury administration in adult Wistar rats increases blood pressure while under chronic treatment at levels similar to those found in exposed individuals adaptive mechanisms restrain such blood pressure increment.
In fact, recently we showed that mercury exposure could accelerated the hypertension development of young SHR.26, 27 However, this effect was accompanied by a vasoprotective mechanism, which includes an increase of NO, against the early establishment of hypertension in resistance arteries of young SHR. Therefore, we evaluated the effect of mercury exposure in in vivo young SHR to elucidate how this metal changes blood pressure at the same time that vasoprotective mechanisms were occurring, as observed previously.27
Thus, we evaluated mercury effects by recording invasive and noninvasive arterial blood pressure. Our results showed an accelerated increase of the SBP (noninvasive recordings) in young SHR exposed to mercury chloride from the second week of treatment. This was not observed in the Wistar group. Invasive recordings after mercury exposure for 30 days also showed that SBP and DBP did not differ between control groups but SBP of SHR treated increased compared to SHR controls. These findings suggested that the mercury exposure of prehypertensive SHRs might influence the time course of hypertension development.
Previous reports showed that mercury exposure causes endothelial dysfunction, reduces NO production and increases oxidative stress in Wistar rats.22, 24, 25, 37 Similar findings of endothelial dysfunction have been reported in other models as the SHR strain.38-40 Therefore, our first attempt was to investigate the role of NO. Considering that NO acts relaxing vascular smooth muscle, we studied its bioavailability in animals exposed to Hg. It is known that eNOS expression is increased in young SHR (approximately 12 weeks)41, 42 and decreased in adult and elderly SHR rats (approximately 36 and 72 weeks, respectively).43 Our findings using L-NAME administration suggested that mercury chloride interfered with the bioavailability of NO at an earlier stage of animal life.
We also investigated plasma nitrate and nitrite as an indicator of NO production.44 The nitrite/nitrate concentration was higher, as expected,45 in the control group of young SHR rats compared with the Wistar control group. However, there was no difference between the treated and untreated Wistar groups (Figure 4). Moreover, only in the SHR group exposed to Hg, a reduction in nitrate/nitrite production was observed suggesting that mercury reduced NO production or increased its degradation (catabolism). Another possible mechanism already reported is the reduced NO production by endothelial damage produced by mercury using a similar treatment.46 This finding reinforced our results, a condition that might contribute to accelerate the increase of the SBP.
It is well established that Hg activates NADPHoxidase and COX consequently increasing free radicals production and producing vasoconstriction.13, 25, 26, 27 NO can react with the superoxide anion forming peroxynitrite, another vasoconstrictor.45 So, we evaluated the involvement of superoxide anion and oxidative stress in animals exposed to Hg. Our findings showed that both Wistar groups, Hg-treated and untreated, had a decrease in systolic blood pressure when treated with Tempol (Figure 5-A), but without difference between the groups. In SHRs, the decrease of SBP was shown only in the group exposed to HgCl2. These results suggest that the superoxide anion was contributing to increase the SBP in young SHR animals exposed to mercury for 30 days.
To evaluate the extent of oxidative stress, we investigated lipid peroxidation. It has also been reported that male SHR adult animals have an increase in oxidative stress.47 However, we did not observe an increase in the lipid peroxidation in the animals exposed to HgCl2 of both SHR groups. In young SHR animals, a previous study showed that they have an activated antioxidant capacity by superoxide dismutase and catalase,48 which explains our findings. Regarding adult Wistar rats, it has been reported in a 30-day mercury exposure that, at the end of the treatment, superoxide dismutase and catalase were higher when compared to the untreated group.49 These findings suggest that the chronic mercury administration to young SHR might activate mechanisms that reduce NO bioavailability and explains the accelerated increase of arterial blood pressure.22, 23, 35, 37
Another mechanism that generates oxidative stress and contributes to increase arterial blood pressure is the renin angiotensin system. Its product, angiotensin II, interacts with AT 1 receptors and increases blood pressure. Therefore, we investigated the modulation of these receptors in animals exposed to Hg. When the animals were treated with Losartan, a decrease of SBP was observed in all groups. However, it was noticed that the decrease of SBP was smaller in the animals exposed to HgCl2.
However, we should emphasize that when comparing untreated rats from both groups ACE activity was increased in the SHRs. This result suggested that ACE activity was interfering only in the young SHRs. Based on these findings it was possible to postulate that in young SHRs the renin angiotensin system is involved in the increase of arterial pressure. In the SHR model in animals with previously established hypertension, we demonstrate that ACE activity levels increase with exposure to mercury.50 However, before establishing hypertension in SHRs the metal exposure caused the opposite effect, a reduction in this enzyme activity. Mercury exposure accelerated the increase of SBP in SHR although the percentage of the losartan effect was reduced and ACE activity was also reduced in this exposed group. Therefore, the enhanced increase in SBP of SHR group exposed to Hg, should be occuring by a mechanism involving oxidative stress and reduction of nitric oxide. So, is there then a link between oxidative stress and the renin-angiotensin system in such conditions? Recently, Saleem et al., (2016)51 suggest a mechanism that might explain such possibility. Superoxide, but not H2O2, acting via Sp3 transcription factor up-regulates angiotensin II AT1 receptor expression and function in human kidney-2 cells. Rizzetti et al., (2018)24 also reported that Hg exposure might play a role upregulating AT-1 receptors by increasing oxidative stress and reducing NO bioavailability.