EVIDENCE FOR PRO-HYPERTENSIVE, PRO-INFLAMMATORY EFFECT OF INTERLEUKIN-10 DURING CHRONIC HIGH SALT INTAKE IN THE CONDITION OF ELEVATED ANGIOTENSIN II LEVEL

Abstract

Interleukin-10 (IL-10) has been suggested to play a protective role in angiotensin II (AngII) induced cardiovascular disorders. This study examined the role of endogenous IL-10 in salt-sensitive hypertension (SSH) and renal injury induced by AngII. Responses to chronic AngII (400 ng/min/kg bw; osmotic minipump) infusion were evaluated in IL-10 gene knockout (IL-10KO) mice, fed with either normal (NS; 0.3% NaCl) or high (HS; 4% NaCl) salt diets and these responses were compared to those in wild type (WT) mice. NS or HS diets were given alone for the first 2 weeks and then with AngII treatment for additional 2 weeks (n=6 in each group). Arterial pressure was continuously monitored by implanted radio-telemetry and 24 hour (hr) urine collection was performed by metabolic cages on the last day of the experimental period. Basal mean arterial pressure (MAP) was lower in IL-10KO than in WT (98±3 vs 113±3, mmHg) mice. MAP responses to NS/HS alone or to the AngII+NS treatment were similar in both strains. However, the increase in MAP induced by the AngII+HS treatment was significantly lower in IL-10KO (15±5% vs 37±3%) compared to WT. Renal tissue eNOS expression (~3 folds) as well as urinary excretion of nitric oxide (NO) metabolites, nitrate/nitrite (1.2±0.1 vs 0.2±0.02, μM/24 hr) were higher in IL-10KO compared to WT. These results indicate that an increase in NO production helps to mitigate SSH induced by AngII and suggest that a compensatory interaction between IL-10 and NO exists in modulating AngII induced responses during HS intake.

Introduction

It is generally known that the anti-inflammatory cytokines effectively down-regulate the generation of the pro-inflammatory cytokines.¹ The anti-inflammatory cytokine, interleukin 10 (IL-10) exerts immune down-regulating action on the generation of pro-inflammatory cytokines such as interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α).^{2, 3} Endogenous IL-10 has been suggested to play a role as a key mediator of vascular protection in atherosclerosis and diabetes.^{4, 5} It is also suggested that IL-10 serves a protective role in Angiotensin II (AngII) induced hypertension and its cardiovascular complications.^{6, 7} AngII chronically induces salt-sensitive hypertension (SSH) and also activates multiple inflammatory mechanisms leading to injury of various organs including the kidney.⁸ However, the role of endogenous IL-10 formation in the pathogenesis of AngII induced SSH and associated renal injury is not yet clearly defined.⁹

Although high salt (HS) intake alone induces no or minimal changes in blood pressure, it exaggerates the hypertensive and renal injury responses to elevated AngII levels.^{8, 10} The mechanisms for such exaggerated responses remain unresolved.⁹ Although HS diet alone does not alter systolic blood pressure, it is able to cause mesangial expansion and kidney fibrosis, without eliciting cell proliferation.⁸ However, a HS diet given in chronic AngII-infused rats leads to greater levels of systolic blood pressure along with marked exacerbation of mesangial expansion, kidney fibrosis, and tubular epithelial cell proliferation.¹¹ These findings could indicate the involvement of pro-inflammatory cytokines, particularly TNF-α in these renal inflammatory responses.¹² AngII infusion in rats subjected to a HS diet causes increases in inflammatory cell infiltration in the tubulointerstitial areas.¹¹ Thus, it is suggested that the production of TNF-α induced by the elevated AngII level is involved in the exaggerated hypertensive and renal injury responses to combined AngII and HS intake.⁹ However, as a modulator of the pro-inflammatory cytokines, the contribution of anti-inflammatory cytokines, particularly the role of IL-10 has not been assessed in such conditions in any previous studies.^{2, 9}

In the present study, we hypothesized that endogenous IL-10 plays a protective role in hypertension and renal injury induced by HS intake in elevated AngII condition by minimizing the production of pro-inflammatory cytokines such as IL-6 and TNF-α. To examine this hypothesis, experiments were conducted to evaluate the changes in blood pressure, pro-inflammatory cytokine (TNF-α, IL-6) levels and renal injury parameters in response to chronic administration of AngII in IL-10 gene knockout (IL-10KO) and wild-type (WT) mice which were fed either normal or HS containing diets. These experiments were primarily conducted to achieve two main aims: a) to determine the extent of salt-sensitive hypertension and associated renal injury parameters (glomerular sclerosis and interstitial fibrosis) induced by AngII treatment in the condition of IL-10 deficiency and b) to determine the impact of endogenous IL-10 deficiency on TNF-α and IL-6 levels in response to HS/AngII treatment.

Materials and Methods

The experiments were performed in accordance with the guidelines and practices established by the Tulane University Animal Care and Use Committee. Male IL-10KO (B6.129P2-Il10tm1Cgn/J; stock no: 002251) mice and their genetic background WT control (C57BL/6J; stock no: 000664) mice (Jackson Laboratory, Bar Harbor, ME) were used in this study. These mice were housed in a temperature- and light-controlled room and allowed free access to standard diet (Ralston-Purina, St. Louis, MO) and tap water. The mice (8–9 wk of age; ∼25 g body wt) were randomly divided into different groups depending on intake of normal salt diet (NS; standard diet containing 0.3% NaCl) and high salt diet (HS, 4% NaCl; Harlan-Teklad, Madison, WI) respectively, comprised of 6 animals in each group. The groups are as follows:

1. NS-WT: Wild type mice given NS diet

2. HS-WT: Wild type mice given HS diet

3. NS-IL-10KO: IL-10KO mice given NS diet

4. HS-IL-10KO: IL-10KO mice given HS diet

These mice were fed on NS or HS diets alone for 2 weeks prior to AngII infusion (400 ng/min/kg body weight, bw; osmotic mini-pumps) for another 2 weeks. Blood pressure in these mice was continuously monitored by telemetry implantation. Urine was collected for 24 hour (hr) using metabolic cages only at the last day of experimental period after the mice have been discontinued from the telemetry recordings. Urinary excretion of sodium and potassium were assessed by flame photometry.^{13, 14, 15} Urinary excretion rate of nitric oxide (NO) metabolites, nitrate/nitrite (U_NOxV) and oxidative stress marker, 8-isoprostane (U_IsoV) were determined by colorimetry¹⁰ and by enzyme immunoassay^{10, 15} respectively. At the end of the experimental period, all the animals were euthanized and blood samples were collected to assess the levels of TNF-α and IL-6 as conducted earlier.¹³ The kidneys were also removed and renal tissue homogenates were analyzed for TNF-α and IL-6 levels and endothelial nitric oxide synthase (eNOS) protein. The formalin-fixed paraffin-embedded kidney sections were analyzed for renal injury parameters, glomerulosclerosis using Periodic-acid-Schiff (PAS) staining^{8, 13, 16} and interstitial fibrosis using Gomori’s trichrome staining.^{14, 17} Detailed experimental methods are available in the On-line Data Supplement.

Statistical Analyses

All results are expressed as means ± SE. Statistical analysis was performed using Sigmastat software (Systat Software, Chicago, IL). Comparison of the responses within the same group as well as between the groups was conducted using the repeated-measures ANOVA and Dunnett multiple comparisons test. P ≤ 0.05 is considered as significant.

Results

Blood pressure response

The basal level of mean arterial pressure (MAP) was lower in IL-10KO mice than WT mice (98±3 vs 113±3, mmHg; P<0.05) as illustrated in Figure 1. HS intake alone for 2 weeks prior to AngII infusion did not cause any effects on MAP in either strain of mice (Fig. 1). Chronic AngII infusion caused similar increases in MAP in NS-WT and NS-IL-10KO mice (Absolute Δ, 26±6 vs 26±7, mmHg; Fig. 2A and Percent Δ, 26±8% vs 23±5%; Fig. 2B). However, AngII treatment in HS-IL-10KO mice induced a smaller increase in MAP (Absolute Δ, 16±5 vs 40±3, mmHg; Fig. 2A and Percent Δ, 15±5% vs 37±3%; Fig. 2B) compared to that in HS-WT mice. These responses are calculated from the baseline MAP values on the day, just prior to the start of AngII treatment. The reduction in AngII induced MAP response in HS-IL-10KO is similar in magnitude both in absolute (Fig. 2A) as well as in percent (Fig. 2B) terms compared to that in HS-WT. It was noted that MAP started to increase on the 3^rd day of AngII treatment which was similar in both IL-10KO and WT mice.

Figure 1.

Mean arterial pressure (MAP) responses in IL-10KO and WT mice fed with either normal (NS) or high salt (HS) diets before and during angiotensin II (AngII; 400 ng/min/kg bw) treatment. The values at 0 day represent basal values at the start of the experiment. n=6 in NS-WT; n=5 in HS-WT; n=5 in NS-IL-10KO; n=6 in HS-IL-10KO.

Figure 2.

A - Absolute changes in mean arterial pressure (MAP) in response to normal (NS) or high salt (HS) diets before and during angiotensin II (AngII; 400 ng/min/kg bw) treatment. ‘Basal’ represent the values at the start of experimental protocol; NS/HS represents the values just prior to the day of AngII mini-pump implantation; NS/HS+AngII represents the value on the last day of the experimental period. B - Percent changes in MAP due to AngII treatment; The percent responses are calculated considering the values on the day just prior to AngII infusion as ‘Baseline’ values. ‘AngII’ represent values on the last day of AngII treatment. *, P<0.05 vs NS/HS or Baseline value, #, P<0.01 vs HS-WT group; @, P<0.001 vs WT group.

Renal excretory responses

The values for excretory parameters in urine collected on the last day of the experimental period are given in Table S1 in the On-line Data Supplement section. The mean values of U_NOxV and U_ISOV are also given in Table S1 and are illustrated in Figure 3. IL-10KO groups showed higher U_NOxV (Fig. 3A) and U_ISOV (Fig. 3B) values than WT groups. HS groups generally showed higher values of these parameters in both the strains except that U_ISOV was not increased in HS-IL-10KO group which could be the effect of marked increase in NO production in that group. As identical NS or HS diets were given and the quantity of their food intake was similar both in WT and in IL-10KO, it is reasonable to conclude that dietary factor(s) other than salt content did not contribute to the differences in U_NOxV between these strains of mice.^{10, 18}

Figure 3.

Mean values of urinary excretion rates of nitrate/nitrite (U_NoxV; A) and 8-isoprostane (U_IsoV; B) at the last day of the experimental period with AngII (400 ng/min/kg bw) treatment. *, P<0.05 vs NS-WT; #, P<0.05 vs HS-WT; @, P<0.05 vs NS-IL-10 KO.

Renal eNOS expression

The images of western blot analysis for total eNOS protein in the renal tissue are shown in Figure 4A and their densitometer values are illustrated in Figure 4B. Protein expression of eNOS was higher both in NS-IL-10KO (~2 folds) and HS-IL-10KO (~3 folds) compared to NS-WT and HS-WT respectively. The density of the β-actin band was not different between samples, indicating equal loading in all lanes.

Figure 4.

Endothelial nitric oxide synthase (eNOS) enzyme protein expression in the renal tissue. A - Representative images of total eNOS expression. B - Mean densitometer value (arbitrary units) of eNOS expression. *, P<0.05 vs NS-WT; #, P<0.05 vs HS-WT; @, P<0.05 vs NS-IL-10KO. β-actin was used as loading control.

TNF-α and IL-6 levels

Figure 5 depicts plasma and renal tissue levels of TNF-α (Fig. 5 A&B) and IL-6 (Fig. 5 C&D) at the last day of experimental period. TNF-α level was higher in plasma (69±6 vs 34±4, pg/mL) and in renal tissue (208±15 vs 95±11, pg/mg protein) in HS-WT compared to NS-WT mice. On the other hand, TNF-α level was lower in plasma (30±3 vs 180±44, pg/mL) and in renal tissue (206±23 vs 277±62, pg/mg protein) in HS-IL-10KO compared to that in NS-IL-10KO mice. However, IL-6 level was lower in plasma (13±1 vs 26±6, pg/mL) and higher in renal tissue (148±29 vs 71±12, pg/mg protein) in HS-WT compared to that in NS-WT mice. IL-6 level was higher in plasma (16±2 vs 8±2, pg/mL) but lower in renal tissue (108±18 vs 198±34, pg/mg protein) in HS-IL-10KO, compared to that in NS-IL-10KO mice.

Figure 5.

Tumor necrosis factor-alpha (TNF-α) levels (A - in plasma; B - in renal tissue) and interleukin-6 (IL-6) levels (C - in plasma; D - in renal tissue) at the last day of the experimental period with AngII treatment (400 ng/min/kg bw). *, P<0.05 vs NS-WT; #, P<0.05 vs HS-WT; @, P<0.05 vs NS-IL-10KO.

Glomerular sclerosis

The representative images of the PAS-stained kidney sections providing the extent of glomerular sclerosis are given in Figure 6A and the percent of the sclerotic areas are illustrated in Figure 6B. The percent sclerotic area in NS-IL-10KO was higher than that in NS-WT mice (25±1% vs 14±2%). However, the percent sclerotic area was not statistically different between HS-WT and HS-IL-10KO mice (16±3 % and 22±3 %).

Figure 6.

The extent of renal injury (glomerular sclerosis and interstitial fibrosis) at the end of experimental period with AngII (400 ng/min/kg bw) treatment. A & C illustrates the representative photomicrographs, and B & D illustrates the mean values of the percent areas of sclerotic area and fibrotic area in the renal tissues respectively. *, P<0.05 vs NS-WT.

Renal interstitial fibrosis

The representative images of Gomori’s trichrome-stained sections providing the extent of renal interstitial fibrosis are given in Figure 6C and the percent of the fibrotic areas are illustrated in Figure 6B. The fibrotic area in the renal interstitium were higher in NS-IL-10KO (11±1% vs 8±1%) compared to NS-WT mice. There was no significant difference between the fibrotic area between HS-WT (10±1%) and HS-IL-10KO (11±0.4%) mice.

Discussion

It is generally known that chronic HS intake alone induces no or minimal changes in blood pressure in normal conditions but it exaggerates the hypertensive and renal injury responses to elevated AngII level in many rodent models.^8–10 In the present study, it has been demonstrated that such HS induced exaggerated hypertensive response (commonly termed as salt-sensitive hypertension, SSH) to chronic AngII administration (400 ng/min/kg bw) is very minimal, rather absent in IL-10KO mice compared to the corresponding blood pressure response in WT mice (Fig. 1 & 2). Although AngII induced increase in MAP during NS intake was similar in both the strains, the observed exaggerated MAP response to AngII administration with HS intake in WT mice, was virtually absent in IL-10 KO mice (Fig. 2 A&B). This finding indicates that the endogenous level of IL-10 provides a pro-hypertensive role in the condition of elevated AngII level during HS intake. Although AngII induced exaggerated hypertension during HS intake is mechanistically linked with the development of oxidative stress condition,^8–10 a pro-inflammatory process is also suggested recently in the pathophysiology of SSH and associated renal injury.⁹ The striking absence of AngII induced salt-sensitivity in IL-10KO mice in the present study indicate that endogenous IL-10 may also contribute to such pro-inflammatory process in the development of SSH. Although widely regarded as one of the anti-inflammatory cytokines, IL-10 has also been recognized for inducing pro-inflammatory reactions in many cellular functions.¹⁹ In the present study, it is also observed that combined AngII treatment with chronic HS intake generally down-regulated the formation of pro-inflammatory cytokines (TNF-α and IL-6) in the renal tissues of IL-10KO mice, as opposed to their increases during similar treatment in WT mice (Fig. 5). Thus, these findings indicate that the endogenous formation of IL-10 is involved in a pro-inflammatory response during chronic HS intake in the condition of elevated AngII level.

In the present study, it is noted that the basal MAP is lower in IL-10KO mice than that in WT mice. Such lower MAP in IL-10KO compared to WT mice was also reported earlier both in conscious²⁰ (recoded by radio-telemetry) and in anesthetized mice²¹ (recorded from carotid artery catheter). However, a previous study⁷ in conscious mice reported no differences in the basal MAP or the increases in MAP (recorded by tail-cuff plethysmography) in response to a high dose of AngII (~2.5 times higher than that used in the present study) treatment between IL-10KO and WT mice. Such difference in the findings in that study⁷ compared to the present and earlier studies^{20, 21} could be related to the differences in the blood pressure measuring techniques as well as the differences in the doses of AngII. Although the reason for a low MAP in IL-10KO was not clearly identified in earlier studies,^{20, 21} it was suggested to be a nonspecific effect of IL-10 gene ablation or could be due to lower blood volume associated with lower erythrocyte count observed in IL-10KO than that in WT.²¹ In the present study, however, it is observed that U_NOxV as well as the renal expression of eNOS was higher in IL-10KO indicating an enhanced NO production in this strain compared to WT. Such increase in NO production seems to have contributed to low MAP in IL-10KO mice. As total eNOS protein expression is high in IL-10KO mice, it is conceivable that an increase in U_NOxV (a marker for increased NOS activity) in this strain of mice is, at least in part, due to the increase in eNOS activity. We have considered to examine eNOS protein expression over other NOS isoforms as it was shown earlier that the NO derived from eNOS mainly regulates baseline vascular resistance, renal blood flow and modulates AngII induced hypertensive responses in mice.^{22, 23} Although both eNOS and nNOS isoforms are present in the renal tissue,²⁴ a contributing role of nNOS in the regulation of blood pressure is not very supportive as it has been observed that blood pressure in nNOS knockout mice is similar to that in WT mice.²³ It is also reported²² that the iNOS isoform is mostly absent or minimally present in the mice kidney. Thus, it is conceivable that an increase in NO production mostly via enhanced eNOS activity results in low MAP and the attenuated AngII induced hypertensive response in HS-IL-10KO mice. However, further investigation may be needed to determine the specific roles of other NOS isoforms in inducing NO production in the condition of IL-10 deficiency.

The findings in the present study may indicate an inhibitory role of IL-10 on the formation of NO as eNOS expression is upregulated in IL-10KO mice. However, such characterization of IL-10 induced effect on NO formation may not be as simple as it is also observed that NOS inhibition decreases IL-10 levels both in the plasma and in the renal tissue.¹³ Although the IL-10 level is not measured in the present study, we have observed earlier²⁵ that its’ level is lower in plasma but higher in renal tissue of eNOS knockout mice compared to that in WT mice. Previous in-vitro studies²⁶ also demonstrated that the NOS activity as well as NO production in uterine tissues from pregnant rats was inhibited by pre-incubation with IL-10 indicating a reciprocal relationship between IL-10 and NO production. The present study also demonstrates that endogenous IL-10 level regulates NO production as well as tissue eNOS expression, particularly in the kidney. Generally, it has been suggested from previous studies ^{27, 28, 29} that pro-inflammatory cytokine (s) released from activated T-cells contribute to the development of hypertension and the anti-inflammatory cytokine, particularly IL-10, released from T-regulatory cells helps to mitigate this hypertensive response.^{6, 30, 31} However, the findings in our present study do not support a direct role of IL-10 in the development of salt-sensitive hypertension, though an anti-hypertensive effect of IL-10 has been suggested earlier.^{6, 7, 31} It is to be emphasized here that the activation of T-cells produces a variety of pro-and anti-inflammatory cytokines, depending on the specific nature of the inflammatory insults. The specific impact of the production of these cytokines in the overall regulation of blood pressure is very complex, which is yet a subject of various investigations. The novel finding in the present study indicates a reciprocal relationship that exists between IL-10 and NO production influencing the regulation of blood pressure during chronic HS intake at elevated renin-angiotensin system.

In the present study, U_ISOV (an indirect marker for oxidative stress) was seen higher in IL-10KO compared to WT. This is in coherence with an earlier report⁷ suggesting the role of endogenous IL-10 in attenuating increases in vascular superoxide and endothelial dysfunction in disease conditions associated with diabetes and hypertension. The beneficial effects of IL-10 in endothelial dysfunction and other microvascular inflammatory complications have also been demonstrated earlier.^{2, 4, 5, 32} It was reported that patients with congestive heart failure register a decreased level of IL-10 in serum³³ and that the progression of vascular injury in several cardiovascular disease models has been retarded by exogenous administration of IL-10.¹ AngII-induced inflammation, oxidative stress and vascular dysfunction in IL-10KO mice were also shown to be reduced by transferring cultured Treg cells isolated from WT mice that produce IL-10.³¹

The percent areas of glomerular sclerosis as well as interstitial fibrosis (renal injury parameters) are greater in AngII treated IL-10KO compared to that in WT in the present study. These findings are consistent with earlier reports that endogenous IL-10 deficiency was involved in both sclerosis and fibrosis of renal tissue induced by many factors such as ureteral obstruction³⁴ and ischemia-reperfusion injury.³⁵ A reduction in IL-10 level was observed during AngII administration⁶ and AngII–induced vascular effects were prevented by exogenously applied IL-10.³⁰ In the present study, it has been observed that the renal tissue levels of TNF-α and IL-6 are higher in NS-IL-10KO compared to those in NS-WT supporting the notion that IL-10 exerts an inhibitory role in the generation of pro-inflammatory cytokines.^{1, 2, 13} However, TNF-α level was lower in HS-IL-10KO compared to NS-IL-10KO. As the eNOS protein expression as well as the U_NOxV is markedly increased in HS-IL-10KO, the increase in NO production could be a contributing factor in such low level of TNF-α in this strain. It seems that NO production reciprocally regulates pro-inflammatory cytokines as it is reported that NOS inhibition increase TNF-α^{13, 15} in mice and NO production enhances anti-inflammatory activity in some disease conditions.³⁶ It is also noted that the level of IL-6 is lower in plasma but higher in the renal tissue of NS-IL-10KO compared to that in NS-WT indicating that the changes in the plasma level of IL-6 may not always reflect tissue production of this cytokine.^{33, 37} The reason for such variability in the regional IL-6 level in these AngII treated mice is not yet clearly understood. However, it is also reported that many conditions including AngII treatment enhances soluble IL-6 receptors (sIL-6R) in the serum which, upon binding to IL-6, exerts biological actions in different organs or cell type.^{38, 39} Although no direct data is yet available, it is possible that an increase in plasma sIL-6R may occur during HS/AngII treatment which may bind with IL-6 to reduce its concentration in the plasma. Further experiments may be needed to examine this possibility. It should also be emphasized here that such variability in the levels of pro- and anti-inflammatory cytokines in different tissues may not be unexpected as the production of these cytokines depends on the local or global inflammatory responses in different conditions.

It is also interesting to note that blood pressure was generally higher in WT than that in IL-10KO, yet the renal injury parameters are generally higher in IL-10KO than those in WT, indicating that AngII induced renal injury may occur independent of an increase in blood pressure as reported⁴⁰ previously. It should be emphasized that both blood pressure dependent as well as independent factors are contributing in the development of AngII induced renal injury. Thus, it is not unexpected that a similar degree of renal injury can be associated with a variable degree of blood pressure in the different groups of mice as observed in the present study. The findings in the present study indicate a comparatively lesser degree of renal injury in IL-10KO mice than that would be generally expected in a condition of such deficiency in anti-inflammatory cytokine. This is mostly due to an increase in NO production in this IL-10KO strain that exerts ant-inflammatory action⁴¹ to provide a protective role against AngII induced renal injury. This is consistent with a report from Walley et al⁴² that NO down-regulates proinflammatory protein and mRNA expression during acute lung injury by an effect upstream of the activation of NF-kB, which binds to the promoter region of the pro-inflammatory cytokine genes.

In conclusion, the results of this investigation demonstrate that there is an increase in NO generation due to enhanced eNOS activity that occurs in the mice lacking IL-10 gene. Such enhanced production of NO helps to mitigate hypertensive and renal injury responses to chronic AngII and HS intake in the condition of IL-10 deficiency. These novel findings provide the evidence that IL-10 can act as pro-hypertensive, pro-inflammatory cytokine during chronic HS intake in the conditions in which AngII level is elevated.

Perspective

The implied pro-hypertensive role of IL-10 in SSH induced by AngII would have significant impact on the therapeutic approaches in the management of inflammatory renal injury associated with many hypertensive conditions. Such pro-hypertensive action of IL-10 achieved by reducing NO formation may also be a vital component in the pathophysiology of many critical conditions including sepsis. Acute early phase of sepsis is usually characterized by an excess of pro-inflammatory cytokines as well as NO production, while IL-10 level is predominantly increased during the secondary phase in which NO production is significantly attenuated.⁴³ Such an increase in IL-10 production in the late stage of sepsis helps to reduce pro-inflammatory cytokines as well as NO formation which would have further detrimental effects in critically ill patients.

Novelty and Significance.

What is new?

1. The role of IL-10 (anti-inflammatory cytokine) in the development of salt-sensitivity has not been addressed earlier in a comprehensive manner. This present study specifically investigates the blood pressure responses to chronic HS intake and AngII treatment in the condition of IL-10 deficiency. The results demonstrate that IL-10 deficiency itself do not induce salt–sensitivity, rather attenuates hypertensive response induced by chronic AngII treatment with HS intake. This is a novel finding as many previous studies implicated the opposite notion that a decrease in IL-10 may propagate AngII induced cardiovascular dysfunction.

2. The observation that IL-10 deficiency enhances endogenous NO production as well as eNOS protein expression induced by chronic AngII treatment and HS intake is also a novel finding in this study. This reciprocal relationship between endogenous IL-10 and NO production during AngII treatment or HS intake was not demonstrated in any previous study.

What is Relevant?

This finding of a reciprocal compensatory interaction between NO and IL-10 is significant as it is indicated that an inhibition of IL-10 activity, rather than IL-10 replacement, would be an adequate therapeutic target for the management of salt sensitive hypertension induced by elevated renin-angiotensin system.

Summary

The findings in the present study document a reciprocal compensatory interaction between NO and IL-10 in modulating AngII induced responses during HS intake. These findings also provide the evidence that IL-10 can act as pro-hypertensive, pro-inflammatory during chronic HS intake in the conditions in which AngII level is elevated.