Salivary, blood and plasma nitrite concentrations in periodontal patients and healthy individuals before and after periodontal treatment.

BACKGROUND: To date, no study has employed ozone-based reductive chemiluminescence to compare nitrite concentration in the saliva of periodontal disease (PD) and healthy individuals or in the various blood compartments of the same individuals before and after periodontal treatment. We evaluated nitrite concentrations in whole, submandibular, and parotid saliva, as well as in whole blood, erythrocytes, and plasma of healthy volunteers and patients with chronic periodontitis. METHODS: Data obtained for the PD and control groups were compared before and 3 months after periodontal therapy. RESULTS: At baseline, stimulated whole saliva nitrite concentration was lower in PD patients (mean=57.3 ± 9.8 µmol/L) as compared with healthy individuals (92.5 ± 13.6 µmol/L, P<0.05). PD and periodontal treatment did not affect submandibular or parotid saliva nitrite concentrations. PD patients presented higher baseline whole blood nitrite concentration (238.4 ± 45.7 µmol/L) as compared with values recorded 3 months after therapy (141.3 ± 20.1 nmol/L, P<0.05). PD patients' erythrocytes exhibited higher baseline nitrite concentration (573.1 ± 97.8 nmol/L) as compared with three months after therapy (298.7 ± 52.1 nmol/L, P<0.05). Again, PD and PD treatment did not impact plasma nitrite concentration. CONCLUSIONS: PD patients had lower nitrite concentration in whole saliva, and this situation remained unchanged after periodontal treatment. Nevertheless, erythrocytes and whole blood nitrite levels diminished after periodontal treatment.

Effect of metabolic syndrome risk factors and MMP-2 genetic variations on circulating MMP-2 levels in childhood obesity.

Matrix metalloproteinase-2 is involved in the development of the adipose tissue, and associated with cardiovascular diseases. Metabolic risk factors (MRFs) and functional polymorphisms in the MMP-2 gene may affect its expression and activity. We investigated whether traditional MRFs and two MMP-2 gene polymorphisms (C(-1306)T; rs243865, and C(-735)T; rs2285053) affect circulating MMP-2 levels in children and adolescents, and whether MMP-2 polymorphisms and/or haplotype are associated with susceptibility to childhood obesity. We studied 114 healthy controls, 43 obese, and 83 obese with ≥ 3 MRFs children and adolescents. Genotypes were determined by Taqman allele discrimination assay and real-time PCR. Plasma MMP-2 was measured using zymography. We found positive correlations between MMP-2 concentrations and mean blood pressure in all children and adolescents group (r = 0.132; P < 0.05) and in obese children and adolescents (r = 0.247; P < 0.01). We found that the CC genotype for the C(-1306)T polymorphism was more common in subjects with higher MMP-2 concentrations in controls (P = 0.003) and in the obese group (P = 0.013). The CT genotype (OR = 0.40; P < 0.01) and the T allele (OR = 0.48; P < 0.01) for the C(-735)T polymorphism were less common in obese children and adolescents than in controls. The haplotypes distribution did not show significant differences between control and obese (P > 0.05). Ours findings show that blood pressure is associated with circulating MMP-2 concentrations, and that the CC genotype for the C(-1306)T polymorphism was more common subjects (controls and obese) with higher MMP-2 concentrations, whereas the CT genotype and the T allele for the C(-735)T polymorphism are less common in obesity.

Oral contraceptive containing chlormadinone acetate and ethinylestradiol reduces plasma concentrations of matrix metalloproteinase-2 in women with polycystic ovary syndrome.

Biochemical markers of cardiovascular disease, including matrix metalloproteinases (MMPs), are altered in women with polycystic ovary syndrome (PCOS), with many of these alterations thought to be due to excess androgen concentrations. Despite oral contraceptives (OCs) being the first-line pharmacological treatment in women with PCOS and the importance of MMPs in many physiological conditions and pathological states, including cardiovascular diseases, no study has yet evaluated whether OCs alter plasma concentrations of MMPs. We therefore assessed whether treatment with an OC containing the anti-androgenic progestogen alters MMP profiles in women with PCOS. We analysed 20 women with PCOS who wanted hormonal contraception (OC-PCOS group), 20 ovulatory women who required hormonal contraception (OC-control group) and 20 ovulatory women who wanted non-hormonal contraception (non-OC-control group). OC consisted of cyclic use of 2 mg chlormadinone acetate/30 µg ethinylestradiol for 6 months. Plasma concentrations of MMP-2, MMP-9, TIMP-1 and TIMP-2 were measured by gelatin zymography or enzyme-linked immunoassays. OC treatment for 6 months significantly reduced plasma MMP-2 concentrations in the OC-control and OC-PCOS groups and TIMP-2 and TIMP-1 concentrations levels in the OC-control group (all p < 0.05), but had no effects on MMP-9 concentrations or on MMP-2/TIMP-2 and MMP-9/TIMP-1 ratios in any group (all p > 0.05). These findings indicated that long-term treatment with an OC containing chlormadinone acetate plus ethinylestradiol reduced plasma MMP-2 concentrations in both healthy and PCOS women. As the latter have imbalances in circulating matrix MMPs, treatment of these women with an OC may be beneficial.

Imbalanced circulating matrix metalloproteinases in polycystic ovary syndrome.

Altered levels of matrix metalloproteinases (MMPs) may reflect relevant pathogenetic mechanisms of disease conditions. The objective of this study was to compare the plasma levels of MMPs and tissue inhibitors of MMPs (TIMPs) in polycystic ovary syndrome (PCOS) patients with those found in healthy ovulatory controls and to examine whether the levels of these biomarkers are associated with clinical and biochemical features of this syndrome. Sixty-five healthy ovulatory subjects (controls) and 80 patients with PCOS were include in this study. MMP-2, MMP-8, MMP-9, TIMP-1, TIMP-2 concentrations were measured in plasma samples by gelatin zymography or enzyme-linked immunoassays. MMP-2, MMP-8, MMP-9, and TIMP-1 levels were similar in PCOS patients and in healthy controls (P > 0.05). PCOS patients had lower plasma TIMP-2 levels than healthy controls (P < 0.05). We found higher MMP-2/TIMP-2 and MMP-9/TIMP-1 ratios in PCOS patients than in healthy controls (all P < 0.05). Testosterone levels correlated positively with the MMP-9/TIMP-1 ratio and negatively with TIMP-2 levels (r = 0.26, P < 0.01 and r = -0.21, P = 0.02, respectively). In addition, only testosterone was an independent predictor of TIMP-2 levels (estimate = -0.35, P = 0.04) and the MMP-9/TIMP-1 ratio (estimate = 0.01, P = 0.04). We found evidence indicating that the balance between MMPs and TIMPs in women with PCOS is altered, probably due to androgen excess found in these women.

Quercetin restores plasma nitrite and nitroso species levels in renovascular hypertension.

Quercetin has antioxidants properties which may increase nitric oxide (NO) bioavailability. However, the effects of quercetin on NO status have been poorly studied. We evaluated whether quercetin improves the plasma levels of NO metabolites in two-kidney one-clip (2K1C) hypertensive rats and assessed its effect on endothelial function. Sham-operated and 2K1C rats were treated with quercetin (10 mg(-1) kg(-1) day(-1) by gavage) or vehicle for 3 weeks. Systolic blood pressure (SBP) was monitored weekly. Vascular responses to acetylcholine (Ach) and sodium nitroprusside (SNP) were assessed in hindquarter vascular bed. Plasma nitrate levels were assessed by Griess reagent and plasma nitrite and nitroso species (S, N-nitroso species) were assessed by ozone- based chemiluminescence. Aortic NADPH oxidase activity and superoxide production were evaluated. While quercetin had no effects in control normotensive rats (P > 0.05), it significantly reduced SBP in 2K1C rats (P < 0.05). At the end of treatment, plasma nitrate levels were similar in all experimental groups (P > 0.05). However, plasma nitrite and the nitroso species levels were significantly lower in 2K1C rats when compared with controls (P < 0.05). Quercetin treatment restored plasma nitrite and nitroso species levels to those found in the sham-vehicle group (P < 0.05). While quercetin treatment induced no significant changes in responses to SNP (P > 0.05), it restored the vascular responses to Ach. Quercetin significantly attenuated 2K1C-hypertension-induced increases in NADPH oxidase activity and vascular superoxide production (P < 0.05). These results suggest that the antihypertensive effects of quercetin were associated with increased NO formation and improved endothelial function, which probably result from its antioxidant effects.

Assessment of vascular effects of tamoxifen and its metabolites on the rat perfused hindquarter vascular bed.

Tamoxifen has been suggested to produce beneficial cardiovascular effects, although the mechanisms for these effects are not fully known. Moreover, although tamoxifen metabolites may exhibit 30-100 times higher potency than the parent drug, no previous study has compared the effects produced by tamoxifen and its metabolites on vascular function. Here, we assessed the vascular responses to acetylcholine and sodium nitroprusside on perfused hindquarter vascular bed of rats treated with tamoxifen or its main metabolites (N-desmethyl-tamoxifen, 4-hydroxy-tamoxifen, and endoxifen) for 2 weeks. Plasma and whole-blood thiobarbituric acid reactive substances (TBARS) concentrations were determined using a fluorometric method. Plasma nitrite and NOx (nitrite + nitrate) concentrations were determined using an ozone-based chemiluminescence assay and Griess reaction, respectively. Treatment with tamoxifen reduced the responses to acetylcholine (pD(2) = 2.2 +/- 0.06 and 1.9 +/- 0.05 after vehicle and tamoxifen, respectively; P < 0.05), while its metabolites improved these responses (pD(2) = 2.5 +/- 0.04 after N-desmethyl-tamoxifen, 2.5 +/- 0.03 after 4-hydroxy-tamoxifen, and 2.6 +/- 0.08 after endoxifen; P < 0.01). Tamoxifen and its metabolites showed no effect on endothelial-independent responses to sodium nitroprusside (P > 0.05). While tamoxifen treatment resulted in significantly higher plasma and whole blood lipid peroxide levels (37% and 62%, respectively; both P < 0.05), its metabolites significantly decreased lipid peroxide levels (by approximately 50%; P < 0.05). While treatment with tamoxifen decreased the concentrations of markers of nitric oxide formation by approximately 50% (P < 0.05), tamoxifen metabolites had no effect on these parameters (P > 0.05). These results suggest that while tamoxifen produces detrimental effects, its metabolites produce counteracting beneficial effects on the vascular system and on nitric oxide/reactive oxygen species formation.

Increased circulating levels of matrix metalloproteinase (MMP)-8, MMP-9, and pro-inflammatory markers in patients with metabolic syndrome.

BACKGROUND: Metabolic syndrome (MetS) predisposes to cardiovascular complications. Increased concentrations of pro-inflammatory mediators and imbalanced concentrations of matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) may reflect the pathophysiology of MetS. We compared the circulating levels of MMPs, TIMPs, and inflammatory mediators in MetS patients with those found in healthy controls. METHODS: We studied 25 healthy subjects and 25 MetS patients. The plasma levels of pro-MMP-2 and pro-MMP-9 were determined by gelatin zymography. The plasma concentrations of MMP-8, MMP-3, TIMP-1, TIMP-2, monocyte chemoattractant protein-1 (MCP-1), interleukin-6 (IL-6), intercellular adhesion molecule (sICAM-1), and sP-selectin were measured by ELISA kits. RESULTS: We found higher sP-selectin, sICAM-1, MCP-1, and IL-6 (all P<0.05) concentrations in MetS patients compared with healthy controls. No differences in pro-MMP-2, MMP-3, and TIMP-2 levels were found (all P>0.05). However, we found higher pro-MMP-9, MMP-8, and TIMP-1 levels in MetS patients compared with healthy controls (all P<0.05). CONCLUSIONS: Patients with MetS have increased circulating concentrations of pro-MMP-9, MMP-8, and TIMP-1 that are associated with increased concentrations of pro-inflammatory mediators and adhesion molecules. These findings suggest that MMPs may have a role in the increased cardiovascular risk of MetS patients. Pharmacological interventions targeting MMPs, especially MMP-9 and MMP-8 deserve further investigation in MetS patients.

Enhanced concentrations of relevant markers of nitric oxide formation after exercise training in patients with metabolic syndrome.

Metabolic syndrome (MetS) denotes a clustering of risk factors that may affect nitric oxide (NO) bioavailability and predispose to cardiovascular diseases, which are delayed by exercise training. However, no previous study has examined how MetS affects markers of NO formation, and whether exercise training increases NO formation in MetS patients. Here, we tested these two hypotheses. We studied 48 sedentary individuals: 20 healthy controls and 28 MetS patients. Eighteen MetS patients were subjected to a 3-month exercise training (E+group), while the remaining 10 MetS patients remained sedentary (E-group). The plasma concentrations of nitrite, cGMP, and ADMA (asymmetrical dimethylarginine; an endogenous nitric oxide synthase inhibitor), and the whole blood nitrite concentrations were determined at baseline and after exercise training using an ozone-based chemiluminescence assay, and commercial enzyme immunoassays. Thiobarbituric acid reactive species (TBA-RS) were measured in the plasma to assess oxidative stress using a fluorometric method. We found that, compared with healthy subjects, patients with MetS have lower concentrations of markers of NO formation, including whole blood nitrite, plasma nitrite, and plasma cGMP, and increased oxidative stress (all P<0.05). Exercise training increased the concentrations of whole blood nitrite and cGMP, and decreased both oxidative stress and the circulating concentrations of ADMA (both P<0.05). These findings show clinical evidence for lower endogenous NO formation in patients with MetS, and for improvements in NO formation associated with exercise training in MetS patients.

Nitric oxide formation is inversely related to serum levels of antiangiogenic factors soluble fms-like tyrosine kinase-1 and soluble endogline in preeclampsia.

Deficient NO formation has been implicated in hypertensive disorders of pregnancy. However, no previous study has compared the circulating nitrite concentrations in healthy pregnant women with those found in hypertensive disorders of pregnancy. Moreover, 2 antiangiogenic factors produced in the placenta (soluble fms-like tyrosine kinase-1 and soluble endogline) may affect NO formation during pregnancy. Here, we hypothesized that lower concentrations of markers of NO formation exist in hypertensive disorders of pregnancy and that inverse relationships exist between these markers and soluble fms-like tyrosine kinase-1 or soluble endogline. In this cross-sectional study, we compared 58 healthy pregnant women with 56 gestational hypertensive subjects and 45 preeclamptic patients. We measured plasma and whole blood nitrite concentrations using an ozone-based chemiluminescence assay and serum soluble fms-like tyrosine kinase-1 and soluble endogline concentrations using enzyme immunoassays. Whole blood nitrite levels were significantly lower in gestational hypertensive subjects and preeclamptic patients (-36% and -58%, respectively; both P<0.05) compared with healthy pregnant women. The plasma nitrite levels were approximately 37% lower in both groups with hypertensive disorders of pregnancy compared with the group with normotensive pregnancies (both P<0.05). As expected, we found higher circulating soluble fms-like tyrosine kinase-1 and soluble endogline concentrations in preeclampsia compared with gestational hypertensive subjects or with healthy pregnancies (both P<0.05). We found negative correlations between antiangiogenic factors and plasma or whole blood nitrite concentrations (Spearman's r from -0.175 to -0.226; all P<0.05). Our results show clinical evidence for impaired NO formation in preeclampsia or gestational hypertension. The negative correlations between markers of NO formation and antiangiogenic factors in preeclamptic patients suggest an inhibitory effect for these factors on NO formation.