Mercury Exposure Increases Circulating Net Matrix Metalloproteinase (MMP)-2 and MMP-9 Activities

Mercury (Hg) exposure causes health problems that may result from increased oxidative stress and matrix metalloproteinase (MMP) levels. We investigated whether there is an association between the circulating levels of MMP-2, MMP-9, their endogenous inhibitors (the tissue inhibitors of metalloproteinases; TIMPs) and the circulating Hg levels in 159 subjects environmentally exposed to Hg. Blood and plasma Hg were determined by inductively coupled plasma-mass spectrometry (ICP-MS). MMP and TIMP concentrations were measured in plasma samples by gelatin zymography and ELISA respectively. Thiobarbituric acid-reactive species (TBARS) were measured in plasma to assess oxidative stress. Selenium (Se) levels were determined by ICP-MS because it is an antioxidant. The relations between bioindicators of Hg and the metalloproteinases levels were examined using multivariate regression models. While we found no relation between blood or plasma Hg and MMP-9, plasma Hg levels were negatively associated with TIMP-1 and TIMP-2 levels, and thereby with increasing MMP-9/TIMP-1 and MMP-2/TIMP-2 ratios, thus indicating a positive association between plasma Hg and circulating net MMP-9 and MMP-2 activities. These findings provide a new insight into the possible biological mechanisms of Hg toxicity, particularly in cardiovascular diseases.

Comparative assessment of matrix metalloproteinase (MMP)-2 and MMP-9, and their inhibitors, tissue inhibitors of metalloproteinase (TIMP)-1 and TIMP-2 in preeclampsia and gestational hypertension

Objectives: To compare the circulating levels of matrix metalloproteinase (MMP)-2, MMP-9, tissue inhibitors of metalloproteinase (TIMP)-1, TIMP-2, and the MMP-9/TIMP-1 and MMP-2/TIMP-2 ratios in preeclampsia and gestational hypertension with those found in normotensive pregnancies. Design and methods: We studied 83 pregnant women (30 healthy pregnant women with uncomplicated pregnancies, 26 with gestational hypertension, and 27 with preeclampsia) and 30 healthy nonpregnant women in a cross-sectional study. MMP and TIMP concentrations were measured in plasma samples by gelatin zymography and ELISA, respectively. Results: We found higher plasma pro-MMP-9 levels, and higher pro-MMP-9/TIMP-1 ratios in women with gestational hypertension (95%-CI: 1.031 to 2.357, and 0.012 to 0.031, respectively), but not with preeclampsia, compared with those found in normotensive pregnant women (95%-CI: 0.810 to 1.350, and 0.006 to 0.013, respectively; both P<0.05). We found no significant differences in pro-MMP-2 levels (P>0.05). Conclusions: The higher net MMP-9 (but not MMP-2) activity in gestational hypertension compared with normotensive pregnancy suggests that MMP-9 plays a role in the pathophysiology of gestational hypertension. Conversely, the lack of such alterations in preeclampsia is consistent with the notion that different pathophysiological mechanisms are involved in these hypertensive disorders. (C) 2008 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Assessment of matrix metalloproteinase (MMP)-2, MMP-8, MMP-9, and their inhibitors, the tissue inhibitors of metalloproteinase (TIMP)-1 and TIMP-2 in obese children and adolescents

Objectives: To compare the circulating levels of matrix metalloproteinase (MMP)-8, pro-MMP-2, pro-MMP-9, and total MMP-9, their endogenous inhibitors, the tissue inhibitors of metalloproteinases (TIMP)-1 and TIMP-2, and the MMP-8/TIMP-1, MMP-9/TIMP-1, and MMP-2/TIMP-2 ratios in normotensive obese children and adolescents with those found in non obese children and adolescents. Design and methods: We studied 40 obese and 40 non obese (controls) children and adolescents in this cross-sectional study. MMP and TIMP concentrations were measured in plasma samples by gelatin zymography and ELISA. Results: Obese children and adolescents had higher circulating MMP-8 concentrations, lower plasma TIMP-1 concentrations, and higher MMP-8/TIMP-1 ratios than non obese controls (P < 0.05). We found no differences in pro-MMP-9 or total MMP-9 levels, or in MMP-9/TIMP-1 ratios between groups (P > 0.05). While we found no significant differences in pro-MMP-2 levels (P > 0.05) obese Subjects had higher TIMP-2 concentrations and lower pro-MMP-2/TIMP-2 ratios (P < 0.05) than non obese controls. Conclusions: In conclusion, we found evidence indicating higher net MMP-8 (but not MMP-9 and MMP-2) activity in childhood obesity. The increased MMP-8 levels found in obese children suggest a possibly relevant pathophysiological mechanism that may be involved in the increase of cardiovascular risk associated with childhood obesity. (c) 2009 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Differentially expressed genes in eutopic and ectopic endometrium of women with endometriosis

Objective: To elucidate the potential mechanisms involved in the physiopathology of endometriosis. We analyzed the differential gene expression profiles of eutopic and ectopic tissues from women with endometriosis. Design: Prospective laboratory study. Setting: University hospital. Patient(s): Seventeen patients in whom endometriosis was diagnosed and 11 healthy fertile women. Intervention(s): Endometrial biopsy specimens from the endometrium of healthy women without endometriosis and from the eutopic and ectopic endometrium tissues of patients with endometriosis were obtained in the early proliferative phase of the menstrual cycle. Main Outcome Measure(s): Six paired samples of eutopic and ectopic tissue were analyzed by subtractive hybridization. To evaluate the expression of genes found by rapid subtraction hybridization methods, we measured CTGF, SPARC, MYC, MMP and IGFBP1 genes by real-time polymerase chain reaction in all samples. Result(s): This study identified 291 deregulated genes in the endometriotic lesions. Significant expression differences were obtained for SPARC, MYC, and IGFBP1 in the peritoneal lesions and for MMP3 in the ovarian endometriomas. Additionally, significant differences were obtained for SPARC and IGFBP1 between the peritoneal and ovarian lesions. No significant differences were found for the studied genes between the control and the eutopic endometrium. Conclusion(s): This study identified 291 genes with differential expression in endometriotic lesions. The deregulation of the SPARC, MYC, MMP3, and IGFBP1 genes may be responsible for the loss of cellular homeostasis in endometriotic lesions. (Fertil Steril(R) 2010;93:1750-73. (C) 2010 by American Society for Reproductive Medicine.)

Hypothermia during endotoxemic shock in female mice lacking inducible nitric oxide synthase

The present study was undertaken to evaluate: (1) whether lipopolysaccharide LPS-incluced hypothermic responses may be altered during two estrous cycle phases, proestrus and diestrus, and after ovariectomy, followed by hormonal supplementation and (2) whether nitric oxide (NO) plays a role on LPS-induced hypothermia responses in female mice. Experiments were performed on adult female wild-type (WT) C57BL and inducible NO synthase knockout (KO) mice weighing 18 to 30 g. Endotoxemia was induced by intraperitoneal LIPS administration from Escherichia coli at a nonlethal dose of 10 mg/kg, and body temperature was measured by biotelemetry. Hormonal replacement was performed in ovariectomized mice through 17 beta-estradiol Silastic capsules (100 mu g) and s.c. injection of progesterone (0.5 mg per animal). We observed that during the diestrus phase, mice presented more intensive hypothermia than during proestrus phase, and hormonal supplementation with 17 beta-estradiol and progesterone attenuated hypothermia in ovariectomized mice. During diestrus and ovariectomy, KO mice had higher hypothermic response when compared with the WT group. During proestrus, the lack of statistical difference between KO and WT mice could be consequent of lower ovarian hormones plasma levels. After hormonal replacement, hypothermia was reverted in KO groups probably because of higher ovarian hormonal levels. In summary, the results demonstrated that NO release by inducible NO synthase has an important thermoregulatory role in LPS-incluced hypothermia in female mice. Besides, this involvement is directly dependent on the presence of ovarian hormones and their respective levels.

Expression of matrix metalloproteinase 2 (MMP-2) and tissue inhibitor of metalloproteinase 2 (TIMP-2) in medullary thyroid carcinoma: Prognostic implications

Background: Metalloproteinase 2 (MMP-2) and tissue inhibitor of metalloproteinase 2 (TIMP-2) participate in the degeneration of the extracellular matrix and are associated with carcinogenesis. MMP-2 is one of the main metalloproteinases active in neoplasia and is a marker of the malignant phenotype. Since the biological behavior of medullary thyroid carcinoma (MTC) varies widely, the present study was undertaken to determine if there is a correlation between the clinical evolution of MTC and the immunohistochemically detected expression of these enzymes in thyroid surgical specimens containing MTC. If so, their expression would be a novel indicator of the prognosis of MTC. Methods: Thirty-seven patients with MTC who had undergone thyroid surgery were followed for an average of 73 months. Immunohistochemical staining for metalloproteinase-related enzymes was performed in surgical paraffin blocks. The clinical status of the patients after surgery and at the end of the study period was characterized to determine correlations between these and the immunohistochemical markers. A value of p < 0.05 was considered statistically significant. Results: At the end of the study period, 15 patients (40.5%) were alive and without evidence of MTC, 17 (45.9%) had persistent MTC, and 5 (13.5%) had a relapse of their neoplasia. Four patients (10.8%) died during the course of the study. There was a significant correlation (p = 0.0005) between the immunohistochemical staining for MMP-2 and the clinical condition of the patients at the end of the study period, and a correlation between the state of apparent cure compared to persistence of MTC after thyroid surgery (p = 0.0207). No significant correlations were observed between either TIMP-2 expression or immune marking of metastatic lymph nodes and the clinical variables studied. Conclusion: Immunohistochemical expression of MMP-2 in thyroid surgical specimens from patients with MTC is a novel indicator of the prognosis of this cancer.

Videodensitometric analysis of advanced carotid plaque: correlation with MMP-9 and TIMP-1 expression

Background: Matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of MMP (TIMP) promote derangement of the extracellular matrix, which is ultimately reflected in plaque images seen on ultrasound. Videodensitometry can identify structural disturbances in plaques. Objectives: To establish the correlations between values determined using videodensitometry in B-mode ultrasound images of advanced carotid plaques and the total expression of MMP-9 and TIMP-1 in these removed plaques. Methods: Thirty patients underwent ultrasonic tissue characterization of carotid plaques before surgery, using mean gray level (MGL), energy, entropy and homogeneity. Each patient was assigned preoperatively to one of 2 groups: group I, symptomatic patients (n = 16; 12 males; mean age 66.7 +/- 6.8 years), and group II, asymptomatic patients (n = 14; 8 males; mean age 67.6 +/- 6.81 years). Tissue specimens were analyzed for MMP-9 and TIMP-1 expression. Nine carotid arteries were used as normal tissue controls. Results: MMP-9 expression levels were elevated in group II and in normal tissues compared to group I (p < 0.001). TIMP-1 levels were higher in group II than in group I, and significantly higher in normal tissues than in group I (p = 0.039). The MGL was higher in group II compared to group I (p = 0.038). Energy had greater values in group II compared to group I (p = 0.02). There were no differences between patient groups in homogeneity and entropy. Energy positively correlated with MMP-9 and TIMP-1 expression (p = 0.012 and p = 0.031 respectively). Homogeneity positively correlated with MMP-9 and TIMP-1 expression (p = 0.034 and p = 0.047 respectively). There were no correlations between protein expression and MGL or entropy. Conclusions: Videodensitometric computer analysis of ultrasound scanning images can be used to identify stable carotid plaques, which have higher total expression levels of MMP-9 and TIMP-1 than unstable plaques.

Effect of Laser Phototherapy on the Release of TNF-alpha and MMP-1 by Endodontic Sealer-Stimulated Macrophages

Objective: Our aim was to analyze the effect of laser phototherapy on the secretory activity of macrophages activated by interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS), and stimulated by substances leached from an epoxy resin-based sealer (AH-Plus) and a calcium hydroxide-based sealer (Sealapex). Background Data: Laser phototherapy can modulate the inflammatory process, improving wound healing. This type of therapy could be useful for modulating postoperative symptoms seen after endodontic treatment. Materials and Methods: Cytotoxicity was indirectly assessed by measuring mitochondrial activity. Macrophages were stimulated by the leached substances or not (controls), and the groups were then irradiated or not. The secretion of pro-inflammatory cytokines (TNF-alpha and MMP-1) was analyzed using ELISA. Two irradiations at 6-h intervals were done with an As-Ga-Al diode laser (780 nm, 70 mW, spot size 4.0 mm(2), 3 J/cm(2), for 1.5 sec) in contact mode. Results: The sealers were non-cytotoxic to macrophages. The production of TNF-alpha was significantly decreased by laser phototherapy, regardless of experimental group. The level of secretion of MMP-1 was similar in all groups. Conclusion: Based on the conditions of this study we concluded that in activated macrophages, laser phototherapy impairs the secretion of the pro-inflammatory cytokine TNF-alpha, but has no influence on MMP-1 secretion.

Low-Intensity Electrical Stimulation Counteracts the Effects of Ovariectomy on Bone Tissue of Rats: Effects on Bone Microarchitecture, Viability of Osteocytes, and Nitric Oxide Expression

Low Intensity Electrical Stimulation (LIES) has been used for bone repair, but little is known about its effects on bone after menopause. Osteocytes probably play a role in mediating this physical stimulus and they could act as transducers through the release of biochemical signals, such as nitric oxide (NO). The aim of the present study was to investigate the effects of LIES on bone structure and remodeling, NOS expression and osteocyte viability in ovariectomized (OVX) rats. Thirty rats (200-220 g) were divided into 3 groups: SHAM, OVX, and OVX subjected to LIES (OVX + LIES) for 12 weeks. Following the protocol, rats were sacrificed and tibias were collected for histomorphometric analysis and immunohistochemical detection of endothelial NO synthase (eNOS), inducible NOS (iNOS), and osteocyte apoptosis (caspase-3 and TUNEL). OVX rats showed significant (p < 0.05 vs. SHAM) decreased bone volume (10% vs. 25%) and trabecular number (1.7 vs. 3.9), and increased eroded surfaces (4.7% vs. 3.2%) and mineralization surfaces (15.9% vs. 7.7%). In contrast, after LIES, all these parameters were significantly different from OVX but not different from SHAM. eNOS and iNOS were similarly expressed in subperiosteal regions of tibiae cortices of SHAM, not expressed in OVX, and similarly expressed in OVX + LIES when compared to SHAM. In OVX, the percentage of apoptotic osteocytes (24%) was significantly increased when compared to SHAM (11%) and OVX + LIES (8%). Our results suggest that LIES counteracts some effects of OVX on bone tissue preserving bone structure and microarchitecture, iNOS and eNOS expression, and osteocyte viability.

Circulating matrix metalloproteinase-8 (MMP-8) and MMP-9 are increased in chronic periodontal disease and decrease after non-surgical periodontal therapy

Background: Periodontal disease shares risk factors with cardiovascular diseases and other systemic inflammatory diseases. The present study was designed to assess the circulating matrix metalloproteinases (MMPs) from chronic periodontal disease patients and, subsequently, after periodontal therapy. Methods: We compared the plasma concentrations of MMP-2. MMP-3, MMP-8, MMP-9, tissue inhibitor of metalloproteinase-1 (TIMP-1) and TIMP-2, and total gelatinolytic activity in patients with periodontal disease (n =28) with those of control subjects (n = 22) before and 3 months after non-surgical periodontal therapy. Results: Higher plasma MMP-3, MMP-8, and MMP-9 concentrations were found in periodontal disease patients compared with healthy controls (all P<0.05), whereas MMP-2, TIMP-1, and TIMP-2 levels were not different. Treatment decreased plasma MMP-8 and MMP-9 concentrations by 35% and 39%, respectively (both P<0.02), while no changes were found in controls. MMP-2, MMP-3, TIMP-1, and TIMP-2 remained unaltered in both groups. Plasma gelatinolytic activity was higher in periodontal disease patients compared with controls (P<0.001) and decreased after periodontal therapy (P<0.05). Conclusions: This study showed increased circulating MMP-8 and MMP-9 levels and proteolytic activity in periodontal disease patients that decrease after periodontal therapy. The effects of periodontal therapy suggest that it may attenuate inflammatory chronic diseases. (C) 2009 Published by Elsevier B.V.

Influence of ovariectomy and masticatory hypofunction on mandibular bone remodeling

Introduction: This study was designed to examine the effect of masticatory hypofunction and estrogen deficiency on mandible bone mass and compare this site with spine and femoral bone. Methods: Twenty-four rats were ovariectomized (OVX) or Sham-operated (Sham) and analyzed after feeding with hard diet (Hard) or soft diet (Soft). They were divided into four groups: (GI)Sham-Hard; (GII)OVX-Hard; (GIII)Sham-Soft and (GIV)OVX-Soft. Bone mineral density (BMD) was measured in the spine and femur in the baseline and at the end of the study, and Delta BMD (final BMD - baseline BMD) was calculated. In mandible bone, BMD and histomorphometry were analyzed at the end of the experiment. Results: Sham rats showed higher spine (GI: 13.5%vs GII: 0.74%, P < 0.01; GIII: 10.67%vs GIV: -4.36%, P < 0.001) and femur Delta BMD (GI: 14.43%vs GII: 4.42%, P < 0.01; GIII: 10.58%vs GIV: 0.49%, P < 0.001) than OVX, but no difference was observed in mandible BMD among these groups (P > 0.05). Soft-diet groups showed decreased mandible BMD compared with hard-diet groups (GIV vs GII, P < 0.01; GIII vs GI, P < 0.01). Similarly, mandibular condyle histomorphometry showed that soft-diet groups presented a significant decrease in trabecular thickness and volume (GIV vs GII, P < 0.05; GIII vs GI, P < 0.01) compared with hard diet. Conclusion: Our results suggest that mandibular bone loss resulted from decreased of mechanical loading during mastication, and was not affect by estrogen depletion.

Prognostic relevance of TTF-1 and MMP-9 expression in advanced lung adenocarcinoma

Background: The thyroid transcription factor-1 (TTF-1) is a tissue-specific transcription factor that Could playan important role in cell differentiation and morphogenesis of lung tumors. Matrix metalloproteinase-9 (MMP-9) is a protease commonly expressed in non-small cell lung cancer, conferring angiogenic and metastatic potential. Methods: We assessed TTF-1 and MMP-9 tumor expression by immunohistochemistry in 51 patients with lung adenocarcinoma, stage 11113 or IV, treated with platinum regimens. A bicategorical prognostic model was obtained using the Kaplan-Meier method, COX regression, and conjunctive consolidation. Results: The median expression of TTF-1 was 30.0% (range: 0-85.9%). All tumors expressed MMP-9 (median: 78.7%: range: 15.2-96.1%). Median survival was 41.6 weeks, with estimated 1- and 2-year survival rates of 45.0% and 22.0%, respectively. Poor performance status (Karnofsky scale) - hazards ratio(HR): 1.03. 95% confidence interval (CI): 1.01-1.06: low TTF-1 expression (<40%) - FIR: 4.00, 95% CI: 1.75-9.09: and high MMP-9 expression (>= 80%) - HR: 2.82, 95% CI: 1.30-6.08 were independent prognostic factors. Patients could be stratified in three death risk groups according to markers expression: low risk (high TTF-1 and low MMP-9; median survival: 127.6 weeks), intermediate risk (low TTF-1 OF high MMP-9; median survival: 39.0 weeks): and high risk (low TTF-1 and high MMP-9: median survival: 16.4 weeks). Conclusion: TTF-1 and MMP-9 tumor expression as detected by immunohistochemistry may allow identification of different, clinically meaningful, prognostic groups of advanced lung adenocarcinoma patients treated with platinum regimens. (C) 2008 Elsevier Ireland Ltd. All rights reserved.

Increased activities of cardiac matrix metalloproteinases matrix metalloproteinase (MMP)-2 and MMP-9 are associated with mortality during the acute phase of experimental Trypanosoma cruzi infection

The strong inflammatory reaction that occurs in the heart during the acute phase of Trypanosoma cruzi infection is modulated by cytokines and chemokines produced by leukocytes and cardiomyocytes. Matrix metalloproteinases (MMPs) have recently emerged as modulators of cardiovascular inflammation. In the present study we investigated the role of MMP-2 and MMP-9 in T. cruzi-induced myocarditis, by use of immunohistochemical analysis, gelatin zymography, enzyme-linked immunosorbent assay, and real-time polymerase chain reaction to analyze the cardiac tissues of T. cruzi-infected C57BL/6 mice. Increased transcripts levels, immunoreactivity, and enzymatic activity for MMP-2 and MMP-9 were observed by day 14 after infection. Mice treated with an MMP inhibitor showed significantly decreased heart inflammation, delayed peak in parasitemia, and improved survival rates, compared with the control group. Reduced levels of cardiac tumor necrosis factor-alpha, interferon-gamma, serum nitrite, and serum nitrate were also observed in the treated group. These results suggest an important role for MMPs in the induction of T. cruzi-induced acute myocarditis.

Severity dependent increases in circulating cardiac troponin I and MMP-9 concentrations after experimental acute pulmonary thromboembolism

Background: Making the diagnosis of acute pulmonary thromboembolism (APT) and assessing its severity is very challenging, While cardiac troponin I (cTnI) concentrations are promising in risk stratification, no previous study has examined whether there is a linear relation between cTnI concentrations and the severity of APT. Moreover, matrix metalloprotemases (MMPs) are involved in the pathophysiology of APT. However, it is unknown whether the increases in MMP concentrations after APT reflect the severity of this condition. We examined whether the circulating concentrations of these biomarkers increase in proportion to the severity of experimental APT induced in anesthetized dogs. Methods: APT was induced with autologous blood clots (saline, 1, 3, or 5 ml/kg) injected into the right atrium. Hemodynamic evaluations were carried out for 120 min. Gelatin zymography of MMP-2 and MMP-9 from plasma samples were performed and serum cTnI concentrations were determined at baseline and 120 min after APT. Results: While no significant increases in pro-MMP-2 concentrations were found after APT, pro-MMP-9 concentrations increased by 80% only after 5 ml/kg of clot embolization. Serum cTnI and plasma pro-MMP-9 concentrations correlated positively with pulmonary vascular resistance (P=0.007 and rs=0.833 for troponin 1, and P=0.034 and rs=0.684 for pro-MMP-9) and with pulmonary artery pressure (P=0.005 and rs=0.610 for troponin 1, and P=0.022 and rs=0.720 for pro-MMP-9). Conclusions: Circulating cTnI and pro-MMP-9 increase in proportion to the severity of APT, although the increases in plasma pro-MMP-9 are less clear with less severe APT. These findings may be relevant for clinical APT. (C) 2007 Elsevier B.V. All rights reserved.

Matrix metalloproteinase (MMP)-9 genotypes and haplotypes in preeclampsia and gestational hypertension

Background: Abnormal production of matrix metalloproteinases (MMPs), especially MMP-9, may play a role in hypertensive disorders of pregnancy. These alterations may result from functional genetic polymorphisms in the promoter region of MMP-9 gene, which are known to change MMP-9 expression. We examined whether 2 MMP-9 polymorphisms (C(-) (1562) T and (CA)n) and haplotypes are associated with preeclampsia and/or gestational hypertension. Methods: We studied 476 pregnant women: 176 healthy pregnant (HP), 146 pregnant with gestational hypertension (GH), and 154 pregnant with preeclampsia (PE). Genomic DNA was extracted from whole blood and genotypes for C(- 1562) T and (CA)n polymorphisms were determined by PCR-RFLP. Haplotype frequencies were inferred using the PHASE ver. 2.1 program. Results: For the g.-90(CA)13-25 polymorphism, no significant differences were found in genotype and allele distributions when PE or GH groups were compared with HP group. However, the CT genotype and T allele for g.-1562C>T polymorphism were more commonly found in GH subjects compared with the HP group (both P<0.05). Conversely, we found no differences in genotypes or allele distributions for the g.-1562C>T polymorphism when the PE and the HP groups were compared. No significant differences were found in overall distributions of haplotype frequencies when the GH or the PE group was compared with the HP group. Conclusions: The C(- 1562) T polymorphism in MMP-9 gene is associated with gestational hypertension, but not with preeclampsia. These findings may help to explain the higher plasma MMP-9 levels previously reported in GH compared with HP. (C) 2010 Elsevier B.V. All rights reserved.