Análise da expressão imuno-histoquímica da ciclofilina A, EMMPRIN e MMP-7 na doença periodontal

Periodontal disease is an infectious disease resulting from the immunoinflammatory response of the host to microorganisms present in the dental biofilm which causes tissue destruction. The objective of this study was to evaluate the immunohistochemical expression of cyclophilin A (CYPA), extracellular matrix metalloproteinase inducer (EMMPRIN) and matrix metalloproteinase 7 (MMP-7) in human specimens of clinically healthy gingiva (n=32), biofilm-induced gingivitis (n=28), and chronic periodontitis (n=30). Immunopositivity for CYPA, EMMPRIN and MMP-7 differed significantly between the three groups, with higher percentages of staining in chronic periodontitis specimens, followed by chronic gingivitis and healthy gingiva specimens (p < 0.001). Immunoexpression of CYPA and MMP-7 was higher in the intense inflammatory infiltrate observed mainly in cases of periodontitis. Analysis of possible correlations between the immunoexpression of EMMPRIN, MMP-7 and CYPA and between the expression of these proteins and clinical parameters (probing depth and clinical attachment loss) showed a positive correlation of CYPA expression with MMP-7 (r = 0.831; p < 0.001) and EMMPRIN (r = 0.289; p = 0.006). In addition, there was a significant positive correlation between probing depth and expression of MMP-7 (r = 0.726; p < 0.001), EMMPRIN (r = 0.345; p = 0.001), and CYPA (r = 0.803; p < 0.001). These results suggest that CYPA, EMMPRIN and MMP-7 are associated with the pathogenesis and progression of periodontal disease

Calcaneal Tendon Regions Exhibit Different MMP-2 Activation After Vertical Jumping and Treadmill Running

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Finasteride treatment alters MMP-2 and-9 gene expression and activity in the rat ventral prostate

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Tissue inhibitor of metalloproteinase-2 (TIMP-2) location in the ventral, lateral, dorsal and anterior lobes of rat prostate by immunohistochemistry

Matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) play a major role in extracellular matrix component degradation in several normal and abnormal tissue situations; they are also found in human seminal plasma. MMPs have been found in rat prostate secretions and are nearly lobe specific in expression pattern. The aim of this study was to evaluate whether TIMP-2, like other semen components, is expressed differently from different rat prostatic lobes. Immunohistochemical staining was performed in both young and adult rat ventral (VP), lateral (LP), dorsal (DP), and anterior (AP) prostatic lobes and confirmed by western blotting. TIMP-2 expression was found in the epithelial cells in the following sequence: LP > AP > DP > VP, in both young and adult rats. In this study, 100% of adult LP presented histological signs of prostatitis, where TIMP-2 immunostaining was positive in normal epithelium even with intraluminal neutrophils, but was reduced or absent in the epithelium with intraepithelial leukocytes or with periductal stroma disorganization associated with mononuclear cell infiltration. However, TIMP-2 expression in LP was not induced by prostatitis, since younger rat LPs were also strongly TIMP-2 positive. The distal and intermediate VP regions were TIMP-2 negative, but the proximal regions were strongly stained. Western blotting results confirmed the high TIMP-2 expression in the LP lobe. Thus, TIMP-2 is expressed differently between the prostatic lobes and is another nearly lobe-specific protein, which plays a role in the regulation of MMP activity in seminal plasma and glandular homeostasis. TIMP-2 is also another regional ductal variation of VP. Further studies should address whether TIMP-2 expression is related to the highest incidence of rat LP prostatitis and adenocarcinoma. © 2006 International Federation for Cell Biology.

MMP-2 and MMP-9 localization and activity in the female prostate during estrous cycle

The gerbil female prostate undergoes morphological and physiological changes resulting from hormonal fluctuations that occur during the reproductive cycle. These repetitive cycles of glandular growth and regression are followed by an extensive reconstruction and remodeling of prostate stroma throughout the reproductive life of the female gerbil. The objective of this study was to evaluate the effect that the hormonal fluctuations of the reproductive cycle have on the stromal remodeling and the expression and activity of matrix metalloproteinases MMP-2 and -9 in the adult female gerbil prostate. For this, serological, ultrastructural, immunohistochemical and biochemical methods were employed. The results showed that the major stromal alteration coincide with the peak of estradiol, which occurs in estrus, and with the peak of progesterone, occurring during diestrus II. MMP-2 and -9 presented a similar pattern of expression and activity during estrous cycle. The estrus was the phase of greater expression and activity of MMP-2 and -9. On the other hand, in DI and DII, the tissue expression and activity of MMP-2 and -9 was very weak. These results are important since they suggest the involvement of estradiol and progesterone in regulating the expression and activity of MMP-2 and -9 in adult gerbil female prostate. © 2011 Elsevier Inc.

MMP-2 and TIMP-2 in the prostates of male and female mongolian gerbils: Effects of hormonal manipulation

TIMPs in the prostates of male and female gerbils and evaluated the effects of testosterone on the expression of these enzymes. Ventral prostates from male gerbils that were either intact or had been castrated for 7 or 21 days, along with prostates from female gerbils that were either intact or had been treated with testosterone for 7 or 21 days, were submitted to histological, stereological and immunohistochemical analyses. Stereology of prostatic components showed significant alterations of tissue compartments in the ventral male prostate after castration, especially after 21 days, with a significant increase in stroma. Administration of testosterone led to disorganization in the female prostate, with a significant increase in collagen fibers and smooth muscle cells after 21 days, along with the development of epithelial lesions such as PINs. MMP-2 increased after 21 days of castration in males; however, the TIMP-2 immunoreaction for this group was weak or absent. In females, the expression of MMP-2 appeared to decrease after 7 days of treatment with testosterone, but after 21 days, both epithelium and stroma showed a stronger reaction for MMP-2 than the controls. The expression of TIMP-2 in the treated females was similar to its expression in the castrated males. We conclude that the distribution of MMPs and TIMPs in both male and female prostates is altered by androgen manipulation, but the mechanism of stromal regulation appears to be distinct between genders because both the lack of T in castrated males and the excess levels of T in treated females lead to the same effect.

Effect of phosphoric acid on the degradation of human dentin matrix

This study determined if dentin proteases are denatured by phosphoric acid (PA) used in etch-and-rinse dentin adhesives. Dentin beams were completely demineralized with EDTA for 30 days. We acid-etched experimental groups by exposing the demineralized dentin beams to 1, 10, or 37 mass% PA for 15 sec or 15 min. Control beams were not exposed to PA but were incubated in simulated body fluid for 3 days to assay their total endogenous telopeptidase activity, by their ability to solubilize C-terminal crosslinked telopeptides ICTP and CTX from insoluble dentin collagen. Control beams released 6.1 ± 0.8 ng ICTP and 0.6 ± 0.1 ng CTX/mg dry-wt/3 days. Positive control beams pre-incubated in p-aminophenylmercuric acetate, a compound known to activate proMMPs, released about the same amount of ICTP peptides, but released significantly less CTX. Beams immersed in 1, 10, or 37 mass% PA for 15 sec or 15 min released amounts of ICTP and CTX similar to that released by the controls (p > 0.05). Beams incubated in galardin, an MMP inhibitor, or E-64, a cathepsin inhibitor, blocked most of the release of ICTP and CTX, respectively. It is concluded that PA does not denature endogenous MMP and cathepsin activities of dentin matrices. © 2013 International & American Associations for Dental Research.

Platelet-Rich Plasma And Chronic Wounds: Remaining Fibronectin May Influence Matrix Remodeling And Regeneration Success

Background: Platelet-rich plasma has been largely used as a therapeutic option for the treatment of chronic wounds of different etiologies. The enhanced regeneration observed after the use of platelet-rich plasma has been systematically attributed to the growth factors that are present inside platelets' granules. Aim: We hypothesize that the remaining plasma and platelet-bound fibronectin may act as a further bioactive protein in platelet-rich plasma preparations. Methods: Recent reports were analyzed and presented as direct evidences of this hypotheses. Results: Fibronectin may directly influence the extracellular matrix remodeling during wound repair. This effect is probably through matrix metalloproteinase expression, thus exerting an extra effect on chronic wound regeneration. Conclusions: Physicians should be well aware of the possible fibronectin-induced effects in their future endeavors with PRP in chronic wound treatment. © 2013 International Society for Cellular Therapy.

Expressão de MMP-13 e ativação de vias de sinalização intracelular em dois modelos de doença periodontal induzida experimentalmente em ratos

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Extracellular matrix remodeling of the testes through the male reproductive cycle in Teleostei fish

During the fish reproductive cycle, testes undergo morphological changes related to germinal epithelium and remodeling of extracellular matrix components (ECM). ECM is degraded mainly by action of matrix metalloproteinases (MMPs). Due to the natural renewal of ECM in fish testes, we choose Pimelodus maculatus to study remodeling of ECM throughout reproductive cycle, using picrosirius (to identify type I, II, III collagen) and reticulin (type III collagen), and to immunolocalize MT1-MMP (membrane type 1-matrix metalloproteinase) and MMP-2 in testis cells. Testes were classified in four reproductive phases: regenerating, development, spawning capable and regressing. Picrosirius and reticulin demonstrated a differential distribution of total collagen fibers during the reproductive cycle. Immunohistochemistry showed MT1-MMP only in acidophilic granulocyte cells mainly inside blood vessels, in connective tissue of capsule close to the germinal compartment, and also infiltrated in interstitial connective tissue. MMP-2 was detected in fibroblast and endothelial cells of interstitial and capsule blood vessels, in epithelial cells of capsule, and in acidophilic granulocyte cells at same description for MT1-MMP. The fish testes ECM were remodeled throughout reproductive cycle in according to morphophysiological alterations. During reproductive season (spawning capable), the interstitium increased in total collagen fibers (type I, II, III). After spermiation period (regression and regenerating), the amount of collagen fibers decreased in response to action of MMPs on collagen degradation and other interstitial components (not assessed in this study). MMPs seem to be indispensable components for natural cyclic events of ECM remodeling of fish testes and for guarantee tissue homeostasis throughout reproductive cycle.

Functional Polymorphism Located in MMP-9 Gene Promoter Is Strongly Associated with Obesity

The adipose tissue expansion is accompanied by remodeling of extracellular matrix performed by matrix metalloproteinases (MMPs). Higher plasma and tissue MMP-9 levels are found in obese; therefore, we evaluated if the functional C-1562T polymorphism (rs3918242) located in promoter region of the MMP-9 gene is associated with obesity in women. We studied 112 lean and 114 obese women. Plasma MMP-9 and tissue inhibitor of MMP-9 (TIMP)-1 were measured using enzyme-linked immunosorbent assay. We found different genotype frequencies between lean and obese women (p = 0.008), prevailing T-allele in obese (2.3-fold). However, although obese women present higher levels of plasma MMP-9, lack of modulation by the polymorphism was found (all p > 0.05). Our findings suggest that C-1562T polymorphism may contribute to pathogenetic mechanisms involved in the development of obesity in women.

Comprehensive Evaluation of the Effects of Enalapril on Matrix Metalloproteinases Levels in Hypertension

Angiotensin-converting enzyme inhibitors (ACEi) may downregulate matrix metalloproteinases (MMPs). We examined whether enalapril affects MMP-2, MMP-8, and MMP-9 levels and activity, and their endogenous inhibitors (tissue inhibitors of MMPs, TIMP-1 and TIMP-2) levels in hypertensive patients. Moreover, we assessed the effects of enalaprilat on MMP-9 and TIMP-1 secretion by human endothelial cells (HUVECs). Thirty-eight hypertensive patients received enalapril for 8 weeks and were compared with thirty-eight normotensive controls. Blood samples were collected at baseline and after treatment. Plasma ACE activity was determined by a fluorimetric assay. Plasma MMP-2, MMP-8, MMP-9, TIMP-1, and TIMP-2 were measured by ELISA and gelatin zymography. A fluorogenic peptide cleavage assay was used to measure MMP activity. HUVECs cells were stimulated by phorbol-12-myristate-13-acetate (PMA) and the effects of enalaprilat (10(-10) to 10(-6) M) on MMP-9 and TIMP-1 levels were determined. Enalapril decreased blood pressure and ACE activity in hypertensive patients (P < 0.05), but had no effects on plasma MMP-2, MMP-8, MMP-9, TIMP-1, and TIMP-2 levels, or MMP activity. Enalaprilat had no effects on PMA-induced increases in MMP-9 and TIMP-1 secretion by HUVECs or on MMP activity. We show consistent evidence, both in vivo and in vitro, that enalapril does not affect MMPs and TIMPs levels in hypertensive patients.

Increased expression of MMP-9 and IL-8 are correlated with poor prognosis of Bladder Cancer

Background: Extracellular matrix homeostasis is strictly maintained by a coordinated balance between the expression of metalloproteinases (MMPs) and their inhibitors. The purpose of this study was to investigate whether the expression of MMP-9, MMP-2 and its specific inhibitors, are expressed in a reproducible, specific pattern and if the profiles are related to prognosis in Bladder Cancer (BC). Methods: MMP-9, MMP-2 and its specific inhibitors expression levels were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) in fresh-frozen malignant tissue collected from 40 patients with BC submitted to transurethral resection of bladder. The control group consisted of normal bladder tissue from five patients who had undergone retropubic prostatectomy to treat benign prostatic hyperplasia. Results: MMP-9 was overexpressed in 59.0 % of patients, and MMP-2, TIMP-1, TIMP-2, MMP-14, RECK and IL-8 was underexpressed in most of the patients. Regarding prognostic parameters we observed that high-grade tumors exhibited significantly higher levels of MMP-9 and IL-8 (p = 0.012, p = 0.003). Invasive tumors (pT1-pT2) had higher expression levels of MMP-9 than superficial tumors (pTa) (p = 0.026). The same was noted for IL-8 that was more expressed by invasive tumors (p = 0.015, p = 0.048). Most importantly tumor recurrence was related with higher levels of both MMP-9 (p = 0.003) and IL-8 (p = 0.005). Conclusion: We have demonstrated that the overexpression of MMP-9 and higher expression of IL-8 are related to unfavorable prognostic factors of urothelial bladder cancer and tumor recurrence and may be useful in the follow up of the patients.

Underexpression of MMP-2 and its Regulators, TIMP2, MT1-MMP and IL-8, is Associated with Prostate Cancer

Objective: Extracellular matrix homeostasis is strictly maintained by a coordinated balance between the expression of metalloproteinases (MMPs) and their regulators. The purpose of this study was to investigate whether MMP-2 and its specific regulators, TIMP-2, MT1-MMP and IL-8, are expressed in a reproducible, specific pattern and if the profiles are related to prognosis and clinical outcome of prostate cancer (PCa). Materials and Methods: MMP-2, TIMP-2, MT1-MMP and IL-8 expression levels were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) in freshly frozen malignant and benign tissue specimens collected from 79 patients with clinically localized PCa who underwent radical prostatectomies. The control group consisted of 11 patients with benign prostate hyperplasia (BPH). The expression profile of the MMP-2 and its regulators were compared using Gleason scores, pathological stage, pre-operative PSA levels and the final outcome of the PCa. Results: The analysis of 79 specimens of PCa revealed that MMP-2, TIMP-2, MT1-MMP and IL-8 were underexpressed at 60.0%, 72.2%, 62.0% and 65.8%, respectively, in malignant prostatic tissue in relation to BPH samples. Considering the prognostic parameters, we demonstrated that high Gleason score tumors (>= 7) over-expressed MMP-2 (p = 0.048) and TIMP-2 (p = 0.021), compared to low Gleason score tumors (< 7). Conclusion: We have demonstrated that MMP-2 and its regulators are underexpressed in PCa. Alternatively, overexpression of MMP-2 and TIMP-2 was related to higher Gleason score tumors. We postulate that alterations in metalloproteinase expression may be important in the control of tissue homeostasis related to prostate carcinogenesis and tumor behavior.

Epistasis among eNOS, MMP-9 and VEGF maternal genotypes in hypertensive disorders of pregnancy

Polymorphisms of the endothelial nitric oxide synthase (eNOS), matrix metalloproteinase-9 (MMP-9) and vascular endothelial growth factor (VEGF) genes were shown to be associated with hypertensive disorders of pregnancy. However, epistasis is suggested to be an important component of the genetic susceptibility to preeclampsia (PE). The aim of this study was to characterize the interactions among these genes in PE and gestational hypertension (GH). Seven clinically relevant polymorphisms of eNOS (T-786C, rs2070744, a variable number of tandem repeats in intron 4 and Glu298Asp, rs1799983), MMP-9 (C-1562T, rs3918242 and -90(CA)(13-25), rs2234681) and VEGF (C-2578A, rs699947 and G-634C, rs2010963) were genotyped by TaqMan allelic discrimination assays or PCR and fragment separation by electrophoresis in 122 patients with PE, 107 patients with GH and a control group of 102 normotensive pregnant (NP) women. A robust multifactor dimensionality reduction analysis was used to characterize gene-gene interactions. Although no significant genotype combinations were observed for the comparison between the GH and NP groups (P>0.05), the combination of MMP-9-1562CC with VEGF-634GG was more frequent in NP women than in women with PE (P<0.05). Moreover, the combination of MMP-9-1562CC with VEGF-634CC or MMP-9-1562CT with VEGF-634CC or-634GG was more frequent in women with PE than in NP women (P<0.05). These results are obscured when single polymorphisms in these genes are considered and suggest that specific genotype combinations of MMP-9 and VEGF contribute to PE susceptibility. Hypertension Research (2012) 35, 917-921; doi:10.1038/hr.2012.60; published online 10 May 2012