EDWIN ROGER PARRA CUENTAS
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conferenceObject Increased decorin and type V collagen in SSc pulmonary fibrosis(2012) TEODORO, W.; VELOSA, A. P.; MARCELINO, A.; MARTIN, P.; CARRASCO, S.; GOLDENSTEIN-SCHAINBERG, C.; PARRA, E.; YOSHINARI, N.; CAPELOZZI, V.Objective: To evaluate COL V and decorin expression in pulmonary tissue and to characterize biochemical profile of COLV from lung fibroblasts culture from SSc patients. Method: We evaluated COL V and decorin expression and tridimensional reconstruction (3D) of 6 patients with SSc without pulmonary hypertension that underwent surgical lung biopsy and as control was obtained lung fragments from 6 normal individuals who died from trauma. COL V amount in lung sections was evaluated with immunofluorescence. To biochemical characterization of COL V from lung fibroblasts culture was used quantitative immunoblot. Results: It was found that the structure of COLV fibers was distorted and strongly thickened in lung tissue from SSc patients compared with thin fibers pattern in the healthy controls. Decorin was distributed around COL V fibrils in the bronchovascular interstitium and vascular walls. Histomorphometric analysis of SSc lung demonstrated increased expression of both COL V and decorin when compared to the control (p<0.01). The semiquantitative imunoblot detected an increased high molecular weight COLV fraction in patients when compared to the control. Conclusion: The over expression and unusual organization of COLV fibers with biochemical changes associated to increased decorin indicates that matrix signalization pathway is involved in COLV fibrillogenesis process in SSc pulmonary fibrosis.conferenceObject Collagen V induces differentiation of rabbit adipose tissue-derived stem cells in chondrocyte-like phenotype(2012) TEODORO, W.; CRUZ, I. Brindo da; VELOSA, A. P.; CARRASCO, S.; GOLDENSTEIN-SCHAINBERG, C.; FULLER, R.; PARRA, E.; CAPELOZZI, V.Objective: Stimulated mesenchymal stem cells (MSCs) have capacity of differentiation in many cell types. It is being used in degenerative diseases treatment protocols. We evaluated the collagen V (COL V) and collagen XI (COL XI) influence in the differentiation of rabbits adipose tissue-derived MSCs in a chondrocyte-like cell phenotype. Method: MSCs isolated of New Zealand rabbits adipose-tissue were maintained in culture by 4 weeks. COLV, COLXI and COLV/XI (10μg/ml) were added to culture during 72 h. The cells aggregates were stained with Toluidine blue, Alcian blue and Picrosirius. Chondrocyte-like phenotype was confirmed by immunofluorescence to CD34, vimentin and collagens I, II and III. Results: MSCs stimulated with COLV expressed proteoglicans and collagen, when compared with COLXI and COLV/XI and control. In the presence of COLV, MSCs was capable to increase collagen II expression confirming its chondro-cyte-like cell phenotype. In contrast, MSCs cultured with COLXI and COLV/XI express collagen I and III. Conclusion: The data suggest that COLV may facilitate the differentiation of rabbit adipose tissue-derived stem cells into a chondrocyte-like phenotype. Further studies are urged in order to evaluate the influence of COLV in the ability of chondrocytes to remodel osteoarthritic joint surface at ultrastructural and molecular levels.conferenceObject Fibrogenesis failure of type V collagen observed in pulmonary and cutaneous fibroblast culture reinforce the pathogenic participation of this collagen in the pathway of systemic sclerosis(2012) TEODORO, W. R.; MORAIS, J.; MARTIN, P.; VELOSA, A. P. P.; CARRASCO, S.; SOUZA, R. B. C.; KATAYAMA, M. L.; GOLDEINSTEIN-SCHAINBERG, C.; PARRA, E. R.; CAPELOZZI, V. L.; YOSHINARI, N. H.Introduction: Unusual type V collagen (COLV) accumulation was demonstrated in systemic sclerosis (SSc) by our group. In this regard, this study analyzed tridimensional reconstruction (3D), biochemical and molecular profile of COLVα1 and COLVα2 chains in pulmonary and cutaneous fibroblasts culture from patients with SSc. Materials and Methods: Pulmonary and cutaneous fibroblasts for culture were obtained from 7 patients with SSc and from six controls respectively. COLV 3D reconstruction was performed by confocal microscopy. COLVα1 and COLVα2 gene expression was performed by RT-PCR and COLV protein expression by immunoblotting. Results: COL V 3D reconstruction showed distorted and strongly thickened fibers with irregular bundles resulting in a dense network in lung and skin fibroblast cultures from SSc patients compared to the thin fibers from fibroblast controls. Collagen quantification showed significant increased COLV fiber expression in SSc cutaneous and pulmonary fibroblasts (P<0.01) compared with the respective controls. In the same way, molecular evaluation demonstrated an increased significance (P=0.05) of COLVα1 and COLVα2 mRNA expression in cutaneous and pulmonary fibroblasts from SSc patients to that of control groups. The immunoblotting analysis demonstrated the increased weight of the molecular COLV chains. Conclusion: COLV overexpression and an unusual organization of these fibers including molecular and biochemical changes, suggest an interference process of the COLV fibrillogenesis in patients with SSc, reinforcing the participation of this collagen in SSc pathogenesis and open new therapeutic perspectives for these patients.conferenceObject INFLUENCE OF ALPHA 2 DO COLLAGEN V OVEREXPRESSION IN PHYSIOPHATOLOGY OF FIBROSIS SYSTEMIC SCLEROSIS PATIENTS(2014) MORAIS, J.; MARTIN, P.; VELOSA, A. P. P.; ANDRADE, P. C.; CRUZ, I. B.; MIRACCA, E. C.; FAC, F. A. C. Barrence; CARRASCO, S.; GOLDEINSTEIN-SCHAINBERG, C.; NAGAI, M. A.; PARRA, E. R.; CAPELOZZI, V. L.; TEODORO, W. R.conferenceObject HLA-B27 PREVALENCE IN A COHORT OF BRAZILIAN PATIENTS WITH PSORIATIC ARTHRITIS AND ANKYLOSING SPONDYLITIS(2014) GOLDENSTEIN-SCHAINBERG, C.; CARRASCO, S.; SAAD, C. G.; MORAES, J. C. B.; GONCALVES, C. R.; SAMPAIO-BARROS, P.; PARRA, E. R.conferenceObject DYNAMIC COLLAGEN V REMODELING IS RELATED TO SKIN THICKENING IN SSc(2012) MARTIN, P.; TEODORO, W. R.; VELOSA, A. P.; CARRASCO, S.; MORAIS, J. de; CHRISTMANN, R. B.; PARRAS, E. R.; CAPELOZZI, V. L.; YOSHINARI, N. H.Background. Normal physiological properties of skin, one of the primary organs affected in SSc, depends on collagen Types I (COL I), III (COLIII) and V (COLV) assembly forming heterotypic fibres. COLV regulates fibril diameter and loss of this function could result in tissue fibrosis. In this way, our aim was to evaluate the histological and molecular profiles of COLI, COLIII and COLV in SSc skin and its correlation with skin thickening and disease activity. Methods. Skin biopsies of 18 patients (5 at early and 13 at late disease stage) and 10 healthy controls were studied. Assessment of skin thickening was performed using the modified Rodnan skin score (MRSS) and disease activity was calculated by Valentini Disease Activity Index. Quantification of COLI, COLIII and COLV was evaluated by histomorphometry in dermis and quantitative RT–PCR in dermal fibroblast culture. Results. A higher expression of abnormal COLV was observed in dermis of patients with early disease when compared with control group and late disease. The COLIII content was also higher in early SSc when compared with healthy controls and late SSc. On the other hand, the amount of COLI was higher in late disease when compared with control and early SSc. A positive correlation between COLV and MRSS (r = 0.42, P = 0.04) as well as disease activity (r = 0.45, P = 0.03) was observed, but there was no correlation between COLI and COLIII expression and these parameters. COLV α-1 and COLV α-2, as well as COLI α-1 and COLIII α-1 mRNA expression were higher in SSc when compared with control group. Conclusion. We found increased COLIII and COLV deposition in early SSc and increased COLI expression in late SSc indicating that collagen remodelling in SSc is a dynamic process. The fact that abnormal COLV expression decreases in later disease stages could explain why skin thickening sometimes improves spontaneously with time. Besides, COLV is correlated to MRSS and disease activity. These findings include COLV as an important regulator of cutaneous thickness in SSc and may add this protein as a new target for future treatments.conferenceObject COLVa2 a Biomarker of Vasculopathy in Scleroderma?(2013) MORAIS, J.; MARTIN, P.; CAMARGO, I. C.; KATAYAMA, M. L.; CARRASCO, S.; GOLDEINSTEIN-SCHAINBERG, C.; PARRAS, E. R.; BARRENCE, F.; VELOSA, A. P.; CAPELOZZI, V. L.; TEODORA, W. R.conferenceObject HLA-B27 ANALYSIS BY FLOW CITOMETRY IN BRAZILIAN PATIENTS WITH ANKYLOSING SPONDYLITIS AND PSORIATIC ARTHRITIS(2013) GOLDENSTEIN-SCHAINBERG, C.; CARRASCO, S.; SAAD, C.; GONCALVES, C.; SAMPAIO-BARROS, P.; PARRA, E.conferenceObject TRIDIMENSIONAL RECONSTRUCTION, BIOCHEMICAL AND MOLECULAR PROFILE OF COLLAGEN V IN SKIN AND LUNG FIBROBLASTS CULTURE FROM SSc INDICATE A FAILING IN FIBRILOGENESIS(2012) TEODORO, W.; MORAIS, J.; MARTIN, P.; VELOSA, A. P. P.; CARRASCO, S.; SOUZA, R. B. C.; KATAYAMA, M. L.; GOLDEINSTEIN-SCHAINBERG, C.; PARRA, E. R.; CAPELOZZI, V. L.; YOSHINARI, N. H.Background. The type V collagen (COL V) mutations are involved in collagen vascular diseases, such as SSc, in which an unusual accumulation of this collagen was demonstrated (Pathol Res Pract 2004; 200:681). In this context, our purpose was to analyse the 3D reconstruction, biochemical and molecular profile of COL Vα1 and α2 chains in skin and lung fibroblasts culture from patients with SSc. Methods. Lung biopsies of seven patients, skin of six patients and respective matched controls were obtained from SSc according ACR. For fibroblast culture skin and lung evaluations were used the following score: intense expression (1–4), fibroblast number/field (1, 2) and collagen fibres architecture (1–3). The total evaluations were: mild (3–5), moderate (6, 7) and severe (8, 9). COLV 3D reconstruction was performed by confocal microscopy, COL Vα1 and Vα2 gene expression in fibroblasts of skin and lung was performed in PCR–RT and COLV protein expression by immunoblotting. Results. The structure of COL V fibre in 3D reconstruction showed distorted and strongly thickened fibres in skin and lung fibroblasts with irregular bundles of COL V distributed in parallel and perpendicular arrangements resulting in a dense network in SSc patients compared with thin fibres pattern from the healthy controls. Collagen quantification showed increase of COL V fibres expression in SSc cutaneous fibroblast [82.5 (9.5)% vs 47.5 (9.5)%, P = 0.002] and lung fibroblast 38.87 (2.99)% vs 20.33 (7.50)%, P = 0,002) compared with respective controls. The molecular evaluation demonstrated an increased of COL Vα1 and α2 mRNA expression in SSc fibroblast skin when compared with control [1.375 (0.373) au vs 0.0047 (0.0013) au, P = 0.05). Similar results were observed in lung [1.61 (0.654) vs 0.99 (0.51) au; P = 0.05).The proportion COL Vα1/COL Vα2 mRNA in fibroblast lung and skin was higher in SSc than in controls being the chains ratio 1 : 2. COL V chains from skin and lung fibroblasts presented alteration of molecular weight of the quoted chain. Conclusion. The overexpression and the unusual organization of COLV fibres, besides the biochemical changes, suggest an interference with the fibrillogenesis process in skin and pulmonary fibrosis from SSc patients, reinforcing the participation of this collagen in pathogenesis of SSc and open new therapeutic perspectives for these patients.- Abnormal collagen V deposition in dermis correlates with skin thickening and disease activity in systemic sclerosis(2012) MARTIN, Patricia; TEODORO, Walcy R.; VELOSA, Ana Paula P.; MORAIS, Jymenez de; CARRASCO, Solange; CHRISTMANN, Romy B.; GOLDENSTEIN-SCHAINBERG, Claudia; PARRA, Edwin R.; KATAYAMA, Maria Lucia; SOTTO, Mirian N.; CAPELOZZI, Vera L.; YOSHINARI, Natalino H.Objective: The physiological and mechanical properties of the skin, the primary tissue affected by systemic sclerosis, depend on the assembly of collagen types I, Ill and V, which form heterotypic fibers. Collagen V (COLV) regulates heterotypic fiber diameter, and the maintenance of its properties is important for maintaining normal tissue architecture and function. Based on a COLV-induced experimental SSc model, in which overexpression of abnormal COLV was a prominent feature, we assumed that this abnormality could be present in SSc patients and could be correlated to disease duration, skin thickening and disease activity. Methods: Skin biopsies from 18 patients (6 early-stage and 12 late-stage) and 10 healthy controls were studied. Skin thickening assessment was performed with the Modified Rodnan Skin Score (MRSS), and activity was calculated using the Valentini Disease Activity Index. Morphology, morphometry of COLV deposition in dermis, as well as, quantitative RT-PCR and 3D-reconstruction of the dermal fibroblast culture were performed. Results: Structurally abnormal COLV was overexpressed in SSc skin, mainly in the early stages of the disease, when compared to normal controls and late-stage. A positive correlation between COLV expression and MRSS and disease activity was observed. Collagen V alpha-1 and alpha-2 mRNA expression levels were higher in SSc. Tridimensional reconstruction of SSc dermal heterotypic fibers confirmed the presence of atypical COLV. Conclusion: Increased synthesis of abnormal COLV and its correlation with disease stage, activity and MRSS suggest that this collagen can be a possible trigger involved in the pathogenesis of SSc.