Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 17 de 17
Filter
1.
Braz. oral res. (Online) ; 34: e007, 2020. graf
Article in English | LILACS | ID: biblio-1089397

ABSTRACT

Abstract The aim of this study was to assess the influence of cyclosporine administration on the repair of critical-sized calvaria defects (CSDs) in rat calvaria filled with diverse biomaterials. Sixty animals were divided into two groups: the control (CTR) group (saline solution) and the cyclosporine (CCP) group (cyclosporine, 10 mg/kg/day). These medications were administered daily by gavage, beginning 15 days before the surgical procedure and lasting until the day the animals were euthanized. A CSD (5 mm Ø) was made in the calvaria of each animal, which was allocated to one of 3 subgroups, according to the biomaterial used to fill the defect: coagulum (COA), deproteinized bovine bone (DBB), or biphasic calcium phosphate ceramics of hydroxyapatite and β-phosphate tricalcium (HA/TCP). Euthanasia of the animals was performed 15 and 60 days after the surgical procedure (n = 5 animals/period/subgroup). Bone repair (formation) assessment was performed through microtomography and histometry, while the analyses of the expression of the BMP2, Osteocalcin, and TGFβ1 proteins were performed using immunohistochemistry. The CSDs not filled with biomaterials demonstrated lower bone formation in the CCP group. At 15 days, less bone formation was observed in the CSDs filled with DBB, a smaller volume of mineralized tissue was observed in the CSDs filled with HA/TCP, and the expression levels of BMP2 and osteocalcin were lower in the CCP group compared to the CTR group. The use of cyclosporine impaired bone repair in CSD, and this effect can be partially explained by the suppression of BMP2 and osteocalcin expression.


Subject(s)
Animals , Male , Rats , Osteogenesis/drug effects , Bone Regeneration/drug effects , Cyclosporine/pharmacology , Bone Substitutes/pharmacology , Calcineurin Inhibitors/pharmacology , Skull/drug effects , Skull/pathology , Time Factors , Immunohistochemistry , Random Allocation , Osteocalcin/analysis , Reproducibility of Results , Transforming Growth Factor beta1/analysis , Bone Morphogenetic Protein 2/analysis , X-Ray Microtomography
2.
Braz. oral res. (Online) ; 34: e007, 2020. graf
Article in English | LILACS | ID: biblio-1055531

ABSTRACT

Abstract The aim of this study was to assess the influence of cyclosporine administration on the repair of critical-sized calvaria defects (CSDs) in rat calvaria filled with diverse biomaterials. Sixty animals were divided into two groups: the control (CTR) group (saline solution) and the cyclosporine (CCP) group (cyclosporine, 10 mg/kg/day). These medications were administered daily by gavage, beginning 15 days before the surgical procedure and lasting until the day the animals were euthanized. A CSD (5 mm Ø) was made in the calvaria of each animal, which was allocated to one of 3 subgroups, according to the biomaterial used to fill the defect: coagulum (COA), deproteinized bovine bone (DBB), or biphasic calcium phosphate ceramics of hydroxyapatite and β-phosphate tricalcium (HA/TCP). Euthanasia of the animals was performed 15 and 60 days after the surgical procedure (n = 5 animals/period/subgroup). Bone repair (formation) assessment was performed through microtomography and histometry, while the analyses of the expression of the BMP2, Osteocalcin, and TGFβ1 proteins were performed using immunohistochemistry. The CSDs not filled with biomaterials demonstrated lower bone formation in the CCP group. At 15 days, less bone formation was observed in the CSDs filled with DBB, a smaller volume of mineralized tissue was observed in the CSDs filled with HA/TCP, and the expression levels of BMP2 and osteocalcin were lower in the CCP group compared to the CTR group. The use of cyclosporine impaired bone repair in CSD, and this effect can be partially explained by the suppression of BMP2 and osteocalcin expression.


Subject(s)
Animals , Male , Rats , Osteogenesis/drug effects , Bone Regeneration/drug effects , Cyclosporine/pharmacology , Bone Substitutes/pharmacology , Calcineurin Inhibitors/pharmacology , Skull/drug effects , Skull/pathology , Time Factors , Immunohistochemistry , Random Allocation , Osteocalcin/analysis , Reproducibility of Results , Transforming Growth Factor beta1/analysis , Bone Morphogenetic Protein 2/analysis , X-Ray Microtomography
3.
Acta cir. bras ; 33(2): 156-162, Feb. 2018. tab, graf
Article in English | LILACS | ID: biblio-886258

ABSTRACT

Abstract Purpose: To evaluate the concentration of transforming growth factor beta 1 (TGFB1) levels in a rat pleural effusion obtained by inoculation of intrapleural bacteria or turpentine through thoracentesis. Methods: Thirty-Nine Wistar rats were divided into three groups: Staphylococcus aureus (SA, n = 17); Streptococcus pneumoniae (SP, n = 12); and turpentine (control, n = 10). Pleural fluid was collected through ultrasound-guided thoracentesis 12 h, 24 h, and 36 h after instillation of bacteria or turpentine. Levels of TGFB1 were measured in pleural fluid. Results: At 12 h, mean TGFB1concentrations were 5.3450 pg/mL in the SA group, 5.3449 pg/mL in the SP group, and 5.3450 pg/mL in controls. At 24 h, they were 4.6700 pg/mL in the SA group, 4.6700 pg/mL in the SP group, and 4.6700 pg/mL in controls. At 36 h, they were 4.6699 pg/mL in the SA group and in control. No difference was observed among the groups in mean TGFB1concentration (p = 0.12); however, a significant intragroup reduction in mean TGFB1 was observed between 12 and 24 h (p < 0.01). Conclusion: The transforming growth factor beta 1 concentrations were not useful as a diagnostic tool or an early marker of infected pleural effusion.


Subject(s)
Animals , Male , Rats , Pleural Effusion/diagnosis , Empyema, Pleural/diagnosis , Transforming Growth Factor beta1/analysis , Pleural Effusion/complications , Bacteria/pathogenicity , Biomarkers/analysis , Empyema, Pleural/complications , Empyema, Pleural/microbiology , Rats, Wistar , Disease Models, Animal
4.
Biol. Res ; 51: 58, 2018. graf
Article in English | LILACS | ID: biblio-1011402

ABSTRACT

BACKGROUND: Studies have demonstrated that transforming growth factor beta-1 (TGF-ß1) exhibits oncogenic activity in different types of cancer, including ovarian cancer (OC). However, its regulatory mechanism in OC and whether TGF-ß1 is involved in chemosensitivity regulation remains unclear. Thus, the aim of this study was to investigate the role of TGF-ß1 in OC. METHODS: The OC cell line SKOV3 was employed, and TGF-ß1 overexpression or knockdown vectors were constructed. The cell proliferation of SKOV3 was evaluated with the cell counting kit (CCK8) kit after treatment with different concentrations of cis-platinum. Western blot and protein immunoprecipitation were employed to detect changes in BRCA1 and Smad3 expression and their interactions. Tumor growth in nude mice was evaluated. RESULTS: The results showed that TGF-ß1 knockdown increased chemosensitivity by promoting BRCA1 expression and Smad3 phosphorylation. In vivo studies showed that TGF-ß1 knockdown significantly inhibited the growth of tumors, also by upregulating BRCA1 expression and Smad3 phosphorylation. CONCLUSION: Taken together, our results suggest that TGF-ß1 knockdown inhibits tumor growth and increases chemosensitivity by promotion of BRCA1/Smad3 signaling.


Subject(s)
Humans , Animals , Male , Female , Ovarian Neoplasms/metabolism , Down-Regulation/physiology , Genes, BRCA1/physiology , Smad3 Protein/physiology , Transforming Growth Factor beta1/physiology , Ovarian Neoplasms/pathology , Ovarian Neoplasms/drug therapy , Immunohistochemistry , Cells, Cultured , Blotting, Western , Drug Resistance, Neoplasm/physiology , Tumor Suppressor Proteins/physiology , Cell Line, Tumor , Cell Proliferation , Smad3 Protein/analysis , Transforming Growth Factor beta1/analysis , Gene Knockdown Techniques , Real-Time Polymerase Chain Reaction , Mice, Inbred BALB C
5.
São Paulo; s.n; s.n; 2018. 93 p. ilus, graf, tab.
Thesis in Portuguese | LILACS | ID: biblio-967928

ABSTRACT

O fator de crescimento transformante beta tipo 1, TGF-ß1, é uma proteína extracelular homodimérica secretada por vários tipos celulares, que pode ter ação parácrina ou endócrina. Essa proteína está envolvida em processos celulares de diferenciação, proliferação, mobilidade e formação de matriz extracelular. Além disso, é parte importante dos processos de regeneração tecidual, atuando, de maneira decisiva, no reparo, atraindo macrófagos e fibroblastos para o local da injúria e estimulando a angiogênese. Assim, considerando o papel desse peptídeo no processo regenerativo, o uso de TGF-ß1 como proteína terapêutica na área de Bioengenharia Tecidual é bastante promissor. Apesar disso, a venda dessa proteína, para fins terapêuticos, é inexistente no mercado e a proteína recombinante vendida, que só pode ser utilizada em pesquisas científicas, não é produzida nacionalmente e chega a custar R$200.000,00/mg. Nesse contexto, o objetivo do presente trabalho é desenvolver uma metodologia de produção do fator recombinante TGF-ß1 em células de ovário de hamster chinês (CHO), visando à obtenção de níveis altos de rendimento, e, futuramente, a transferência da tecnologia de produção para a iniciativa privada, tornando possível seu uso na Medicina Regenerativa, sozinho ou em combinação com outros fatores de crescimento. O cDNA de TGF-ß1 foi amplificado a partir de um banco de cDNA humano e clonado no vetor proprietário pNU1 de expressão de mamífero. A construção pNU1/TGF-ß1 foi utilizada para transfectar estavelmente células CHO DG44 e uma estratégia de co-amplificação foi utilizada para selecionar células transfectantes com maior número de cópias da sequência correspondente a TGF-ß1. Estas culturas foram submetidas ao processo de amplificação gênica com concentrações crescentes de metotrexato. Ensaios de Western Blot e ELISA foram realizados utilizando-se o meio condicionado pelas populações selecionadas e por clones superprodutores. Entre os 41clones obtidos, cinco apresentaram maiores níveis de produção de TGF-ß1, entre 1.000 e 2.000 ng/mL. Estes clones foram selecionados para a realização de testes de atividade in vitro utilizando-se células A549, que permitem avaliar a transição epitélio-mesênquima. Um ensaio de cicatrização de feridas em peles do dorso de camundongos foi padronizado e utilizado para avaliar a atividade in vivo do clone que apresentou melhor resultado in vitro. A proteína TGF-ß1 foi parcialmente purificada por HPLC em uma coluna de afinidade. Portanto, a proteína TGF-ß1 humana recombinante foi produzida, apresentando atividade biológica in vitro e in vivo, sendo capaz de reparar eficientemente feridas cutâneas. Essa iniciativa pode oferecer aos pacientes uma alternativa para o tratamento de lesões teciduais, acelerando a cicatrização de feridas e o reparo de tecidos


The transforming growth factor beta 1, TGF-ß1, is a homodimeric extracellular protein secreted by several cell types, which may have paracrine or endocrine action. This protein is involved in cellular processes of differentiation, proliferation, mobility and formation of extracellular matrix. In addition, it is an important part of the tissue regeneration processes, acting decisively on repair, attracting macrophages and fibroblasts to the site of injury and stimulating angiogenesis. Therefore, considering the role of this peptide in the regenerative process and the use of TGF-ß1 as a therapeutic protein in the field of Tissue Bioengineering is very promising. Despite this, the sale of this protein for therapeutic purposes is nonexistent in the market and the recombinant protein available in the market, which can only be used in scientific research, is not produced nationally and the costs are in the order of R$ 200,000.00/mg. In this context, the objective of the present work is to develop a methodology for the production of the TGF-ß1 recombinant factor in Chinese hamster ovary (CHO) cells, aiming at obtaining high yields, and, in the future, transfering the production technology to the private initiative, allowing its use in Regenerative Medicine, alone or in combination with other growth factors. The TGF-ß1 cDNA was amplified from a human cDNA library and cloned into the proprietary pNU1 mammalian expression vector. The pNU1/TGF-ß1 construct was used to stably transfect CHO DG44 cells, and a co-amplification strategy was used to select transfectant cells with the largest number of gene copies. These cultures were subjected to the process of gene amplification with methotrexate. Western Blot and ELISA were used to assay the conditioned medium obtained from the selected cell populations and from overproducing cell clones. Among the 41 clones obtained, five presented higher levels of TGF-ß1 production, between 1,000 and 2,000 ng/mL. These clones were selected for in vitro activity testing using A549 cells to evaluate the epithelial-mesenchymal transition. Awound healing assay on mouse dorsal skin was standardized and used to evaluate the in vivo activity of the cell clone which displayed the highest result in vitro. The TGF-ß1 protein was partially purified by HPLC on an affinity column. Therefore, the recombinant human TGF-ß1 protein was produced and shown to display biological activity both in vitro and in vivo, being able to eficiently repair cutaneous wounds. This initiative may provide patients with an alternative treatment for tissue damage, accelerating wound healing and tissue repair


Subject(s)
Animals , Mice , CHO Cells/chemistry , Transforming Growth Factor beta1/analysis , Enzyme-Linked Immunosorbent Assay/instrumentation , Blotting, Western/instrumentation , Regenerative Medicine/trends , Latent TGF-beta Binding Proteins
6.
Braz. oral res. (Online) ; 31: e51, 2017. tab, graf
Article in English | LILACS | ID: biblio-952089

ABSTRACT

Abstract The objective of this study was to evaluate the expression of matrix metalloproteinase 9 (MMP-9) and transforming growth factor beta (TGF-β1) in periapical lesion samples correlated with the intensity of the inflammatory infiltrate and thickness of the epithelial lining. Forty-five cases of periapical lesions (23 periapical granulomas and 22 radicular cysts) were subjected to morphological and immunohistochemical analyses using anti-MMP-9 and anti-TGF-β1 antibodies. The data were analyzed using the following tests: non-parametric Mann-Whitney, chi-square, Fisher's exact test and Spearman's correlation test (P<0.05). Analysis of inflammatory infiltrate revealed that 78% of periapical granulomas presented infiltrate grade III, in contrast with 32% of radicular cysts (P<0.001). Morphological evaluation of the epithelial thickness in radicular cysts revealed the presence of atrophic epithelium in 86% of the cysts. The immunostaining of MMP-9 was score 2 in 67% of the granulomas and 77% of the cysts. Both lesions were predominantly score 1 for TGF-β1. Significant differences were confirmed between the expression scores of TGF-β1 and MMP-9 in periapical granulomas (p = 0.004) and in radicular cysts (p < 0.001). Expression of TGF-β1 was different for periapical granulomas and radicular cysts. This immunoregulatory cytokine seems more representative in asymptomatic lesions. The extracellular matrix remodeling process dependent on MMP-9 seems to be similar for both periapical granulomas and radicular cysts. TGF-β1 and MMP-9 may play an important role in the maintenance of periapical lesions.


Subject(s)
Humans , Male , Female , Adult , Periapical Granuloma/metabolism , Radicular Cyst/chemistry , Matrix Metalloproteinase 9/analysis , Transforming Growth Factor beta1/analysis , Periapical Granuloma/immunology , Periapical Granuloma/pathology , Biopsy , Severity of Illness Index , Immunohistochemistry/methods , Radicular Cyst/immunology , Radicular Cyst/pathology , Statistics, Nonparametric , Epithelial Cells/pathology
7.
Braz. j. med. biol. res ; 49(10): e5373, 2016. graf
Article in English | LILACS | ID: lil-792522

ABSTRACT

Stem cells from human exfoliated deciduous teeth (SHEDs) have great potential to treat various dental-related diseases in regenerative medicine. They are usually maintained with 10% fetal bovine serum (FBS) in vitro. Modified platelet-rich plasma (mPRP) would be a safe alternative to 10% FBS during SHEDs culture. Therefore, our study aimed to compare the proliferation and differentiation of SHEDs cultured in mPRP and FBS medium to explore an optimal concentration of mPRP for SHEDs maintenance. Platelets were harvested by automatic blood cell analyzer and activated by repeated liquid nitrogen freezing and thawing. The platelet-related cytokines were examined and analyzed by ELISA. SHEDs were extracted and cultured with different concentrations of mPRP or 10% FBS medium. Alkaline phosphatase (ALP) activity was measured. Mineralization factors, RUNX2 and OCN, were measured by real-time PCR. SHEDs were characterized with mesenchymal stem cells (MSCs) markers including vimentin, CD44, and CD105. mPRP at different concentrations (2, 5, 10, and 20%) enhanced the growth of SHEDs. Moreover, mPRP significantly stimulated ALP activity and promoted expression of RUNX2 and OCN compared with 10% FBS. mPRP could efficiently facilitate proliferation and differentiation of SHEDs, and 2% mPRP would be an optimal substitute for 10% FBS during SHEDs expansion and differentiation in clinical scale manufacturing.


Subject(s)
Humans , Animals , Cattle , Cell Proliferation/physiology , Dental Pulp/cytology , Mesenchymal Stem Cells/cytology , Platelet-Rich Plasma , Tooth, Deciduous/cytology , Alkaline Phosphatase/antagonists & inhibitors , Analysis of Variance , Cell Culture Techniques/methods , Cell Differentiation/physiology , Cells, Cultured , Core Binding Factor Alpha 1 Subunit/analysis , Culture Media , Enzyme-Linked Immunosorbent Assay , Platelet-Derived Growth Factor/analysis , Real-Time Polymerase Chain Reaction , Reproducibility of Results , Time Factors , Transforming Growth Factor beta1/analysis
8.
Braz. j. med. biol. res ; 49(10): e5526, 2016. graf
Article in English | LILACS | ID: lil-792523

ABSTRACT

Pseudomonas aeruginosa is one of the common colonizing bacteria of the human body and is an opportunistic pathogen frequently associated with respiratory infections. Inactivated P. aeruginosa (IPA) have a variety of biological effects against inflammation and allergy. Transforming growth factor-β (TGF-β) signaling plays a critical role in the regulation of cell growth, differentiation, and development in a wide range of biological systems. The present study was designed to investigate the effects of IPA on TGF-β/Smad signaling in vivo, using a hypoxia-induced pulmonary hypertension (PH) rat model. Sprague Dawley rats (n=40) were exposed to 10% oxygen for 21 days to induce PH. At the same time, IPA was administered intravenously from day 1 to day 14. Mean pulmonary artery pressure (mPAP) and the right ventricle (RV) to left ventricle plus the interventricular septum (LV+S) mass ratio were used to evaluate the development of PH. Vessel thickness and density were measured using immunohistochemistry. Primary arterial smooth muscle cells (PASMCs) were isolated and the proliferation of PASMCs was assayed by flow cytometry. The production of TGF-β1 in cultured supernatant of PASMCs was assayed by ELISA. The expression levels of α-smooth muscle actin (α-SMA), TGF-β1 and phospho-Smad 2/3 in PASMCs were assayed by western blot. Our data indicated that IPA attenuated PH, RV hypertrophy and pulmonary vascular remodeling in rats, which was probably mediated by restraining the hypoxia-induced overactive TGF-β1/Smad signaling. In conclusion, IPA is a promising protective treatment in PH due to the inhibiting effects on TGF-β1/Smad 2/3 signaling.


Subject(s)
Animals , Male , Hypertension, Pulmonary/metabolism , Hypertension, Pulmonary/prevention & control , Hypoxia/metabolism , Myocytes, Smooth Muscle/physiology , Pseudomonas aeruginosa/physiology , Smad Proteins/metabolism , Transforming Growth Factor beta1/metabolism , Actins/analysis , Actins/metabolism , Blotting, Western , Cell Proliferation/physiology , Enzyme-Linked Immunosorbent Assay , Flow Cytometry , Hypertension, Pulmonary/etiology , Hypoxia/complications , Immunohistochemistry , Rats, Sprague-Dawley , Reproducibility of Results , Signal Transduction/physiology , Smad Proteins/analysis , Transforming Growth Factor beta1/analysis
9.
Braz. oral res. (Online) ; 30(1): e93, 2016. graf
Article in English | LILACS | ID: biblio-952019

ABSTRACT

Abstract The aim of the present study was to evaluate the expression of transforming growth factor-β1 (TGF-β1) and osteonectin (ON) in pulp-like tissues developed by tissue engineering and to compare it with the expression of these proteins in pulps treated with Ca(OH)2 therapy. Tooth slices were obtained from non-carious human third molars under sterile procedures. The residual periodontal and pulp soft tissues were removed. Empty pulp spaces of the tooth slice were filled with sodium chloride particles (250-425 µm). PLLA solubilized in 5% chloroform was applied over the salt particles. The tooth slice/scaffold (TS/S) set was stored overnight and then rinsed thoroughly to wash out the salt. Scaffolds were previously sterilized with ethanol (100-70°) and washed with phosphate-buffered saline (PBS). TS/S was treated with 10% EDTA and seeded with dental pulp stem cells (DPSC). Then, TS/S was implanted into the dorsum of immunodeficient mice for 28 days. Human third molars previously treated with Ca(OH)2 for 90 days were also evaluated. Samples were prepared and submitted to histological and immunohistochemical (with anti-TGF-β1, 1:100 and anti-ON, 1:350) analyses. After 28 days, TS/S showed morphological characteristics similar to those observed in dental pulp treated with Ca(OH)2. Ca(OH)2-treated pulps showed the usual repaired pulp characteristics. In TS/S, newly formed tissues and pre-dentin was colored, which elucidated the expression of TGF-β1 and ON. Immunohistochemistry staining of Ca(OH)2-treated pulps showed the same expression patterns. The extracellular matrix displayed a fibrillar pattern under both conditions. Regenerative events in the pulp seem to follow a similar pattern of TGF-β1 and ON expression as the repair processes.


Subject(s)
Humans , Animals , Mice , Stem Cells/drug effects , Calcium Hydroxide/pharmacology , Osteonectin/analysis , Dental Pulp/drug effects , Transforming Growth Factor beta1/analysis , Time Factors , Calcium Hydroxide/therapeutic use , Immunohistochemistry , Osteonectin/drug effects , Cells, Cultured , Reproducibility of Results , Tissue Engineering/methods , Dental Pulp/cytology , Dentin/drug effects , Guided Tissue Regeneration/methods , Extracellular Matrix/drug effects , Transforming Growth Factor beta1/drug effects , Tissue Scaffolds , Odontoblasts/drug effects
10.
Yonsei Medical Journal ; : 1597-1603, 2015.
Article in English | WPRIM | ID: wpr-177064

ABSTRACT

PURPOSE: The aim of this study was to examine the effects of all-trans retinoic acid (ATRA) on diabetic nephropathy. MATERIALS AND METHODS: We measured amounts of urinary albumin excretion (UAE) after administrating ATRA to Otsuka Long-Evans Tokushima Fatty (OLETF) rats. In order to understand the mechanism of action for ATRA, we administrated ATRA to examine its inhibitory action on the production of transforming growth factor-beta1 (TGF-beta1), protein kinase C (PKC), and reactive oxidative stress (ROS) in cultured rat mesangial cells (RMCs). RESULTS: After 16 weeks of treatment, UAE was lower in the ATRA-treated OLETF rats than in the non-treated OLETF rats (0.07+/-0.03 mg/mgCr vs. 0.17+/-0.15 mg/mgCr, p<0.01). After incubation of RMCs in media containing 30 or 5 mM of glucose, treatment with ATRA showed time- and dose-dependent decreases in TGF-beta1 levels and ROS. Moreover, ATRA treatment showed a dose-dependent decrease in PKC expression. CONCLUSION: ATRA treatment suppressed UAE and TGF-beta1 synthesis, which was mediated by significant reductions in PKC activity and ROS production. Our results suggest that ATRA has a potential therapeutic role for diabetic nephropathy.


Subject(s)
Animals , Diabetes Mellitus, Type 2/complications , Diabetic Nephropathies/complications , Mesangial Cells/metabolism , Oxidative Stress/drug effects , Rats , Rats, Inbred OLETF , Reactive Oxygen Species/metabolism , Transforming Growth Factor beta1/analysis , Tretinoin/pharmacology
11.
Clinics ; 67(11): 1259-1263, Nov. 2012. ilus, tab
Article in English | LILACS | ID: lil-656714

ABSTRACT

OBJECTIVE: Pleural tuberculosis is the most frequently occurring form of extra pulmonary disease in adults. In up to 40% of cases, the lung parenchyma is concomitantly involved, which can have an epidemiological impact. This study aims to evaluate the pleural and systemic inflammatory response of patients with pleural or pleuropulmonary tuberculosis. METHODS: A prospective study of 39 patients with confirmed pleural tuberculosis. After thoracentesis, a high resolution chest tomography was performed to evaluate the pulmonary involvement. Of the 39 patients, 20 exhibited only pleural effusion, and high resolution chest tomography revealed active associated-pulmonary disease in 19 patients. The total protein, lactic dehydrogenase, adenosine deaminase, vascular endothelial growth factor, interleukin-8, tumor necrosis factor-α, and transforming growth factor-β1 levels were quantified in the patient serum and pleural fluid. RESULTS: All of the effusions were exudates with high levels of adenosine deaminase. The levels of vascular endothelial growth factor and transforming growth factor-β1 were increased in the blood and pleural fluid of all of the patients with pleural tuberculosis, with no differences between the two forms of tuberculosis. The tumor necrosis factor-α levels were significantly higher in the pleural fluid of the patients with the pleuropulmonary form of tuberculosis. The interleukin-8 levels were high in the pleural fluid of all of the patients, without any differences between the forms of tuberculosis. CONCLUSION: Tumor necrosis factor-α was the single cytokine that significantly increased in the pleural fluid of the patients with pulmonary involvement. However, an overlap in the results does not permit us to suggest that cytokine is a biological marker of concomitant parenchymal involvement. Although high resolution chest tomography can be useful in identifying these patients, the investigation of fast acid bacilli and cultures for M. tuberculosis in the sputum is recommended for all patients who are diagnosed with pleural tuberculosis.


Subject(s)
Adult , Humans , Middle Aged , Young Adult , Biomarkers/analysis , Pleural Effusion/metabolism , Tuberculosis, Pleural/metabolism , Adenosine Deaminase/analysis , Cytokines/analysis , Disease Progression , Enzyme-Linked Immunosorbent Assay , Exudates and Transudates/chemistry , Oxidoreductases/analysis , Prospective Studies , Pleural Effusion , Transforming Growth Factor beta1/analysis , Tuberculosis, Pleural , Tuberculosis, Pulmonary/metabolism , Tumor Necrosis Factor-alpha/analysis , Vascular Endothelial Growth Factor A/analysis
12.
São Paulo; s.n; s.n; 2011. 185 p. tab, graf, ilus.
Thesis in Portuguese | LILACS | ID: biblio-847632

ABSTRACT

A causa de morte da maioria das pacientes com câncer de mama se deve à doença metastática desenvolvida a partir do tumor primário. A degradação dos componentes da matriz extracelular (MEC), um dos principais eventos do processo metastático, é regulada pelo balanço entre as atividades das metaloproteinases de matriz (MMPs) e dos seus inibidores, tanto os inibidores teciduais (TIMPs) como o inibidor associado à membrana (RECK). Contudo, ainda existe pouca informação sobre os mecanismos moleculares responsáveis pela manutenção deste balanço. No presente trabalho, foi investigado o envolvimento de TGF-ß1 (Transforming Growth Factor-ß1), uma citocina multifuncional é capaz tanto de inibir o crescimento celular, quanto de promover invasão e metástase, dependendo do estadiamento e do tipo de tumor, na regulação da expressão de MMPs, TIMPs e RECK, em modelo de câncer de mama. Primeiramente, examinou-se os níveis de expressão de mRNA das isoformas e receptores de TGF-ß, em um painel de cinco linhagens de carcinoma mamário humano, com diferentes potenciais invasivos e metastáticos, por qRT-PCR. Os resultados obtidos demonstraram uma correlação positiva entre a expressão dessas moléculas, e a progressão do caráter invasivo e metastático celular. Em seguida, a linhagem altamente invasiva, MDA-MB-231, foi tratada com diferentes concentrações de TGF-ß1 recombinante. Esta citocina foi capaz de modular a expressão gênica de MMPs (MMP-2 e MMP-9) e de seus inibidores (TIMP- 2 e RECK). Tanto ERK½, quanto p38MAPK mostraram-se envolvidas neste mecanismo. Foi demonstrado que a inibição da atividade de ERK½ alterou a expressão das proteínas MMP-9, TIMP-2 e RECK, enquanto o bloqueio de p38 MAPK afetou os níveis protéicos de MMP-2 e TIMP-2. O aumento do potencial migratório e invasivo da linhagem MDA-MB-231, induzido por TGF-ß1, mostrou-se também dependente da atividade de MMPs, ERK½ e p38MAPK. Dada a ausência de informações sobre o papel de RECK em modelo mamário, a função deste inibidor de MMPs também foi investigada. Primeiramente, analisou-se a expressão de RECK ao longo do desenvolvimento da mama e, posteriormente, em 1040 amostras tumorais de mama humana, através da metodologia de Tissue Microarray, tendo sido possível demonstrar que a alta expressão de RECK associa-se a menor tempo de sobrevida global e livre de doença em 10 anos. Os resultados obtidos indicaram que a expressão da proteína RECK, em oposição ao verificado em outros tipos de tumores, está relacionada ao fenótipo mais agressivo de tumores de mama. Entretanto, a análise funcional de RECK, realizada por meio da utilização de vetores shRNA específicos para a inibição desta proteína, demonstrou que RECK também atua como um inibidor de invasão celular e da expressão de MMP-9, na linhagem MDA-MB-231. Em conjunto, os resultados obtidos neste trabalho contribuíram para a elucidação dos mecanismos moleculares de regulação de RECK, por clássicas moléculas associadas ao processo de tumorigênese (TGF-ß1 e MAPKs), bem como para o esclarecimento de suas funções em modelo mamário, sugerindo-o como mais um promissor candidato a marcador prognóstico e alvo molecular para a terapia do câncer de mama


The metastatic disease is the main mortality cause of breast cancer patients. The metastatic process involves a complex cascade of events, including the organized breakdown of the extracellular matrix (ECM) compounds. The degradation of ECM is tightly regulated by the balance between the activities of matrix metalloproteinases (MMPs) and their inhibitors, the tissue inhibitors (TIMPs) and the membrane-associated inhibitor (RECK). Among the several molecules released and activated by ECM remodeling, TGF-ß1 (Transforming Growth Factor-ß1) is a multifunctional cytokine able to regulate both cell growth inhibition and invasion and metastasis promotion, depending on the tumor stage and type. Since the molecular mechanisms involved in the ECM remodeling control are still not completed understood, in this study, we investigated the involvement of TGF-ß1 in regulating of MMPs, TIMPs and RECK expression, in the breast cancer model. By qRT-PCR, we first examined the gene expression levels of TGF-ß isoforms and receptors, in a panel of five human breast cancer cell lines displaying different degrees of invasiveness and metastatic potential. Our results suggest a positive correlation between the mRNA expression of these molecules and the breast cancer progression. Moreover, the highly invasive breast cancer cell line MDA-MB-231 was treated with different concentrations of recombinant TGF-ß1. We described that this cytokine was able to modulate the gene expression of MMPs (MMP-2 and MMP-9) and MMPs inhibitors (TIMP-2 and RECK) at both the mRNA and protein levels, with ERK½ and p38 MAPK being involved in this molecular mechanism. However, while ERK½ activity inhibition altered MMP-9, TIMP-2 and RECK expression, the p38 MAPK blockage affected the protein levels of MMP-2 and TIMP-2. Finally, we reposted that the TGF-ß1-enhanced migration and invasion capacities of MDA-MB- 231 cells were blocked by MMPs, ERK½ and p38 MAPK inhibitors. Analysis of the RECK function in the breast model was also an objective of this study. We analyzed RECK expression during mammary gland development. We evaluated the RECK protein profile in 1040 breast tumor tissue samples using Tissue Microarray assays. We demonstrated that high expression levels of RECK were associated with shorter overall and disease-free survival in 10 years. Moreover, we verified that RECK is a biomarker of poor prognosis mainly for patients diagnosed with less aggressive breast tumor. Therefore, in contrast to other tumor types, our results indicate that high protein expression levels of RECK are related to a more aggressive phenotype. In fact, the RECK functional analysis, performed by using of shRNA vectors, showed that RECK function remains as an inhibitor of cellular invasion and MMP-9 expression, in MDA-MB-231 cells. Taken together, our results contribute to better understanding of the molecular mechanisms associated to RECK regulation by TGF-ß1 and MAPK as well as to clarify its role in breast model. Thus, we suggests RECK as a new and promising prognostic marker and molecular target candidate for breast cancer therapy


Subject(s)
Breast Neoplasms/pathology , Matrix Metalloproteinases/analysis , Receptors, Transforming Growth Factor beta/administration & dosage , Transforming Growth Factor beta1/analysis , Gene Expression/genetics , Mammary Analogue Secretory Carcinoma/prevention & control , Matrix Metalloproteinase 17/analysis , Matrix Metalloproteinase 2 , RNA, Messenger/analysis , Tissue Inhibitor of Metalloproteinases/analysis
13.
Article in English | WPRIM | ID: wpr-73118

ABSTRACT

Diabetic nephropathy is the most serious complication in diabetes mellitus. It is known that oxidative stress and inflammation play a central role in the development of diabetic nephropathy. In this study, we investigated that ferulic acid (FA) known as anti-oxidative agent could effect on diabetic nephropathy by anti-oxidative and anti-inflammatory mechanism. We examined the effects of FA in obese diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats and non-diabetic control Long-Evans Tokushima Otsuka (LETO) rats. We treated FA to experimental rats from 26 to 45 weeks of age. We evaluated ACR, MDA and MCP-1 in 24 h urine and examined renal histopathology and morphologic change in extracted kidneys from rats. Also, we evaluated the ROS production and MCP-1 levels in cultured podocyte after FA treatment. In the FA-treated OLETF rats, blood glucose was significantly decreased and serum adiponectin levels were increased. Urinary ACR was significantly reduced in FA-treated OLETF rats compared with diabetic OLETF rats. In renal histopathology, FA-treated OLETF rats showed decreased glomerular basement membrane thickness, glomerular volume, and mesangial matrix expansion. FA treatment decreased oxidative stress markers and MCP-1 levels in 24 h urine of rats and supernatants of cultured podocyte. In conclusion, it was suggested that FA have protective and therapeutic effects on diabetic nephropathy by reducing oxidative stress and inflammation.


Subject(s)
Animals , Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Antioxidants/therapeutic use , Cells, Cultured , Chemokine CCL2/genetics , Collagen/analysis , Coumaric Acids/therapeutic use , Diabetes Mellitus, Type 2/complications , Diabetic Nephropathies/complications , Gene Expression/drug effects , Kidney/drug effects , Malondialdehyde/urine , Podocytes/drug effects , Rats , Rats, Inbred OLETF , Reactive Oxygen Species/metabolism , Transforming Growth Factor beta1/analysis
14.
Braz. oral res ; 24(2): 182-188, Apr.-June 2010. ilus, graf
Article in English | LILACS | ID: lil-553905

ABSTRACT

Cyclosporin A (CyA) induces gingival overgrowth via its stimulatory effects on expression of transforming growth factor-beta1 (TGF-â1) and collagen. It is not known whether CyA has a direct effect on gingival fibroblasts or induces its effect indirectly via stimulation of myofibroblast transdifferentiation. The present study was undertaken to examine the in vivo and in vitro effect of CyA on myofibroblast transdifferentiation. Rats were treated for 60 days with a daily subcutaneous injection of CyA, and the gingival overgrowth tissue was analyzed by immunohistochemistry. In vitro, fibroblasts from normal gingiva (NG) were cultured in the presence of different concentrations of CyA, and subjected to semi-quantitative reverse transcriptase-polymerase chain reaction and western blot. Although CyA treatment stimulated TGF-â1 expression by NG fibroblasts, it lacked to induce expression and production of isoform á of smooth muscle actin (á-SMA), the specific myofibroblast marker. The expression levels of connective tissue growth factor (CTGF), which has been considered a key molecule to promote the transdifferentiation of myofibroblasts via TGF-â1 activation, were unaffected by CyA. Our results demonstrate that CyA-induced gingival overgrowth is not associated with activation of myofibroblast transdifferentiation, since CyA is not capable to increase CTGF expression.


Subject(s)
Adult , Animals , Humans , Male , Rats , Cell Transdifferentiation/drug effects , Connective Tissue Growth Factor/metabolism , Cyclosporine/pharmacology , Fibroblasts/drug effects , Gingival Overgrowth/chemically induced , Immunosuppressive Agents/pharmacology , Actins/metabolism , Blotting, Western , Cell Culture Techniques , Culture Media , Collagen/metabolism , Connective Tissue Growth Factor/analysis , Fibroblasts/cytology , Fibroblasts/metabolism , Gingival Overgrowth/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Transforming Growth Factor beta1/analysis , Transforming Growth Factor beta1/metabolism
15.
J. appl. oral sci ; 18(1): 83-91, Jan.-Feb. 2010. ilus
Article in English | LILACS | ID: lil-545031

ABSTRACT

Myoepithelial cells have an important role in salivary gland tumor development, contributing to a low grade of aggressiveness of these tumors. Normal myoepithelial cells are known by their suppressor function presenting increased expression of extracellular matrix genes and protease inhibitors. The importance of stromal cells and growth factors during tumor initiation and progression has been highlighted by recent literature. Many tumors result from the alteration of paracrine growth factors pathways. Growth factors mediate a wide variety of biological processes such as development, tissue repair and tumorigenesis, and also contribute to cellular proliferation and transformation in neoplastic cells. OBJECTIVES: This study evaluated the expression of fibroblast growth factor-2 (FGF-2), transforming growth factor â-1 (TGFâ-1), platelet-derived growth factor-A (PDGF-A) and their respective receptors (FGFR-1, FGFR-2, TGFâR-II and PDGFR-á) in myoepithelial cells from pleomorphic adenomas (PA) by in vivo and in vitro experiments. MATERIAL AND METHODS: Serial sections were obtained from paraffin-embedded PA samples obtained from the school's files. Myoepithelial cells were obtained from explants of PA tumors provided by surgery from different donors. Immunohistochemistry, cell culture and immunofluorescence assays were used to evaluate growth factor expression. RESULTS: The present findings demonstrated that myoepithelial cells from PA were mainly positive to FGF-2 and FGFR-1 by immunohistochemistry and immunofluorescence. PDGF-A and PDGFR-á had moderate expression by immunohistochemistry and presented punctated deposits throughout cytoplasm of myoepithelial cells. FGFR-2, TGFâ-1 and TGFâR-II were negative in all samples. CONCLUSIONS: These data suggested that FGF-2 compared to the other studied growth factors has an important role in PA benign myoepithelial cells, probably contributing to proliferation of ...


Subject(s)
Adult , Female , Humans , Male , Young Adult , Adenoma, Pleomorphic/pathology , /analysis , Platelet-Derived Growth Factor/analysis , Protein-Serine-Threonine Kinases/analysis , Receptor, Fibroblast Growth Factor, Type 1/analysis , /analysis , Receptor, Platelet-Derived Growth Factor alpha/analysis , Receptors, Transforming Growth Factor beta/analysis , Salivary Gland Neoplasms/pathology , Transforming Growth Factor beta1/analysis , Actins/analysis , Cells, Cultured , Calcium-Binding Proteins/analysis , Cell Nucleus/ultrastructure , Cytoplasm/ultrastructure , Epithelial Cells/pathology , Fluorescent Antibody Technique , Immunohistochemistry , /analysis , Lip Neoplasms/pathology , Microfilament Proteins/analysis , Muscle Cells/pathology , Muscle Proteins/analysis , Muscle, Smooth/pathology , Palatal Neoplasms/pathology , Vimentin/analysis , Young Adult
16.
Article in English | WPRIM | ID: wpr-157447

ABSTRACT

For the purpose of determining the pathogenic role of transforming growth factor-beta1 (TGF-beta1) in the mechanism of chronic rheumatic heart disease, we evaluated the expression of TGF-beta1, proliferation of myofibroblasts, and changes in extracellular matrix components including collagen and proteoglycan in 30 rheumatic mitral valves and in 15 control valves. High TGF-beta1 expression was identified in 21 cases (70%) of rheumatic mitral valves, whereas only 3 cases (20%) of the control group showed high TGF-beta1 expression (p<0.001). Additionally, increased proliferation of myofibroblasts was observed in the rheumatic valves. High TGF-beta1 expression positively correlated with the proliferation of myofibroblasts (p=0.004), valvular fibrosis (p< 0.001), inflammatory cell infiltration (p=0.004), neovascularization (p=0.007), and calcification (p<0.001) in the valvular leaflets. The ratio of proteoglycan to collagen deposition inversely correlated with TGF-beta1 expression in mitral valves (p=0.040). In conclusion, an ongoing inflammatory process, the expression of TGF-beta1, and proliferation of myofibroblasts within the valves have a potential role in the valvular fibrosis, calcification, and changes in the extracellular matrix that lead to the scarring sequelae of rheumatic heart disease.


Subject(s)
Adult , Aged , Chronic Disease , Collagen/metabolism , Female , Fibrosis , Humans , Immunohistochemistry , Male , Middle Aged , Mitral Valve/pathology , Proteoglycans/metabolism , Rheumatic Heart Disease/metabolism , Transforming Growth Factor beta1/analysis
17.
Article in English | WPRIM | ID: wpr-9479

ABSTRACT

Maternal chorioamnionitis has been associated with abnormal lung development. We examined the effect of maternal chorioamnionitis on the expression of transforming growth factor-beta1 (TGF-beta1) in the lungs of preterm infants. A total of 63 preterm (< or =34 weeks) infants who were intubated in the delivery room were prospectively enrolled. Their placentas were examined for the presence of chorioamnionitis. Bronchoalveolar lavage (BAL) fluid and cells were obtained shortly after birth. TGF-beta1 was measured in BAL fluid and TGF-beta1 mRNA expression was determined by reverse transcription polymerase chain reaction (RT-PCR) in BAL cells. TGF-beta1 mRNA expression in BAL cells showed a positive correlation with gestational age (r=0.414, p=0.002). TGF-beta1 mRNA expression was significantly decreased in the presence of maternal chorioamnionitis (0.70+/-0.12 vs. 0.81+/-0.15, p=0.007). Adjustment for gestational age, birth weight, and delivery mode did not nullify the significance. TGF-beta1 mRNA expression was marginally significantly decreased in preterm infants who developed bronchopulmonary dysplasia (BPD) later (0.75+/-0.11 vs. 0.82+/-0.15, p=0.055). However, adjustment for gestational age, patent ductus arteriosus (PDA), and maternal chorioamnionitis nullified the significance. These results might be an indirect evidence that maternal chorioamnionitis may inhibit normal lung development of fetus.


Subject(s)
Birth Weight , Bronchoalveolar Lavage Fluid/chemistry , Bronchopulmonary Dysplasia/etiology , Chorioamnionitis/metabolism , Female , Humans , Infant, Newborn , Infant, Premature , Male , Pregnancy , RNA, Messenger/analysis , Transforming Growth Factor beta1/analysis
SELECTION OF CITATIONS
SEARCH DETAIL