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1.
Medicina (B.Aires) ; 81(3): 346-358, jun. 2021. graf
Article in English | LILACS | ID: biblio-1346469

ABSTRACT

Abstract Hepatocellular carcinoma (HCC) is the most common primary liver tumor. Hexachlorobenzene (HCB) is an endocrine disruptor and a liver tumor promoter. Deregulation of thyroid hormone (TH) homeostasis may play a significant role in early neoplastic transformation. The aim of this study was to evaluate the relation between TH metabolism and the regulation of cell growth in an in vivo and in vitro model. We examined the role of transforming growth factor-β1 (TGF-β1) on TH deiodinase expression and hepatocyte proliferation. An initiation (DEN)/promotion (HCB) tumor model from rat liver and HepG2 cells were used. We evaluated PCNA, p21, p27, SMAD2/3, TGF-β1, deiodinase 1 (D1), D3, protein expression levels; D1 and D3 mRNA expression; TH and TGF-β1, D1, D3, and GST-P protein levels in focal/non-focal areas. In vivo, HCB decreased triiodothyronine (T3) and D1 mRNA levels and increased thyroxine (T4) and D3 mRNA levels in liver from DEN+HCB vs. DEN group. HCB increased protein levels from D3, TGF-β1, and PCNA and decreased D1 in focal-areas. In vitro, HCB increased PCNA, pSMAD 2/3, and TGF-β1 protein levels and mRNA expression and decreased p21 and p27 protein levels. Exogenous T3 treatment prevent HCB induced molecular alterations related to hepatocyte proliferation whereas T4 did not have any effect. These effects were prevented by using a TGF-β1 receptor II inhibitor. Results suggest that alteration of TH homeostasis, through D1 function, play a key role in hepatocyte proliferation and that TGF-β1-SMAD pathway is involved in this process confirming their role in early neoplastic transformation in HCC.


Resumen El hepatocarcinoma (HCC) es un tumor hepático primario. El hexaclorobenceno (HCB) es un disruptor endocrino y un promotor de tumores hepáticos. La desregulación de la homeostasis de las hormonas tiroideas (HT) puede ser un proceso importante para la transformación neoplásica temprana. Nuestro objetivo fue evaluar la relación entre el metabolismo de las HT y la regulación de la prolifera ción celular. Se utilizó un modelo tumoral de iniciación (DEN)/promoción (HCB) de hígado de rata (in vivo) (DEN/ HCB) y células HepG2 (in vitro). Evaluamos los niveles de PCNA, p21, p27, SMAD2/3, TGF-β1, D1, D3, ARNm de D1 y D3, HT y los niveles de TGF-β1, D1, D3 y GST-P en áreas focales/no focales. In vivo, HCB disminuyó los niveles de T3 y ARNm de la D1 y aumentó los niveles de T4 y ARNm de D3 del grupo DEN + HCB frente al grupo DEN. El HCB aumentó los niveles de D3, TGF-β1 y PCNA y disminuyó el D1 en las áreas focales. In vitro, HCB aumentó los niveles de PCNA, pSMAD 2/3 y TGF-β1 y la expresión de ARNm mientras que disminuyó los niveles de p21 y p27. El tratamiento con T3 exógeno previno las alteraciones moleculares relacionadas con la proliferación hepatocitaria. Estos efectos se evitaron utilizando un inhibidor del receptor II de TGF-β1. Los resultados sugieren que la alteración de la homeostasis de HT, a través de la D1 y la vía TGF-β1-SMAD, juega un papel clave en la proliferación celular y en las transformaciones neoplásicas tempranas en el HCC.


Subject(s)
Animals , Rats , Carcinoma, Hepatocellular , Transforming Growth Factor beta1 , Iodide Peroxidase/genetics , Liver Neoplasms , Cell Proliferation
2.
São Paulo; s.n; 2021. 52 p.
Thesis in Portuguese | LILACS, Inca | ID: biblio-1348838

ABSTRACT

Introdução: O câncer de mama é a neoplasia mais comum em mulheres. A maioria deles é diagnosticada em estágios iniciais, quando o tratamento visa a cura. Mas apesar dos avanços no tratamento, metástases à distância podem ocorrer. A biópsia das lesões metastáticas é recomendada para confirmar o status do receptor de estrogênio (RE), receptor de progesterona (RP) e receptor do fator de crescimento epidérmico humano 2 (HER2), por ocorrerem discrepâncias nesses padrões entre tumores primários e metástases em até 40% dos casos. As células tumorais circulantes (CTCs) estão relacionadas às evoluções clínicas do câncer de mama e podem potencialmente desempenhar um papel substituto aos procedimentos invasivos de rebiópsia de metástase. A tecnologia ISET® (Isolation by SizE of Tumor Cells, Rarecells-Diagnostics, Paris, França) não é usualmente empregada para detectar CTCs em pacientes com câncer de mama, embora seja reconhecida como uma ferramenta útil em alguns outros tumores. Existem dados emergentes de que a caracterização da expressão proteica das CTC pode refinar seu valor prognóstico. Sabe-se que o fator de transformação de crescimento (TGF-ß) desempenha um papel na progressão e invasividade do câncer de mama. Objetivos: Comparar a expressão de RE, RP e HER2 em tumores primários, CTCs, metástases e avaliar a expressão do receptor TGF-ß tipo 1 (TGF-ß RI) em CTCs como fator prognóstico para sobrevida global. Metodologia: Estudo realizado no A.C.Camargo Cancer Center, Brasil. As amostras de sangue foram coletadas antes da biópsia guiada por tomografia computadorizada de lesões metastáticas suspeitas e processadas pela metodologia ISET®. Os níveis de expressão proteica das CTCs foram comparados aos de tumores primários e metástases e correlacionados aos resultados clínicos. Todos os dados clínicopatológicos foram obtidos dos prontuários médicos. Resultados: Dos 39 pacientes inicialmente incluídos, 27 tiveram tanto a biópsia de metástases quanto a coleta de sangue e foram considerados para análise. As taxas de concordância para a expressão de RE, RP e HER2 entre tumores primários e metástases foram altas. Não foi observada nenhuma perda de expressão de HER2 nas metástases e os tumores triplo negativos mantiveram o mesmo padrão em todas as metástases (p <0,0001). Quando as metástases e CTCs foram classificadas como triplo negativo (TN) ou não ­ TN, as CTCs determinaram alta especificidade (93%), acurácia (84,2%) e valor preditivo negativo (88%). A sobrevida global mediana de pacientes sem expressão de TGF-ß RI em CTCs foi de 42,6 x 20,8 meses para os positivos, clinicamente relevante, porém sem significância estatística (p> 0,05). Conclusões: No câncer de mama, o papel das CTCs detectadas pelo ISET® ainda não está estabelecido. Com este estudo, sugerimos que esta metodologia possa ser útil para avaliar metástases em casos de tumores não TN, assim como a expressão de TGF-ß RI em CTCs, o que pode impactar a sobrevida. Devido à limitação da amostra, estudos futuros devem se concentrar em subtipos específicos de câncer de mama, ampliando a coorte.


Introduction: Breast cancer (BC) is the most common neoplasm in women. Most of BC are diagnosed in early stages, when treatment aims cure. Despite advances in BC treatment, distant metastases may develop. Biopsy of metastatic lesions is recommended to confirm estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2) status, due to discrepancies in these patterns between primary tumors/metastasis in up to 40% of cases. Circulating Tumor Cells (CTCs) are related to breast cancer outcomes and could potentially play a role surrogating invasive procedures of metastasis rebiopsy. ISET® (Isolation by SizE of Tumor Cells, Rarecells-Diagnostics, Paris, France) technology is not currently employed to detect CTCs in breast cancer patients, although recognized as a useful tool in some other tumors. There are emerging data that characterization of CTC protein expression can refine its prognostic value. Transforming growth factor (TGF)-ß play a role in progression/invasiveness of BC. Objectives: To compare ER, PR and HER2 expression in primary tumors, CTCs, metastases and to evaluate TGF-ß type 1 receptor (TGF- ß RI) expression in CTCs as prognostic factor for overall survival. Methods: Study conducted at the A.C.Camargo Cancer Center, Brazil. Blood samples were processed in ISET® before computed tomography­guided biopsy of suspected metastatic lesions. Protein expression levels in CTCs were compared to those in primary tumors/metastases and correlated to clinical outcomes. All clinicopathological data were obtained from medical records. Results: From the 39 patients initially included, 27 had both biopsy of metastases and blood collection and were considered for analysis. Concordance rates for ER, PR and HER2 expression between primary tumors/metastases were high. No loss of HER2 expression at any metastasis site and retention of the same pattern in all triplenegative (TN) tumors (p <0.0001) were observed. When metastases/CTCs were classified as TN/non­TN, CTCs showed high specificity (93%), accuracy (84.2%) and negative predictive value (88%). The median overall survival of patients with no TGF-ß RI expression in CTCs was 42.6 x 20.8 months for positive ones, clinically relevant but not statistically significant (p>0.05). Conclusions: In BC, the role of CTCs detected by ISET® is not yet established. Here, we could suggest that this methodology may be useful to evaluate metastasis in non-TN cases as also TGF-ß RI expression in CTCs, which may impact survival. Due to sample limitation, future studies must focus on specific subtypes of BC, expanding the cohort.


Subject(s)
Female , Breast Neoplasms , ErbB Receptors , Neoplastic Cells, Circulating , Neoplasm Metastasis , Prognosis , Receptors, Progesterone , Receptors, Estrogen , Survival Analysis , Transforming Growth Factor beta1
3.
São Paulo; s.n; 2021. 71 p. tab, ilus, graf.
Thesis in Portuguese | LILACS, Inca | ID: biblio-1353654

ABSTRACT

As glândulas salivares são estruturas formadas por um sistema de ductos e ácinos responsáveis por secretar saliva. Apesar de raros, os tumores de glândulas salivares compreendem um grupo heterogêneo de lesões, apresentando diferentes características histológicas, sendo de difícil classificação e comportamento clínico diverso. A identificação de novos marcadores moleculares tem sido alvo de pesquisas para melhor compreensão e classificação dessas neoplasias, visto que a avaliação da expressão gênica e suas vias envolvidas permite identificar genes associados à regulação que modula o desenvolvimento neoplásico. Assim, novos achados podem direcionar a aplicação de novas técnicas para o diagnóstico, prognóstico e tratamento terapêutico. Contudo, pouco se sabe sobre a via de sinalização TGFß em neoplasias mais comuns em glândulas salivares, como: Adenoma Pleomórfico (AP), Carcinoma Mucoepidermoide (CME) e Carcinoma Adenoide Cístico (CAC). Diante disso, torna-se necessário ampliar a pesquisa de genes associados para a determinação de um painel de marcadores e, deste modo, fornecer informações que possam contribuir com o diagnóstico dessas neoplasias. O objetivo desse trabalho foi avaliar a expressão gênica relacionada à via de sinalização TGFß por meio da técnica de RT-PCR em tempo real (qPCR) destacando os marcadores TGFß1, ITGB6, SMAD2, SMAD4, FBN1, LTBP1 e c-MYC. Para tanto, foram selecionadas 13 amostras de AP, 17 de CME e 13 de CAC, além de 10 amostras de glândulas salivares não neoplásicas provenientes de cirurgias realizadas no A.C.Camargo Cancer Center no período do ano 2000 a 2015 e fornecidas pelo Biobanco de Tumores. Os resultados indicam que em pacientes com AP há aumento da expressão dos genes TGFß1, LTPB1, c-MYC e FBN1, enquanto a expressão de SMAD2 diminui quando comparados às amostras não neoplásicas. Em pacientes com CME, foi observada expressão aumentada dos genes TGFß1, ITGB6, FBN1 e c-MYC enquanto a expressão dos genes SMAD2 e SMAD4 diminui ao serem comparados às amostras não neoplásicas. Nos pacientes com CAC, foi observada expressão aumentada em quase todos os genes avaliados. Na análise de clusterização hierárquica não foi possível classificar nas diferentes neoplasias de glândula salivar. Para a validação dos resultados de expressão gênica foi realizada uma meta-análise utilizando dados da literatura, sendo possível observar concordância nos valores de expressão dos genes ITGB6, LTBP1 e TGFß1 em amostras de CME e dos genes FBN1, ITGB6, LTBP1, c-MYC, SMAD2 e SMAD4 nas amostras de CAC. Comparando-se a expressão dos genes entre os três tipos de neoplasias estudados, foi observado aumento de expressão dos genes c-MYC, SMAD2 e SMAD4 nos casos de CAC e aumento da expressão do gene ITGB6 nos casos de CME. A análise de sobrevida demonstrou que, em pacientes com Carcinoma Mucoepidermoide foi observado que a ausência de linfonodo comprometido e ausência de recidiva estão associadas a melhor probabilidade de sobrevida global em 5 anos. Nossos resultados sugerem que a expressão diminuída dos genes SMAD2 e SMAD4 parece não interferir na regulação transcricional de c-MYC, especialmente no AP e CME. Considerando os genes ITGB6, TGFß1, LTBP1, FBN1 e c-MYC a expressão aumentada parece ser relevante para a regulação da via de sinalização no processo de tumorigênese. Sendo assim, este estudo contribui para um melhor entendimento da via de sinalização TGFß em neoplasias de glândulas salivares, além de fornecer informações para o desenvolvimento de potenciais marcadores biológicos para essas neoplasias.


Salivary glands are structures formed by a system of ducts and acini responsible for secreting saliva. Although rare, salivary gland tumors comprise a heterogeneous group of lesions, presenting different histological features, difficult classification, and diverse clinical behavior. Identification of new molecular markers has been the subject of researchers for better comprehension and classification of these tumors, since gene expression evaluation and their signaling pathways allow the identification of genes associated with regulation that modulated tumor development. Therefore, new findings can direct the application of new technologies for diagnosis, prognosis, and therapeutic treatment. However, little is known about the TGFß signaling pathway in the most common salivary gland tumors, such as: Pleomorphic adenoma (PA), mucoepidermoid carcinoma (MEC), and adenoid cystic carcinoma (ACC). In addition, it is necessary to expand research of genes and associated genes for determining a panel of markers and, thus, provide information that could be contribute with the diagnostic of these neoplasms. The aim of this study is to evaluate the expression of genes associated with the TGFß signaling pathway by real-time RT-PCR (qPCR) highlighting the markers TGFß1, ITGB6, SMAD2, SMAD4, FBN1, LTBP1, and c-MYC. For this purpose, 13 PA samples, 17 MEC samples, 13 ACC samples, and histologically normal salivary glands samples were selected from surgeries performed at A.C.Camargo Cancer Center between 2000 and 2015. These samples were provided by Tumor Biobank. The results indicate that PA patients presented an increased TGFß1, LTPB1, c-MYC, and FBN1 gene expression whereas SMAD2 expression was decreased when compared to the normal samples. In MEC patients, increased expression of TGFß1, ITGB6, FBN1, and c-MYC genes was observed whereas SMAD2 and SMAD4 genes presented decreased expression. In ACC patients, increased expression in almost all genes was observed. In hierarchical clustering analysis it was not possible to classify the different salivary gland tumors. For the validation of the gene expression results it was carried out a meta-analysis using the literature date, being possible to observe an agreement in the expression values of the genes ITGB6, LTBP1 and TGFß1 in MEC samples and FBN1, ITGB6, LTBP1, c-MYC, SMAD2 and SMAD4 in ACC samples. Comparing gene expression among the three tumor types studied it was observed higher expression of c-MYC, SMAD2 and SMAD4 genes in ACC cases and higher expression of ITGB6 in MEC cases. Survival analysis demonstrated that, in MEC patients it was observed that absence of affected lymph nodes and absence of recurrence are associated with better overall survival in 5 years. Our results suggest that the decreased expression of SMAD2 and SMAD4 genes seems not to interfere with the transcriptional regulation of c-MYC, especially in PA and MEC. Considering ITGB6, TGFß1, LTBP1, FBN1 and c-MYC increased gene expression appears to be relevant for the regulation of the signaling pathway in tumorigenic process. Thus, this study contributes to a better understanding of TGFß signaling pathway in salivary gland tumors, apart from supplying information in development of potential biomarkers for these tumors.


Subject(s)
Humans , Male , Female , Salivary Gland Neoplasms , Carcinoma, Mucoepidermoid , Carcinoma, Adenoid Cystic , Adenoma, Pleomorphic , Gene Expression Profiling , Transforming Growth Factor beta1
4.
Braz. j. med. biol. res ; 54(4): e10692, 2021. tab, graf
Article in English | LILACS | ID: biblio-1153536

ABSTRACT

Fibrosis caused by the increase in extracellular matrix in cardiac fibroblasts plays an important role in the occurrence and development of atrial fibrillation (AF). The aim of this study was to investigate the role of hsa-miR-4443 in AF, human cardiac fibroblast (HCFB) proliferation, and extracellular matrix remodeling. TaqMan Stem-loop miRNA assay was used to measure hsa-miR-4443 expression in patients with persistent AF (n=123) and healthy controls (n=100). Patients with AF were confirmed to have atrial fibrosis by late gadolinium enhancement. At the cellular level, after hsa-miR-4443 mimic and inhibitor were transfected with HCFBs, proliferation, apoptosis, migration, and invasion were analyzed. Lastly, hsa-miR-4443-targeted gene and transforming growth factor (TGF)-β1/α-SMA/collagen pathway were evaluated by dual-luciferase reporter assay and western blot, respectively. In patients with AF, hsa-miR-4443 decreased significantly and collagen metabolism level increased significantly. Logistic regression analysis showed that low hsa-miR-4443 level was a risk factor of AF (P<0.001). The receiver operating characteristic curve revealed that hsa-miR-4443 was useful for predicting AF (area under the curve: 0.828, sensitivity: 0.71, specificity: 0.78, P<0.001). In HCFBs, hsa-miR-4443 targeted thrombospondin-1 (THBS1) and downregulated TGF-β1/α-SMA/collagen pathway. The inhibition of hsa-miR-4443 expression promoted HCFB proliferation, migration, invasion, myofibroblast differentiation, and collagen production. The significant reduction of hsa-miR-4443 can be used as a biomarker for AF. hsa-miR-4443 protected AF by targeting THBS1 and regulated TGF-β1/α-SMA/collagen pathway to inhibit HCFB proliferation and collagen synthesis.


Subject(s)
Humans , Atrial Fibrillation , MicroRNAs/genetics , Fibrosis , Collagen , Contrast Media , Thrombospondin 1/genetics , Cell Proliferation , Transforming Growth Factor beta1 , Fibroblasts , Gadolinium
5.
Article in English | WPRIM | ID: wpr-880869

ABSTRACT

Gap junction (GJ) has been indicated to have an intimate correlation with adhesion junction. However, the direct interaction between them partially remains elusive. In the current study, we aimed to elucidate the role of N-cadherin, one of the core components in adhesion junction, in mediating connexin 43, one of the functional constituents in gap junction, via transforming growth factor-β1(TGF-β1) induction in osteoblasts. We first elucidated the expressions of N-cadherin induced by TGF-β1 and also confirmed the upregulation of Cx43, and the enhancement of functional gap junctional intercellular communication (GJIC) triggered by TGF-β1 in both primary osteoblasts and MC3T3 cell line. Colocalization analysis and Co-IP experimentation showed that N-cadherin interacts with Cx43 at the site of cell-cell contact. Knockdown of N-cadherin by siRNA interference decreased the Cx43 expression and abolished the promoting effect of TGF-β1 on Cx43. Functional GJICs in living primary osteoblasts and MC3T3 cell line were also reduced. TGF-β1-induced increase in N-cadherin and Cx43 was via Smad3 activation, whereas knockdown of Smad3 signaling by using siRNA decreased the expressions of both N-cadherin and Cx43. Overall, these data indicate the direct interactions between N-cadherin and Cx43, and reveal the intervention of adhesion junction in functional gap junction in living osteoblasts.


Subject(s)
Cadherins , Cell Communication , Connexin 43 , Osteoblasts , Transforming Growth Factor beta1
6.
Article in Chinese | WPRIM | ID: wpr-878923

ABSTRACT

This study aimed to assess whether chrysin(ChR) can inhibit epithelial-mesenchymal transition(EMT) of type Ⅱ alveolar epithelial cell and produce anti-pulmonary fibrosis effect by regulating the NF-κB/Twist 1 signaling pathway. Sixty rats were randomly divided into the control group, the bleomycin(BLC) group, BLC+ChR(50 mg·kg~(-1)) group and BLC+ChR(100 mg·kg~(-1)) group, with 15 rats in each group. The pulmonary fibrosis model was induced by intratracheal injection of BLC(7 500 U·kg~(-1)). Rats were orally administered with different doses of ChR after BLC injection for 28 days. The cells were divided into control group, TGF-β1 group(5 ng·mL~(-1)), and TGF-β1+ChR(1, 10, 100 μmol·L~(-1)) groups. The type Ⅱ alveolar epithelial cells were treated with TGF-β1 for 24 h, and then treated with TGF-β1 for 48 h in the presence or absence of different doses of ChR(1, 10 and 100 μmol·L~(-1)). The morphological changes and collagen deposition in lung tissues were analyzed by HE staining, Masson staining and immunohistochemistry. The mRNA and protein expression levels of collagen Ⅰ, E-cadherin, zonula occludens-1(ZO-1), vimentin, alpha smooth muscle actin(α-SMA), inhibitor of nuclear factor kappa B alpha(IκBα), nuclear factor-kappa B p65(NF-κB p65), phospho-NF-κB p65(p-p65) and Twist 1 in lung tissues and cells were detected by qPCR and Western blot, respectively. The animal experiment results showed that as compared with the BLC group, after administration of ChR for 28 days, bleomycin-induced pulmonary fibrosis in rats was significantly relieved, collagen Ⅰ expression in lung tissues was significantly reduced(P<0.05 or P<0.01), and EMT of alveolar epithelial cells was obviously inhibited [the expression levels of E-cadherin and ZO-1 were increased and the expression levels of vimentin and α-SMA were decreased(P<0.05 or P<0.01)], concomitantly with significantly reduced IκBα and p65 phosphorylation level in cytoplasm and decreased NF-κB p65 and Twist 1 expression in nucleus(P<0.05 or P<0.01). The cell experiment results showed that different doses of ChR(1, 10 and 100 μmol·L~(-1)) significantly reduced TGF-β1-induced collagen Ⅰ expression(P<0.05 or P<0.01), significantly inhibited EMT of type Ⅱ alveolar epithelial cells[the expression levels of E-cadherin and ZO-1 were increased and the expression levels of vimentin and α-SMA were decreased(P<0.05 or P<0.01)], and inhibited IκBα and p65 phosphorylation in cytoplasm and down-regulated NF-κB p65 and Twist 1 expression in nucleus induced by TGF-β1(P<0.05 or P<0.01). The results suggest that ChR can reverse EMT of type Ⅱ alveolar epithelial cell and alleviate pulmonary fibrosis in rats, and its mechanism may be associated with reducing IκBα phosphorylation and inhibiting NF-κB p65 phosphorylation and nuclear transfer, thus down-regulating Twist 1 expression.


Subject(s)
Alveolar Epithelial Cells/metabolism , Animals , Epithelial-Mesenchymal Transition , Flavonoids , NF-kappa B/metabolism , Rats , Signal Transduction , Transforming Growth Factor beta1/genetics
7.
Article in Chinese | WPRIM | ID: wpr-878899

ABSTRACT

To investigate the effects of Dahuang Zhechong Pills combined with hepatic arterial chemoembolization(TACE) on tumor index and immune function of patients with primary liver cancer(blood stasis and collaterals blocking type), observe its application values in treatment of such patients, and provide effective treatment means for this disease. From June 2019 to December 2019, 79 patients with confirmed primary liver cancer(blood stasis and collaterals blocking type) treated in Wenzhou Hospital of Traditional Chinese Medicine were included in this study, all of which were grouped with random number table method before inclusion in this study. 40 patients in the control group were treated with TACE, while 39 patients in the observation group were treated with Dahuang Zhechong Pills combined with TACE. The efficacy was compared between two groups after 4 weeks of treatment. The immune function indexes of serum CD4~+ cells, CD4~+/CD8~+, CD3~+ cells of the observation group were higher than those in control group after treatment(P<0.05), and tumor indexes such as serum alpha-fetoprotein(AFP), carbohydrate antigen 199(CA199) and glutamic-pyruvic transaminase(ALT), total bilirubin(TBiL) levels were lower than those in the control group, with statistically significant differences(P<0.05). Plasma vascular endothelial growth factor(VEGF), transforming growth factor-β1(TGF-β1), and matrix metalloprotei-nase-2(MMP-2) levels in the observation group were lower than those in the control group after treatment, with statistically significant differences(P<0.05). The total effective rate of the observation group was 87.18%, higher than 67.50% in the control group, and the benefit rate was 94.87% in the observation group, higher than 85.00% in the control group(P<0.05). The total incidence of adverse reactions such as bone marrow suppression, gastrointestinal reaction, fever, renal function injury and peripheral nerve injury in the observation group was 48.72%, lower than 82.50% in the control group, with statistically significant difference(P<0.05). In summary, the combination of Dahuang Zhechong Pills with TACE could improve immunity, protect liver function, and reduce the risk of metastasis and the incidence of adverse reactions from chemotherapy, so it is worth popularizing for patients with primary liver cancer(blood stasis and collaterals blocking type).


Subject(s)
Carcinoma, Hepatocellular , Chemoembolization, Therapeutic , Drugs, Chinese Herbal , Humans , Liver Neoplasms/drug therapy , Matrix Metalloproteinase 2 , Transforming Growth Factor beta1 , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth Factors
8.
Braz. j. med. biol. res ; 53(8): e9794, 2020. tab, graf
Article in English | LILACS, ColecionaSUS | ID: biblio-1132540

ABSTRACT

Although estrogen has crucial functions for endometrium growth, the specific dose and underlying molecular mechanism in intrauterine adhesion (IUA) remain unclear. In this study, we aimed to investigate the effects of estrogen on epithelial-mesenchymal transition (EMT) in normal and fibrotic endometrium, and the role of estrogen and Wnt/β-catenin signaling in the formation of endometrial fibrosis. CCK-8 and immunofluorescence assay were performed to access the proliferation of different concentrations of estrogen on normal human endometrial epithelial cells (hEECs). qRT-PCR and western blot assay were utilized to explore the effect of estrogen on EMT in normal and fibrotic endometrium, and main components of Wnt/β-catenin signaling pathway in vitro. Hematoxylin and eosin and Masson staining were used to evaluate the effect of estrogen on endometrial morphology and fibrosis in vivo. Our results indicated that the proliferation of normal hEECs was inhibited by estrogen at a concentration of 30 nM accompanied by upregulation of mesenchymal markers and downregulation of epithelial markers. Interestingly, in the model of transforming growth factor β1 (TGF-β1)-induced endometrial fibrosis, the same concentration of estrogen inhibited the process of EMT, which might be partially mediated by regulation of the Wnt/β-catenin pathway. In addition, relatively high doses of estrogen efficiently increased the number of endometrial glands and reduced the area of fibrosis as determined by the reduction of EMT in IUA animal models. Taken together, our results demonstrated that an appropriate concentration of estrogen may prevent the occurrence and development of IUA by inhibiting the TGF-β1-induced EMT and activating the Wnt/β-catenin pathway.


Subject(s)
Humans , Animals , Female , Uterine Diseases , Transforming Growth Factor beta1 , Epithelial-Mesenchymal Transition , Estrogens , Wnt Signaling Pathway
9.
Braz. oral res. (Online) ; 34: e013, 2020. graf
Article in English | LILACS | ID: biblio-1089379

ABSTRACT

Abstract This study evaluated the effect of a cyclopentenone-type PG, 15-Deoxy-Δ12,14-PG J2 (15d-PGJ2), and lectin (ScLL) on the viability of human gingival fibroblasts (HGFs), and on IL-6 and TGFβ-1 release by these fibroblasts, stimulated with lipopolysaccharide (LPS). HGFs were stimulated with LPS 10 μg/ml and treated with 15d-PGJ2 1 and 2 μg/ml, and ScLL 2 and 5 μg/ml, for 1 and 3h, and then evaluated for viability by MTT assay. Supernatant was collected to detect IL-6 and TGFβ-1 release, by ELISA. Positive control was cells kept in Dulbecco's Modified Eagle's Medium, and negative control was those kept in LPS. Data were analyzed by ANOVA and Dunnett's test (α = 0.05). No significant difference was found in viability among experimental groups at 1h (p > 0.05). Percentage of ScLL 5 µg/ml viable cells was similar to that of positive control at evaluated periods (p > 0.05), whereas the other groups had lower levels than the positive control (p < 0.05). IL-6 release was statistically higher for ScLL 5 μg/ml and 15d-PGJ2 2 µg/ml at 1h, compared with the other treated groups and positive control (p < 0.05). No significant differences were found among the groups at 3h (p > 0.05), except for ScLL 2 µg/ml and 15d-PGJ2 1 µg/ml, which showed lower IL-6 release compared with that of negative control (p < 0.05). No significant difference was found among the groups for TGFβ-1 release (p > 0.05). Results indicated that ScLL 5 μg/ml did not interfere in viability, and ScLL 2 µg/ml and 15d-PGJ2 1 µg/ml demonstrated reduced IL-6 release. Tested substances had no effect on TGFβ-1 release.


Subject(s)
Humans , Prostaglandin D2/analogs & derivatives , Lipopolysaccharides/pharmacology , Interleukin-6/metabolism , Plant Lectins/pharmacology , Transforming Growth Factor beta1/metabolism , Fibroblasts/drug effects , Fibroblasts/metabolism , Reference Values , Time Factors , Enzyme-Linked Immunosorbent Assay , Prostaglandin D2/pharmacology , Cell Survival/drug effects , Cells, Cultured , Analysis of Variance , Statistics, Nonparametric , Transforming Growth Factor beta1/drug effects , Gingiva/cytology
10.
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
11.
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
12.
Clinics ; 75: e1769, 2020. tab, graf
Article in English | LILACS | ID: biblio-1142762

ABSTRACT

OBJECTIVES: To determine the effects of three sessions of a passive stretching exercise protocol on the muscles of elderly female rats. METHODS: The effects of the stretching exercises on the soleus muscle were analyzed using immunohistochemistry [tissue inhibitors of matrix metalloproteinases (TIMP), the tumor necrosis factor-alpha (TNF-α), and the gene expression levels using real-time PCR of the transforming growth factor-beta 1 (TGF-β1), collagen type 1 (COL1), and collagen type 3 (COL3)]. Fifteen 26-month-old female Wistar rats were randomly divided into two groups, namely, Stretching (SG, n=8) and Control (CG, n=7). The passive mechanical stretching protocol consisted of a set of 4 1-minute repetitions, with 30 seconds between each repetition (total treatment of 4 minutes), three times a week for 1 week. RESULTS: Immunohistochemical analysis revealed an increase of 71.4% in the TNF-α (p=0.04) gene expression levels for the SG and a 58% decrease in the TGF-β1 gene expression levels (p=0.005) in the SG compared to that in the CG. No significant differences were observed between the groups for the immunostaining of TIMP-1 or the gene expression levels of COL1 and COL3. CONCLUSION: Three sessions of static stretching reduced the gene expression level of TGF-β1, which, owing to its anti-fibrotic role, might contribute to the remodeling of the intramuscular connective tissue of the aging muscle. In addition, immunostaining revealed that TNF-α levels increased in the aging muscle tissue in response to stretching, indicating its effect on stimulating extracellular matrix degradation. These outcomes have important clinical implications in reinforcing the use of stretching exercises in the elderly, considering that the aging muscle presents an infiltration of connective tissue.


Subject(s)
Humans , Animals , Female , Aged , Rats , Muscle Stretching Exercises , Rats, Wistar , Muscle, Skeletal , Collagen Type I/genetics , Collagen Type III/genetics , Transforming Growth Factor beta1
13.
Article in Chinese | WPRIM | ID: wpr-880757

ABSTRACT

OBJECTIVE@#To investigate high-salt exposure-induced polarization of mononuclear macrophages and the changes in proliferation and phenotypic transformation of renal fibroblasts in a co-culture system.@*METHODS@#Cultured mononuclear macrophages were exposed to high salt (161 mmol/L Na +) for 2 h and the surface markers of M0, M1 and M2-type macrophages were detected with RT-qPCR. The culture medium of the macrophages in normal and high-salt groups was collected for detection of the mRNA and protein levels of IL-6 and TGF-β1 using RT-qPCR and ELISA. A co-culture system of high salt-exposed macrophages and renal fibroblasts (NRK-49F) was established using a Transwell chamber, and the changes in proliferation and migration of NRK-49F cells were examined using EdU assay and Transwell assay, respectively. Western blotting was performed to detect the expressions of collagen I, collagen III and collagen α-SMA in NRK-49F cells.@*RESULTS@#The high salt-exposed macrophages showed significantly increased mRNA levels of M2-type macrophage surface markers mannose receptor and arginase (@*CONCLUSIONS@#High-salt exposure induces polarization of mononuclear macrophages into M2-type macrophages and promotes secretion of IL-6 and TGF-β1 by the macrophages to induce the proliferation and phenotypic transformation of NRK-49F cells.


Subject(s)
Cell Proliferation , Coculture Techniques , Fibroblasts , Kidney , Macrophages , Transforming Growth Factor beta1/genetics
14.
Article in Chinese | WPRIM | ID: wpr-878836

ABSTRACT

To investigate the effect of baicalin extracted from Qinbai Qingfei Concentrated Pills on the expressions of TGF-β1, mmp2 and timp2 in mice with pulmonary fibrosis induced by bleomycin. The Biacore technique was used to detect the specific binding between Qinbai Qingfei Concentrated Pills and TGF-β1, and the affinity components were enriched, regenerated and recovered by Biacore fishing. Then ultra-performance liquid chromatography and quadrupole time of flight mass spectrometry(UPLC-Q-TOF-MS) were used to determine whether the monomer was baicalin. Biacore was used to verify the affinity kinetics of baicalin, which was validated by pharmacodynamics in vivo. Totally 30 BALB/C mice were randomly divided into three groups: baicalin group, blank group and model group. The blank group was given sodium chloride injection(0.08 mL·kg~(-1)), while the model group and the baicalin group were injected with 4 mg·kg~(-1) bleomycin. The localization of TGF-β1, mmp2 and timp2 protein in the cells and the mRNA expressions of TGF-β1, mmp2 and timp2 were detected by RT-PCR 14 days later. The results of Biacore affinity analysis showed that the peak of binding response between Qinbai Qingfei Concentrated Pills and TGF-β1 protein reached 1 524.0 RU, with specific binding. The affinity constant K_D of baicalin and TGF-β1 was 1.620 06 μmol·L~(-1), which was determined by SPR kinetic analysis, suggesting a stable binding between baicalin and TGF-β1, which verified the results of angulation. The results of immunohistochemistry showed that the deposition of cellulose in baicalin group was significantly less than that in model group, the mRNA expressions of TGF-β1, mmp2 and timp2 were decreased in baicalin solution compared with the model group. Baicalin combined with TGF-β1 could inhibit the expressions of mmp2 and timp2 and delay the progress of pulmonary fibrosis.


Subject(s)
Animals , Flavonoids , Kinetics , Mice , Mice, Inbred BALB C , Pulmonary Fibrosis , Transforming Growth Factor beta1
15.
Article in Chinese | WPRIM | ID: wpr-826728

ABSTRACT

OBJECTIVE@#To observe the therapeutic effects of different waves of electroacupuncture (EA) on knee osteoarthritis (KOA), and to explore the mechanism of different waves of EA on promoting cartilage repair.@*METHODS@#Ninety- seven patients with KOA were randomly divided into a dilatational wave group (32 cases, 2 cases dropped off), a continuous wave group (32 cases, 2 cases dropped off) and a discontinuous wave group (33 cases, 3 cases dropped off). The same acupoints of Xuehai (SP 10), Liangqiu (ST 34), Dubi (ST 35) and Neixiyan (EX-LE 4) were selected in the three groups. The dilatational wave (frequency of 2 Hz/10 Hz) was used in the dilatational wave group, the continuous wave (frequency of 10 Hz) was used in the continuous wave group, and the discontinuous wave (frequency of 10 Hz) was used in the discontinuous wave group. All the needles were retained for 30 min. All the treatment was given 3 times a week (on Monday, Wednesday and Friday) for 4 weeks. Lysholm knees scoring scale (LKSS) was used to evaluate the knee joint function before and after treatment, and the content of transforming growth factor-β1 (TGF-β1) in the joint effusion before and after treatment was determined by enzyme-linked immunosorbent assay (ELISA).@*RESULTS@#Compared before treatment, the total score and each score of LKSS were increased after treatment in the three groups (all <0.05). The improvements of total score, pain score, instability score, swelling score of LKSS in the continuous wave group and the dilatational wave group were superior to those in the discontinuous wave group (all <0.05). The content of TGF-β1 in the joint effusion in each group was increased after treatment (<0.05), and the improvement in dilatational wave group was superior to thoes in the continuous wave group and the discontinuous wave group (all <0.05).@*CONCLUSION@#The three different waves of EA could all improve the clinical symptoms of KOA, which may promote cartilage repair by increasing TGF-β1 content. The dilatational wave had the best overall effect, which can be used as a clinical optimal treatment.


Subject(s)
Acupuncture Points , Electroacupuncture , Humans , Knee Joint , Osteoarthritis, Knee , Therapeutics , Synovial Fluid , Chemistry , Transforming Growth Factor beta1 , Treatment Outcome
16.
Article in Chinese | WPRIM | ID: wpr-828925

ABSTRACT

OBJECTIVE@#To study the anti- fibrotic effect of human umbilical cord mesenchymal stem cell-derived exosomes (hUCMSC-EXOs) and explore the mechanism.@*METHODS@#Twenty-four C57 BL/6 mice were divided into 4 groups (=6), including the control group treated with intratracheal injection of saline (3 mg/kg); lung fibrosis model group with intratracheal injection of 1.5 mg/mL bleomycin solution (prepared with saline, 3 mg/kg); EXOs1 group with intratracheal injection of 1.5 mg/mL bleomycin solution (3 mg/kg) and hUCMSC-EXOs (100 μg/250 μL, given by tail vein injection on the next day after modeling); and EXOs2 group with intratracheal injection of 1.5 mg/mL bleomycin solution (3 mg/kg) and hUCMSC-EXOs (100 μg/250 μL, given by tail vein injection on the 10th day after modeling). At 21 days after modeling, pulmonary index, lung tissue pathology and collagen deposition in the mice were assessed using HE staining and Masson staining. The expression level of TGF-β1 was detected using ELISA, and vimentin, E-cadherin and phosphorylated Smad2/3 (p-Smad2/3) were detected using immunohistochemical staining. CCK8 assay was used to evaluate the effect of hUCMSCEXOs on the viability of A549 cells, and Western blotting was used to detect the expression levels of p-Smad2/3, vimentin, and E-cadherin in the cells.@*RESULTS@#Compared with those in the model group, the mice treated with hUCMSC-EXOs showed significantly reduced the pulmonary index ( < 0.05), collagen deposition, lung tissue pathologies, lowered expressions of TGF-β1 ( < 0.05), vimentin, and p-Smad2/3 and increased expression of E-cadherin. hUCMSC-EXOs given on the second day produced more pronounced effect than that given on the 11th day ( < 0.05). CCK8 assay results showed that hUCMSC-EXOs had no toxic effects on A549 cells ( > 0.05). Western blotting results showed that hUCMSC-EXOs treatment significantly increased the expression of E-cadherin and decreased the expressions of p-Smad2/3 and vimentin in the cells.@*CONCLUSIONS@#hUCMSC-EXOs can alleviate pulmonary fibrosis in mice by inhibiting epithelialmesenchymal transition activated by the TGF-β1/Smad2/3 signaling pathway, and the inhibitory effect is more obvious when it is administered on the second day after modeling.


Subject(s)
Animals , Epithelial-Mesenchymal Transition , Exosomes , Humans , Mesenchymal Stem Cells , Mice , Pulmonary Fibrosis , Transforming Growth Factor beta1 , Umbilical Cord
17.
Article in Chinese | WPRIM | ID: wpr-828876

ABSTRACT

OBJECTIVE@#To clarify the molecular signaling mechanism underlying the inhibitory effect of metformin on transforming growth factor-β1 (TGF-β1)-stimulated collagen I production in rat biliary fibroblasts.@*METHODS@#Primary biliary fibroblasts were isolated under aseptic condition from 50 Sprague-Dawley rats (half male and half female), and microscopic observation identified no obvious difference in the morphology or viability of the cells from rats with different sexes or body weight. The cells were treated with TGF-β1 (10 ng/mL), Smad3 siRNA+TGF-β1, CTGF siRNA+TGF-β1, metformin (10 mmol/L)+ TGF-β1, or Compound C (10 μmol/L)+metformin+TGF-β1. The expressions of CTGF and collagen I in the treated cells were determined using ELISA kit or Western blotting; the phorsphorylated and total Smad3 and AMPK expressions were detected using immunoblotting.@*RESULTS@#TGF-β1 time- and dose-dependently induced collagen I production in rat biliary fibroblasts. The activated AMPK by metformin dose-dependently inhibited TGF-β1-induced collagen I production. Pre-incubation of cells with the AMPK inhibitor Compound C restored the inhibitory effect of AMPK on TGF-β1-induced collagen I secretion ( < 0.01). Activation of AMPK by metformin significantly reduced TGF-β1-induced collagen I production by suppressing Smad3-driven CTGF expression ( < 0.01), and the application of Compound C reversed such changes in the fibroblasts ( < 0.01).@*CONCLUSIONS@#Metformin inhibits TGF-β1-stimulated collagen I production by activating AMPK and inhibiting Smad3- driven CTGF expression in rat biliary fibroblasts.


Subject(s)
Animals , Cells, Cultured , Collagen , Female , Fibroblasts , Male , Metformin , Rats , Rats, Sprague-Dawley , Signal Transduction , Smad3 Protein , Transforming Growth Factor beta1
18.
Article in Chinese | WPRIM | ID: wpr-828869

ABSTRACT

OBJECTIVE@#To observe the effect of traditional Chinese medicine for intervention of phlegm and blood stasis in regulating TGF-β1/Smad3 signaling and relieving nephropathy in diabetic rats.@*METHODS@#SD rats were divided into blank group (NC), diabetic model group (MC group), intervention of phlegm and blood stasis (RPDBS) group, phlegm-removing (RP) group and blood-removing (DBS) group. Diabetic models were established in all the rats except for those in the blank group. After 4 weeks of feeding, the rats in RPDBS group, RP group and DBS group were given corresponding drug intervention for 8 weeks. HE staining was used to observe the changes in renal histopathology. Western blotting and real-time fluorescence quantitative PCR were used to detect the expression levels of transforming growth factor-β1 (TGF-β1) and Smad3.@*RESULTS@#The structure and arrangement of the glomeruli and renal tubules improved significantly in the treatment groups in comparison with those in the MC group. The expression levels of TGF-β1, Smad3 and p-Smad3 were significantly downregulated at both the protein and mRNA levels in the treatment groups ( < 0.05), and the down-regulation was more obvious in RPDBS group than in RP group and DBS group ( < 0.05).@*CONCLUSIONS@#Intervention of phlegm and blood stasis may inhibit the activation of TGF-β1/Smad3 signaling pathway and delay diabetic nephropathy and fibrosis to protect the renal function in diabetic rats.


Subject(s)
Animals , Diabetes Mellitus, Experimental , Diabetic Nephropathies , Kidney , Rats , Rats, Sprague-Dawley , Signal Transduction , Smad3 Protein , Transforming Growth Factor beta1
19.
Article in Chinese | WPRIM | ID: wpr-828506

ABSTRACT

OBJECTIVE@#To study the anti- fibrotic effect of human umbilical cord mesenchymal stem cell-derived exosomes (hUCMSC-EXOs) and explore the mechanism.@*METHODS@#Twenty-four C57 BL/6 mice were divided into 4 groups (=6), including the control group treated with intratracheal injection of saline (3 mg/kg); lung fibrosis model group with intratracheal injection of 1.5 mg/mL bleomycin solution (prepared with saline, 3 mg/kg); EXOs1 group with intratracheal injection of 1.5 mg/mL bleomycin solution (3 mg/kg) and hUCMSC-EXOs (100 μg/250 μL, given by tail vein injection on the next day after modeling); and EXOs2 group with intratracheal injection of 1.5 mg/mL bleomycin solution (3 mg/kg) and hUCMSC-EXOs (100 μg/250 μL, given by tail vein injection on the 10th day after modeling). At 21 days after modeling, pulmonary index, lung tissue pathology and collagen deposition in the mice were assessed using HE staining and Masson staining. The expression level of TGF-β1 was detected using ELISA, and vimentin, E-cadherin and phosphorylated Smad2/3 (p-Smad2/3) were detected using immunohistochemical staining. CCK8 assay was used to evaluate the effect of hUCMSCEXOs on the viability of A549 cells, and Western blotting was used to detect the expression levels of p-Smad2/3, vimentin, and E-cadherin in the cells.@*RESULTS@#Compared with those in the model group, the mice treated with hUCMSC-EXOs showed significantly reduced the pulmonary index ( < 0.05), collagen deposition, lung tissue pathologies, lowered expressions of TGF-β1 ( < 0.05), vimentin, and p-Smad2/3 and increased expression of E-cadherin. hUCMSC-EXOs given on the second day produced more pronounced effect than that given on the 11th day ( < 0.05). CCK8 assay results showed that hUCMSC-EXOs had no toxic effects on A549 cells ( > 0.05). Western blotting results showed that hUCMSC-EXOs treatment significantly increased the expression of E-cadherin and decreased the expressions of p-Smad2/3 and vimentin in the cells.@*CONCLUSIONS@#hUCMSC-EXOs can alleviate pulmonary fibrosis in mice by inhibiting epithelialmesenchymal transition activated by the TGF-β1/Smad2/3 signaling pathway, and the inhibitory effect is more obvious when it is administered on the second day after modeling.


Subject(s)
Animals , Epithelial-Mesenchymal Transition , Exosomes , Gene Expression Profiling , Gene Expression Regulation , Humans , Mesenchymal Stem Cells , Cell Biology , Mice , Pulmonary Fibrosis , Therapeutics , Transforming Growth Factor beta1 , Genetics , Umbilical Cord , Cell Biology
20.
Article in Chinese | WPRIM | ID: wpr-828366

ABSTRACT

The aim of this paper was to observe the effect of salvianolic acid B(Sal B) on high-glucose induced renal tubular epithelial-mesenchymal transition(EMT) in rats, and to explore its possible mechanisms of prevention and treatment of diabetic nephropathy. The rat renal tubular epithelial NRK-52 E cells were cultured in vitro. The cells were divided into control group, high glucose group, high glucose+10 μmol·L~(-1)Sal B group(Sal B), the above 3 groups were set at 6, 12, 24 and 48 h for dynamic observation; high glucose+Sal B different concentration(1, 5, 10 μmol·L~(-1)) groups, high glucose+5.0 μmol·L~(-1) pioglitazone group, high glucose+10 μmol·L~(-1)Sal B+5 μmol·L~(-1)GW9662 group. The protein expression levels of PPARγ, PTEN, α-SMA, E-cadherin and PI3 K/Akt signaling molecules were determined by Western blot. The mRNA expression of PPARγ and PTEN were detected by Real-time PCR. The viabi-lity of NRK52 E cells was determined by MTT assay. The results showed that as compared with control group, the mRNA and protein expression levels of PPARγ and PTEN in high glucose group gradually reduced, the protein expression levels of α-SMA and p-Akt~((Thr308))gradually increased, and the protein expression of E-cadherin gradually reduced(P<0.05). As compared with high glucose group, when increases in Sal B doses, the mRNA and protein expression levels of PPARγ, PTEN in high glucose + different concentrations of Sal B groups gradually increased, the protein expression levels of α-SMA and p-Akt~((Thr308)) gradually reduced, and the protein expression of E-cadherin gradually increased(P<0.05), however, the effect of 1 μmol·L~(-1)concentration of Sal B on the expression of PPARγ mRNA and protein and PTEN mRNA was not significantly different. As compared with high glucose group, the mRNA and protein expression levels of PPARγ mRNA(except 6 h) and protein(except 6 h), PTEN mRNA(except 6 h) and protein(except 6, 12 h) kept increasing, the protein expression levels of α-SMA and p-Akt~((Thr308))(except 6 h) continued to reduce, the protein expression of E-cadherin kept increasing in high glucose+10 μmol·L~(-1) Sal B dynamic observation group(P<0.05). As compared with high glucose group, Sal B and the pioglitazone(PIO) can greatly enhance the expression of PPARγ, PTEN at mRNA and protein levels, enhance the expression of E-cadherin at protein levels, and reduce the expression of α-SMA, p-Akt~((Thr308))protein level(P<0.05), there was no significant difference between the two groups. However, the expression levels of PPARγ and PTEN mRNA and protein, E-cadherin, α-SMA and p-Akt(Thr308) protein in the Sal B+GW9662 control group were not statistically significant compared with the high glucose group. The effect of Sal B was blocked by the PPARγ antagonist GW9662. It can be concluded that Sal B can suppress the NRK52 E cells induced by high-glucose EMT. The mechanism may be related to the activation of PPARγ with Sal B, and the up-regulation of PTEN expression, and thereby inhibiting the fibrosis effect of PI3 K/Akt signaling pathway.


Subject(s)
Animals , Benzofurans , Epithelial Cells , Epithelial-Mesenchymal Transition , Glucose , Rats , Transforming Growth Factor beta1
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