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1.
Int J Mol Sci ; 22(12)2021 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-34203850

RESUMO

Steroid receptor coactivator-1 (SRC-1) is a transcription coactivator playing a pivotal role in mediating a wide range of signaling pathways by interacting with related transcription factors and nuclear receptors. Aberrantly elevated SRC-1 activity is associated with cancer metastasis and progression, and therefore, suppression of SRC-1 is emerging as a promising therapeutic strategy. In this study, we developed a novel SRC-1 degrader for targeted degradation of cellular SRC-1. This molecule consists of a selective ligand for SRC-1 and a bulky hydrophobic group. Since the hydrophobic moiety on the protein surface could mimic a partially denatured hydrophobic region of a protein, SRC-1 could be recognized as an unfolded protein and experience the chaperone-mediated degradation in the cells through the ubiquitin-proteasome system (UPS). Our results demonstrate that a hydrophobic-tagged chimeric molecule is shown to significantly reduce cellular levels of SRC-1 and suppress cancer cell migration and invasion. Together, these results highlight that our SRC-1 degrader represents a novel class of therapeutic candidates for targeting cancer metastasis. Moreover, we believe that the hydrophobic tagging strategy would be widely applicable to develop peptide-based protein degraders with enhanced cellular activity.


Assuntos
Interações Hidrofóbicas e Hidrofílicas , Coativador 1 de Receptor Nuclear/metabolismo , Proteólise , Transativadores/metabolismo , Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular , Movimento Celular , Humanos , Fator Estimulador de Colônias de Macrófagos/genética , Fator Estimulador de Colônias de Macrófagos/metabolismo , Chaperonas Moleculares/metabolismo , Invasividade Neoplásica , Peptídeos/química , Complexo de Endopeptidases do Proteassoma/metabolismo , Estabilidade Proteica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
2.
Int J Mol Sci ; 22(11)2021 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-34205118

RESUMO

During metastasis, cancer cells that originate from the primary tumor circulate in the bloodstream, extravasate, and form micrometastases at distant locations. Several lines of evidence suggest that specific interactions between cancer cells and endothelial cells, in particular tumor cell adhesion to the endothelium and transendothelial migration, play a crucial role in extravasation. Here we have studied the role of vascular endothelial (VE)-cadherin which is expressed aberrantly by breast cancer cells and might promote such interactions. By comparing different human breast cancer cell lines, we observed that the number of cancer cells that adhered to endothelium correlated with VE-cadherin expression levels. VE-cadherin silencing experiments confirmed that VE-cadherin enhances cancer cell adhesion to endothelial cells. However, in contrast, the number of cancer cells that incorporated into the endothelium was not dependent on VE-cadherin. Thus, it appears that cancer cell adhesion and incorporation are distinct processes that are governed by different molecular mechanisms. When cancer cells incorporated into the endothelial monolayer, they formed VE-cadherin positive contacts with endothelial cells. On the other hand, we also observed tumor cells that had displaced endothelial cells, reflecting either different modes of incorporation, or a temporal sequence where cancer cells first form contact with endothelial cells and then displace them to facilitate transmigration. Taken together, these results show that VE-cadherin promotes the adhesion of breast cancer cells to the endothelium and is involved in the initial phase of incorporation, but not their transmigration. Thus, VE-cadherin might be of relevance for therapeutic strategies aiming at preventing the metastatic spread of breast cancer cells.


Assuntos
Antígenos CD/genética , Neoplasias da Mama/genética , Caderinas/genética , Adesão Celular/genética , Endotélio Vascular/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Técnicas de Cocultura , Endotélio Vascular/patologia , Endotélio Vascular/ultraestrutura , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Células Endoteliais da Veia Umbilical Humana/citologia , Humanos , Imagem Molecular/métodos , Metástase Neoplásica
3.
Nan Fang Yi Ke Da Xue Xue Bao ; 41(6): 931-936, 2021 Jun 20.
Artigo em Chinês | MEDLINE | ID: mdl-34238747

RESUMO

OBJECTIVE: To investigate the regulatory role of the long non-coding RNA (lncRNA) small nucleolar host gene 3 (SNHG3) in proliferation, migration and invasion of human cervical cancer cell line SiHa. OBJECTIVE: Array data were retrieved from GEO database to analyze the expression levels of SNHG3 in cervical cancer and adjacent normal tissues. SiHa cells were transfected with a small interfering RNA (siRNA) targeting SNHG3, and the changes in the transcriptional levels of lncRNA SNHG3 and the epithelial-mesenchymal transition (EMT) markers N-cadherin, Snail, vimentin and E-cadherin were detected using real-time quantitative PCR; the protein expressions of N-cadherin, Snail, vimentin and E-cadherin were determined using Western blotting. Cell counting kit-8 (CCK8) assay was utilized to assess the proliferation capacity of the transfected cells. Wound healing assay and Transwell assay were performed to evaluate the transversal and longitudinal migration and invasion abilities of the cells. OBJECTIVE: SNHG3 was over-expressed in cervical cancer tissues and SiHa cells. In SiHa cells, knocking down SNHG3 significantly inhibited the proliferation (P < 0.001), migration (P < 0.01) and invasion abilities (P < 0.001) of the cells, down-regulated the expression levels of N-cadherin, Snail and vimentin (P < 0.001) and up-regulated the expression of E-cadherin (P < 0.001). OBJECTIVE: SNHG3 may promote the proliferation, migration and invasion of SiHa cells by activating the EMT signaling pathway.


Assuntos
RNA Longo não Codificante , Neoplasias do Colo do Útero , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , RNA Longo não Codificante/genética , RNA Interferente Pequeno , Neoplasias do Colo do Útero/genética
4.
Zhongguo Dang Dai Er Ke Za Zhi ; 23(7): 749-754, 2021 Jul.
Artigo em Chinês | MEDLINE | ID: mdl-34266536

RESUMO

Febrile seizures are the most common nervous system disease in childhood, and most children have a good prognosis. However, some epilepsy cases are easily induced by fever and are characterized by "fever sensitivity", and it is difficult to differentiate such cases from febrile seizures. Epilepsy related to fever sensitivity includes hereditary epilepsy with febrile seizures plus, Dravet syndrome, and PCDH19 gene-related epilepsy. This article mainly describes the clinical manifestations of these three types of epilepsy and summarizes their clinical features in the early stage of disease onset, so as to achieve early identification, early diagnosis, and early intervention to improve prognosis.


Assuntos
Epilepsia , Síndromes Epilépticas , Convulsões Febris , Caderinas/genética , Criança , Epilepsia/etiologia , Epilepsia/genética , Humanos , Mutação , Convulsões Febris/etiologia , Convulsões Febris/genética
5.
Medicine (Baltimore) ; 100(24): e26307, 2021 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-34128868

RESUMO

INTRODUCTION: 22q13.3 deletion syndrome is a well-known syndrome characterized by typical clinical findings including neonatal hypotonia, absent or severely delayed speech, intellectual disability, and other various features, and detection of a heterozygous deletion of chromosome 22q13.3 with the involvement of at least part of SHANK3. It is reported that 10% to 29% of patients with 22q13.3 deletion syndrome present lymphedema. Protein-losing enteropathy (PLE) has never been reported in 22q13.3 deletion syndrome. PATIENT CONCERNS: The patient presented to our institution for refractory hypoalbuminemia and chronic lymphedema in both legs. DIAGNOSIS: The patient manifested intellectual disability, absent speech, tooth grinding, dysmorphic face, and abnormal hands and toenails. Copy-number variation sequencing confirmed the maternal deletion in 22q13.31-q13.33 (chr22:46285592-51244566, hg19). The patient was genetically diagnosed with 22q13.3 deletion syndrome. INTERVENTIONS: Low-fat diets and medium-chain triglycerides supplements were prescribed. The patient was recommended to wear compression garments and elevate legs. OUTCOMES: The symptom of diarrhea was resolved, but hypoalbuminemia persisted. Lower extremities lymphedema was gradually becoming severe. CONCLUSIONS: Primary lymphedema and PLE can occur simultaneously in a patient with 22q13.3 deletion syndrome. The 2 phenotypes could share the same genetic etiology of congenital lymphatic abnormalities. CELSR1 deletion may play a role in lymphatic dysplasia. The case also provides additional proof of the pathogenic effect of CELSR1 on hereditary lymphedema.


Assuntos
Caderinas/genética , Transtornos Cromossômicos/genética , Linfedema/genética , Enteropatias Perdedoras de Proteínas/genética , Deleção Cromossômica , Cromossomos Humanos Par 22/genética , Variações do Número de Cópias de DNA , Feminino , Humanos , Hipoalbuminemia/genética , Deficiência Intelectual/genética , Perna (Membro)/patologia , Adulto Jovem
6.
J Cell Sci ; 134(11)2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-34085693

RESUMO

Urokinase-type plasminogen activator (uPA; encoded by Plau) is a serine proteinase that, in the central nervous system, induces astrocytic activation. ß-Catenin is a protein that links the cytoplasmic tail of cadherins to the actin cytoskeleton, thus securing the formation of cadherin-mediated cell adhesion complexes. Disruption of cell-cell contacts leads to the detachment of ß-catenin from cadherins, and ß-catenin is then degraded by the proteasome following its phosphorylation by GSK3ß. Here, we show that astrocytes release uPA following a scratch injury, and that this uPA promotes wound healing via a plasminogen-independent mechanism. We found that uPA induces the detachment of ß-catenin from the cytoplasmic tail of N-cadherin (NCAD; also known as CDH2) by triggering its phosphorylation at Tyr654. Surprisingly, this is not followed by degradation of ß-catenin because uPA also induces the phosphorylation of the low density lipoprotein receptor-related protein 6 (LRP6) at Ser1490, which then blocks the kinase activity of GSK3ß. Our work indicates that the ensuing cytoplasmic accumulation of ß-catenin is followed by its nuclear translocation and ß-catenin-triggered transcription of the receptor for uPA (Plaur), which in turn is required for uPA to induce astrocytic wound healing.


Assuntos
Ativador de Plasminogênio Tipo Uroquinase , beta Catenina , Caderinas/genética , Receptores de Ativador de Plasminogênio Tipo Uroquinase/genética , Ativador de Plasminogênio Tipo Uroquinase/genética , Cicatrização , beta Catenina/genética
7.
Talanta ; 232: 122442, 2021 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-34074427

RESUMO

The anti-tumor effects of metformin hydrochloride (MH), an initial pharmacologic agent for type 2 diabetes, were reexamined by surface-enhanced Raman scattering (SERS) spectroscopy. A SERS immuno-tag fabricated by decorating silver nanoparticles (AgNPs) with a specific antibody was employed to trace the dynamic expression of the tumor metastasis-related N-cadherin. With the MH action, the N-cadherin expression on the cell membranes decreases, proving that MH has a pharmacological effect on prohibiting cancer cell metastasis. Another AgNP-based nucleus targeting nanoprobe was adopted to culture with the MH acted cells, which can help the label-free SERS collection of the cell nuclei to explore the MH influences on intranuclear genes and proteins. By analyzing the intranuclear SERS spectra, the find is that MH has impacts on the transcription and translation of genes, thus regulates the expression of tumor metastasis-related proteins (N-cadherins). This study presents a proof-of-concept for MH as a potential drug for diabetes patients associated with tumors. The developed plasmonic immune analytical platform can be extended to assess other substances of the cell membrane and applicable for the SERS-based screening of membrane receptor-related drugs at the cellular level.


Assuntos
Diabetes Mellitus Tipo 2 , Nanopartículas Metálicas , Metformina , Caderinas/genética , Membrana Celular , Núcleo Celular , Humanos , Metformina/farmacologia , Prata , Análise Espectral Raman
8.
Arkh Patol ; 83(3): 11-19, 2021.
Artigo em Russo | MEDLINE | ID: mdl-34041891

RESUMO

OBJECTIVE: To determine the level of E-cadherin expression in tumor emboli, to compare it with expression in a tumor, to determine the dependence of E-cadherin expression in tumor emboli on the clinical and morphological characteristics of gastric cancer. MATERIAL AND METHODS: We used samples of surgical material from 280 patients with a verified diagnosis of gastric cancer. E-cadherin expression was determined by immunohistochemical method. The results of the reactions were assessed semi-quantitatively and compared with the main clinical and morphological characteristics of gastric cancer (histological type according to the WHO classification 2019, histological type according to the classification of P. Lauren, clinical stage, depth of invasion (T), number of metastases in lymph nodes (N), presence or/absence of distant metastases (M), tumor localization in the stomach). RESULTS: Among 280 cases of cancer, emboli were detected only in 67 cases, used for further analysis. The rest of the samples were excluded from the analysis, since emboli did not get into the sections during the cutting of immunohistochemical preparations. The expression of E-cadherin in tumor emboli was significantly higher (p<0.001) than in tumor tissue. At the same time, no cases identified where the level of E-cadherin decreased in emboli compared to the tumor. A significant increase in the expression of E-cadherin in tumor emboli compared to the primary tumor was noted for all histological types according to WHO 2019, for intermediate and diffuse types according to the P. Lauren classification (p<0.001). Comparison of expression in emboli and tumors for neoplasms with different depths of invasion (T), different stages and different localizations did not reveal statistically significant differences. An increase in the expression of E-cadherin in emboli compared to tumors was characterized by a higher level of significance in the presence of metastases (N1, N2, N3a, N3b; p<0.001) than in the absence of metastases (N0; p=0.016). CONCLUSION: The study revealed a statistically significant increase in the expression of E-cadherin in tumor emboli compared to the primary tumor, which is evidence of its important role in maintaining the integrity of emboli and tumor dissemination.


Assuntos
Neoplasias Gástricas , Caderinas/genética , Diferenciação Celular , Humanos , Neoplasias Gástricas/genética
9.
Cancer Med ; 10(10): 3346-3357, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33932127

RESUMO

BACKGROUND: Chronic alcohol consumption is more frequently associated with advanced, aggressive hepatocellular carcinoma (HCC) tumors. Alcohol adversely impacts ER/Golgi membrane trafficking and Golgi protein N-glycosylation in hepatocytes; these effects have been attributed (in part) to dysregulated adenosine diphosphate-ribosylation factor (ARF) GTPase signaling. Here, we investigated the role of the ARF GTPase guanine exchange factor PSD4 in HCC progression. METHODS: R-based bioinformatics analysis was performed on publicly available array data. Modulating gene expression was accomplished via lentiviral vectors. Gene expression was analyzed using quantitative real-time PCR and immunoblotting. PSD4 promoter methylation was assessed using quantitative methylation-specific PCR. Phospho-p65(S276)/DNMT1 binding to the PSD4 promoter was analyzed via chromatin immunoprecipitation. We constructed ethanol/DEN-induced and DEN only-induced transgenic murine models of HCC. RESULTS: We identified PSD4 as a hypermethylated, suppressed gene in alcohol-related HCC tumors; however, PSD4 was not dysregulated in all-cause HCC tumors. Certain HCC cell lines also displayed varying degrees of PSD4 downregulation. PSD4 overexpression or knockdown decreased and increased cell migration and invasiveness, respectively. Mechanistically, PSD4 transcription was repressed by TNF-α-induced phospho-p65(S276)'s recruitment of DNA methyltransferase 1 (DNMT1), resulting in PSD4 promoter methylation. PSD4 inhibited pro-EMT CDC42 activity, resulting in downregulation of E-cadherin and upregulation of N-cadherin and vimentin. Hepatocyte-specific PSD4 overexpression reduced ethanol/DEN-induced HCC tumor progression and EMT marker expression in vivo. CONCLUSIONS: PSD4 is a hypermethylated, suppressed gene in alcohol-related HCC tumors that negatively modulated pro-EMT CDC42 activity. Furthermore, we present a novel phospho-NF-κB p65(S276)/DNMT1-mediated promoter methylation mechanism by which TNF-α/NF-κB signaling represses PSD4 transcription in HCC cells.


Assuntos
Álcoois/efeitos adversos , Carcinoma Hepatocelular/genética , Epigênese Genética/genética , Fatores de Troca do Nucleotídeo Guanina/genética , Neoplasias Hepáticas/genética , NF-kappa B/genética , Transcrição Genética/genética , Fator de Necrose Tumoral alfa/genética , Consumo de Bebidas Alcoólicas/genética , Animais , Caderinas/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , DNA (Citosina-5-)-Metiltransferase 1/genética , Metilação de DNA/genética , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Células Hep G2 , Hepatócitos/patologia , Humanos , Neoplasias Hepáticas/patologia , Camundongos , Camundongos Transgênicos , Gravidez , Regiões Promotoras Genéticas/genética , Transdução de Sinais/genética , Fator de Transcrição RelA/genética
10.
J Med Case Rep ; 15(1): 263, 2021 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-33941229

RESUMO

BACKGROUND: Germline pathogenic variants in the cadherin-1 (CDH1) gene cause a predisposition to hereditary diffuse gastric cancer (HDGC). We report an HDGC case in Vietnam and identify a novel mutation in the CDH1 gene. CASE PRESENTATION: A 28-year-old Vietnamese man was diagnosed with HDGC and a novel mutation at c.639G>A. All exons of CDH1 were sequenced in his pedigree, which revealed the c.639G>A mutation in the proband, his father, and uncle. The patient refused treatment and died 4 months after diagnosis. Endoscopic surveillance of the father and the uncle showed structural abnormalities in the father. CONCLUSION: In cases of HDGC, identification of the CDH1 gene mutation is very important for better counseling and more effective strategies to prevent the development of diseases, such as prophylactic gastrectomy for family members with genetic mutations.


Assuntos
Neoplasias Gástricas , Adulto , Grupo com Ancestrais do Continente Asiático , Caderinas/genética , Códon de Terminação , Éxons/genética , Predisposição Genética para Doença , Mutação em Linhagem Germinativa , Humanos , Masculino , Mutação , Linhagem , Neoplasias Gástricas/genética
11.
Anticancer Res ; 41(5): 2383-2395, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33952463

RESUMO

BACKGROUND/AIM: This study aimed to investigate the effect of the new ciprofloxacin chalcone [7-(4-(N-substituted carbamoyl methyl) piperazin-1 yl)] on the proliferation, migration, and metastasis of MCF-7 and MDA-MB-231 breast cancer cell lines. MATERIALS AND METHODS: Cell viability, colony formation and cell migration abilities were analysed. Cell cycle distribution and apoptosis were examined by flow cytometry. The molecular mechanism underlying chalcone's activity was investigated using qRT-PCR and western blotting. RESULTS: This new ciprofloxacin chalcone significantly inhibited proliferation, colony formation, and cell migration abilities of both cancer cell lines. Furthermore, it initiated apoptosis and caused cell cycle arrest at G2/M and S phase in MCF-7 and MDA-MB-231 cell lines, respectively. In addition, it up-regulated the expression of pro-apoptotic factors, p53, PUMA and NOXA, and down-regulated the expression of anti-apoptotic factors, MDM2 and MDM4. At the same time, it inhibited epithelial-mesenchymal transition by increasing the expression of E-cadherin and decreasing the expression of TGF-ß1, SNAI1, TWIST1, MMP2, and MMP9. CONCLUSION: This new ciprofloxacin chalcone exhibited promising apoptotic and anti-metastatic activities against MCF-7 and MDA-MB-231 breast cancer cell lines, and, therefore, is an attractive molecule for drug development in the treatment of breast cancer.


Assuntos
Apoptose/efeitos dos fármacos , Neoplasias da Mama/metabolismo , Chalcona/farmacologia , Ciprofloxacina/farmacologia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Proteínas/metabolismo , Transdução de Sinais/efeitos dos fármacos , Apoptose/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Caderinas/genética , Caderinas/metabolismo , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Chalcona/química , Ciprofloxacina/química , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Células MCF-7 , Estrutura Molecular , Proteínas/genética , Transdução de Sinais/genética , Fatores de Transcrição da Família Snail/genética , Fatores de Transcrição da Família Snail/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismo
12.
EMBO Rep ; 22(6): e51299, 2021 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-33880878

RESUMO

Endothelium protection is critical, because of the impact of vascular leakage and edema on pathological conditions such as brain ischemia. Whereas deficiency of class II phosphoinositide 3-kinase alpha (PI3KC2α) results in an increase in vascular permeability, we uncover a crucial role of the beta isoform (PI3KC2ß) in the loss of endothelial barrier integrity following injury. Here, we studied the role of PI3KC2ß in endothelial permeability and endosomal trafficking in vitro and in vivo in ischemic stroke. Mice with inactive PI3KC2ß showed protection against vascular permeability, edema, cerebral infarction, and deleterious inflammatory response. Loss of PI3KC2ß in human cerebral microvascular endothelial cells stabilized homotypic cell-cell junctions by increasing Rab11-dependent VE-cadherin recycling. These results identify PI3KC2ß as a potential new therapeutic target to prevent aggravating lesions following ischemic stroke.


Assuntos
Células Endoteliais , Fosfatidilinositol 3-Quinases , Junções Aderentes/metabolismo , Animais , Antígenos CD/metabolismo , Caderinas/genética , Caderinas/metabolismo , Permeabilidade Capilar , Células Endoteliais/metabolismo , Endotélio Vascular/metabolismo , Camundongos , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo
13.
Nucleic Acids Res ; 49(8): 4506-4521, 2021 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-33849071

RESUMO

Repressor element-1 silencing transcription factor (REST) or neuron-restrictive silencer factor (NRSF) is a zinc-finger (ZF) containing transcriptional repressor that recognizes thousands of neuron-restrictive silencer elements (NRSEs) in mammalian genomes. How REST/NRSF regulates gene expression remains incompletely understood. Here, we investigate the binding pattern and regulation mechanism of REST/NRSF in the clustered protocadherin (PCDH) genes. We find that REST/NRSF directionally forms base-specific interactions with NRSEs via tandem ZFs in an anti-parallel manner but with striking conformational changes. In addition, REST/NRSF recruitment to the HS5-1 enhancer leads to the decrease of long-range enhancer-promoter interactions and downregulation of the clustered PCDHα genes. Thus, REST/NRSF represses PCDHα gene expression through directional binding to a repertoire of NRSEs within the distal enhancer and variable target genes.


Assuntos
Caderinas/metabolismo , Elementos Facilitadores Genéticos , Regulação da Expressão Gênica/genética , Regiões Promotoras Genéticas , Proteínas Repressoras/metabolismo , Dedos de Zinco , Animais , Caderinas/química , Caderinas/genética , Linhagem Celular Tumoral , Sequenciamento de Cromatina por Imunoprecipitação , Metilação de DNA , Humanos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , Simulação de Dinâmica Molecular , Família Multigênica , Ligação Proteica , Domínios Proteicos , RNA-Seq , Proteínas Repressoras/química , Proteínas Repressoras/genética
14.
Biomaterials ; 272: 120781, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33848809

RESUMO

We aimed to directly convert adult human dermal fibroblasts (aHDFs) into functional endothelial cells (ECs). Lentiviral vectors encoding endothelial transcription factors (TFs) were constructed. We examined whether five TFs (FOXO1, ER71, KLF2, TAL1, and LMO2) used for the generation of mouse induced ECs (iECs) could convert the aHDFs into human iECs. Twenty-eight days after transduction with lentiviral constructs, 32.1 ± 5.1% cells expressed vascular endothelial (VE)-cadherin. Factor screening revealed that only three factors (3F: ER71, KLF2, and TAL1) were necessary to induce VE-cadherin (+) cells (49.4 ± 3.5%). However, whole transcriptome sequencing showed that VE-cadherin (+) cells were not completely reprogrammed. Mature iECs double-positive for VE-cadherin/Pecam1 (DP cells) with a cobblestone appearance were obtained at a frequency of only 5.1 ± 0.6%. Using whole transcriptome analysis, the potential factors that could block the conversion were screened. Among candidates TWIST1-knockdown enhanced efficiency of conversion. Rosiglitazone, an inhibitor of epithelial-mesenchymal transition (EMT), also improved the conversion efficiency. Moreover, a 2nd second-stage conversion process, in which VE-cadherin (+) cells were incubated for additional two weeks, further enhanced the efficiency. The final protocol for 6 weeks yielded a conversion rate of 19.6 ± 3.0% iECs, defined by DP cells depicting the nature of mature ECs in various analyses. Further analyses revealed that the genetic and epigenetic profiles of iECs resembled those of functional ECs. Collectively, aHDFs can be converted into functional ECs through the transduction of ER71, KLF2, and TAL1, combined with two EMT inhibitors (siTWIST1 and rosiglitazone), followed by 2nd stage conversion.


Assuntos
Caderinas , Células Endoteliais , Adulto , Animais , Caderinas/genética , Transição Epitelial-Mesenquimal , Fibroblastos , Humanos , Camundongos , Molécula-1 de Adesão Celular Endotelial a Plaquetas , Fatores de Transcrição/genética
15.
Hum Genet ; 140(7): 1061-1076, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33811546

RESUMO

Teebi hypertelorism syndrome (THS; OMIM 145420) is a rare craniofacial disorder characterized by hypertelorism, prominent forehead, short nose with broad or depressed nasal root. Some cases of THS have been attributed to SPECC1L variants. Homozygous variants in CDH11 truncating the transmembrane and intracellular domains have been implicated in Elsahy-Waters syndrome (EWS; OMIM 211380) with hypertelorism. We report THS due to CDH11 heterozygous missense variants on 19 subjects from 9 families. All affected residues in the extracellular region of Cadherin-11 (CHD11) are highly conserved across vertebrate species and classical cadherins. Six of the variants that cluster around the EC2-EC3 and EC3-EC4 linker regions are predicted to affect Ca2+ binding that is required for cadherin stability. Two of the additional variants [c.164G > C, p.(Trp55Ser) and c.418G > A, p.(Glu140Lys)] are also notable as they are predicted to directly affect trans-homodimer formation. Immunohistochemical study demonstrates that CDH11 is strongly expressed in human facial mesenchyme. Using multiple functional assays, we show that five variants from the EC1, EC2-EC3 linker, and EC3 regions significantly reduced the cell-substrate trans adhesion activity and one variant from EC3-EC4 linker results in changes in cell morphology, focal adhesion, and migration, suggesting dominant negative effect. Characteristic features in this cohort included depressed nasal root, cardiac and umbilical defects. These features distinguished this phenotype from that seen in SPECC1L-related hypertelorism syndrome and CDH11-related EWS. Our results demonstrate heterozygous variants in CDH11, which decrease cell-cell adhesion and increase cell migratory behavior, cause a form of THS, as termed CDH11-related THS.


Assuntos
Anormalidades Múltiplas/genética , Caderinas/genética , Adesão Celular/genética , Anormalidades Craniofaciais/genética , Deformidades Congênitas do Pé/genética , Variação Genética/genética , Deformidades Congênitas da Mão/genética , Hipertelorismo/genética , Sequência de Aminoácidos , Movimento Celular/genética , Feminino , Heterozigoto , Homozigoto , Humanos , Masculino , Linhagem , Fenótipo
16.
Biomed Res Int ; 2021: 5559102, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33860034

RESUMO

Objective: To explore the function of miR-543 in endometrial cells and the possible mechanism of regulating the occurrence and development of intrauterine adhesion. Method: Endometrial epithelial cells and endometrial adenocarcinoma cells were transfected with miR-543 mimics and miR-543 inhibitor as the experimental group and were tested with the control group, using the CCK-8 method, scratch test, and Transwell assay, and flow cytometry was used to detect the proliferation, migration, invasion, and apoptosis of cells. RT-qPCR and Western blot were used to detect the expression of corresponding mRNA and protein. Results: After the overexpression of miR-543, endometrial epithelial cells and endometrial adenocarcinoma cells have reduced migratory, proliferative, and invasive capabilities, while the apoptosis rate has increased significantly. The mRNA expression of CDH2, COL16A1, vimentin, α-SMA and fibronectin decreased, and the protein expression of CDH2, vimentin, and α-SMA also decreased, while the mRNA and protein expression of CDH1 increased. The result after interfering with miR-543 is opposite, and luciferase reporter gene confirms that CDH2 is the target gene of miR-543. Conclusion: During the formation of intrauterine adhesions, the expression of CDH2, COL16A1, vimentin, and α-SMA may be inhibited by the high expression of miR-543, which may affect the degree of fibrosis and collagen content in the intrauterine adhesions, thereby inhibiting the occurrence and development of intrauterine adhesions.


Assuntos
Movimento Celular/genética , MicroRNAs/metabolismo , Aderências Teciduais/genética , Útero/patologia , Antígenos CD/genética , Antígenos CD/metabolismo , Apoptose/genética , Sequência de Bases , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/genética , Endométrio , Células Epiteliais/patologia , Feminino , Fibrose , Regulação Neoplásica da Expressão Gênica , Humanos , MicroRNAs/genética , Invasividade Neoplásica
17.
Science ; 372(6539)2021 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-33859005

RESUMO

Protocadherin-19 (PCDH19) mutations cause early-onset seizures and cognitive impairment. The PCDH19 gene is on the X-chromosome. Unlike most X-linked disorders, PCDH19 mutations affect heterozygous females (PCDH19HET♀ ) but not hemizygous males (PCDH19HEMI♂ ); however, the reason why remains to be elucidated. We demonstrate that PCDH19, a cell-adhesion molecule, is enriched at hippocampal mossy fiber synapses. Pcdh19HET♀ but not Pcdh19HEMI♂ mice show impaired mossy fiber synaptic structure and physiology. Consistently, Pcdh19HET♀ but not Pcdh19HEMI♂ mice exhibit reduced pattern completion and separation abilities, which require mossy fiber synaptic function. Furthermore, PCDH19 appears to interact with N-cadherin at mossy fiber synapses. In Pcdh19HET♀ conditions, mismatch between PCDH19 and N-cadherin diminishes N-cadherin-dependent signaling and impairs mossy fiber synapse development; N-cadherin overexpression rescues Pcdh19HET♀ phenotypes. These results reveal previously unknown molecular and cellular mechanisms underlying the female-specific PCDH19 disorder phenotype.


Assuntos
Caderinas/metabolismo , Disfunção Cognitiva/fisiopatologia , Doenças Genéticas Ligadas ao Cromossomo X/fisiopatologia , Fibras Musgosas Hipocampais/fisiopatologia , Sinapses/fisiologia , Animais , Região CA3 Hipocampal/fisiopatologia , Região CA3 Hipocampal/ultraestrutura , Caderinas/genética , Disfunção Cognitiva/genética , Modelos Animais de Doenças , Epilepsia/genética , Epilepsia/fisiopatologia , Feminino , Genes Ligados ao Cromossomo X , Doenças Genéticas Ligadas ao Cromossomo X/genética , Potenciação de Longa Duração , Masculino , Camundongos , Fibras Musgosas Hipocampais/ultraestrutura , Mutação , Caracteres Sexuais , Sinapses/ultraestrutura , beta Catenina/metabolismo
18.
Nutr Metab Cardiovasc Dis ; 31(5): 1542-1547, 2021 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-33810964

RESUMO

BACKGROUND AND AIMS: Novel genetic determinants associated with coronary artery disease (CAD) have been discovered by genome wide association studies. Variants encompassing the CELSR2- PSRC1-SORT1 gene cluster have been associated with CAD. This study is aimed to investigate the rs629301 polymorphism association with the extent of CAD evaluated by coronary angiography (CAG), and to evaluate its associations with an extensive panel of lipid and lipoprotein measurements in a large Italian cohort of 2429 patients. METHODS AND RESULTS: The patients were collected by four Intensive Care Units located in Palermo and Verona (Italy). Clinical Records were filed, blood samples were collected, lipids and apolipoproteins (apo) were measured in separate laboratories. CAD was defined by the presence of stenotic arteries (>50% lumen diameter) by CAG. The presence of CAD was associated with the rs629301 genotype. Patients with CAD were 78% and 73% (p = 0.007) of the T/T vs. T/G + G/G genotype carriers respectively. T/T genotype was also correlated with the number of stenotic arteries, with a 1.29 (1.04-1.61) risk to have a three-arteries disease. T/T genotype correlated with higher levels of LDL-, non-HDL cholesterol, apoB, apoE and apoCIII, and lower HDL-cholesterol. Logistic Regression confirmed that rs629301was associated with CAD independently from the common risk factors, with a risk similar to that conferred by ten years of age [odds ratios were 1.43 (1.04-1.96) and 1.39 (1.22-1.58) respectively]. CONCLUSIONS: rs629301 risk allele was independently associated with the extension and severity of CAD and positively with apoE and apoB containing lipoproteins.


Assuntos
Caderinas/genética , Angiografia Coronária , Estenose Coronária/diagnóstico por imagem , Estenose Coronária/genética , Polimorfismo de Nucleotídeo Único , Fatores Etários , Idoso , Biomarcadores/sangue , Estenose Coronária/sangue , Estenose Coronária/epidemiologia , Feminino , Estudos de Associação Genética , Predisposição Genética para Doença , Humanos , Itália/epidemiologia , Lipídeos/sangue , Masculino , Pessoa de Meia-Idade , Fenótipo , Valor Preditivo dos Testes , Prognóstico , Medição de Risco , Fatores de Risco , Índice de Gravidade de Doença , Fatores de Tempo
19.
Biomed Res Int ; 2021: 6644827, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33834070

RESUMO

Objective: This study is aimed at understanding the molecular mechanisms and exploring potential therapeutic targets for atrial fibrillation (AF) by multiomics analysis. Methods: Transcriptomics and methylation data of AF patients were retrieved from the Gene Expression Omnibus (GEO). Differentially expressed genes (DEGs) and differentially methylated sites between AF and normal samples were screened. Then, highly expressed and hypomethylated and lowly expressed and hypermethylated genes were identified for AF. Weighted gene coexpression network analysis (WGCNA) was presented to construct AF-related coexpression networks. 52 AF blood samples were used for whole exome sequence. The mutation was visualized by the maftools package in R. Key genes were validated in AF using independent datasets. Results: DEGs were identified between AF and controls, which were enriched in neutrophil activation and regulation of actin cytoskeleton. RHOA, CCR2, CASP8, and SYNPO2L exhibited abnormal expression and methylation, which have been confirmed to be related to AF. PCDHA family genes had high methylation and low expression in AF. We constructed two AF-related coexpression modules. Single-nucleotide polymorphism (SNP) was the most common mutation type in AF, especially T > C. MUC4 was the most frequent mutation gene, followed by PHLDA1, AHNAK2, and MAML3. There was no statistical difference in expression of AHNAK2 and MAML3, for AF. PHLDA1 and MUC4 were confirmed to be abnormally expressed in AF. Conclusion: Our findings identified DEGs related to DNA methylation and mutation for AF, which may offer possible therapeutic targets and a new insight into the pathogenesis of AF from a multiomics perspective.


Assuntos
Fibrilação Atrial/tratamento farmacológico , Fibrilação Atrial/genética , Epigênese Genética , Genômica , Terapia de Alvo Molecular , Idoso , Caderinas/genética , Caderinas/metabolismo , Metilação de DNA/genética , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Humanos , Masculino , Mutação/genética , Reprodutibilidade dos Testes , Sequenciamento Completo do Exoma
20.
Mymensingh Med J ; 30(2): 315-322, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33830108

RESUMO

Astrocytoma is the commonest primary brain tumor. These are feared due to their invasiveness in brain parenchyma so are less amenable to surgical removal and current chemotherapy regimens with a high mortality rate. Cell adhesion molecule (CAM) E-cadherin (CDH1) downregulation has been associated with tumors of different system and organs featuring invasion and metastasis. Therefore, the aim of the study was to find out the level of E-cadherin gene expression in low grade astrocytoma. In this cross-sectional study, 22 formalin fixed paraffin embedded tissue were taken as cases. Three non tumorous brain tissue and 1fresh post-mortem brain tissue were taken as control. Histological features were studied under light microscope with Haematoxylin and Eosin (H&E stain). Expression of CDH1 gene was analyzed by real time - polymerase chain reaction (RT-PCR) by comparative cyclic threshold (Ct) value method. The change in E-cadherin expression was measured by fold change in comparison with the control brain tissue. The data was tabulated and statistical analysis was performed. Among the 22 study cases 8(36.36%) were World Health Organization (WHO) Grade I and 14(63.63%) were WHO Grade II. All tumors showed downregulation of CDH1 gene in comparison with non-tumorous control tissue. The result is statistically significant (p=0.019). So, the study data revealed that downregulation of E-cadherin gene occurs in low grade astrocytoma and tumors of WHO Grade II showed more downregulation than Grade I tumors.


Assuntos
Astrocitoma , Neoplasias Encefálicas , Antígenos CD/genética , Astrocitoma/genética , Neoplasias Encefálicas/genética , Caderinas/genética , Estudos Transversais , Humanos , Reação em Cadeia da Polimerase em Tempo Real
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