RESUMEN
Liver cirrhosis is a condition with high mortality that poses a significant health and economic burden worldwide. The clinical characteristics of liver cirrhosis are complex and varied. Therefore, the evaluation of immune infiltration-involved genes incirrhosis has become mandatory in liver disease research, not only to identify the potential biomarkers but also to provide important insights into the underlying mechanisms of the disease. In this study, we aimed to investigate the expression profile of cytokine genes in peripheral blood mononuclear cells (PBMCs) of HCV patients and identify the gene expression signature associated with advanced cirrhosis. A cross-sectional study of 90 HCV genotype 4 patients, including no fibrosis patients (F0, n = 24), fibrotic patients (F1-F3, n = 36), and cirrhotic patients (F4, n = 30) has been conducted. The expression of cytokine genes was analyzed by quantitative real-time PCR in the subjects' PBMCs, and the serum level of TGFß2 was measured by ELISA. Our findings showed that the expression level of the TGIF1 transcript was lower in cirrhotic and fibrotic patients compared to no fibrosis patients (p = 0.046 and 0.022, respectively). Also, there was an upregulation of the TGFß1 gene in cirrhotic patients relative to fibrotic patients (p = 0.015). Additionally, the cirrhotic patients had higher expression levels of the TGF-ß2 transcript and elevated levels of the TGF-ß2 protein than patients with no cirrhosis or fibrosis. According to the ROC analysis, TGFß1, TGIF1 transcripts, and TGFß2 protein have a good discriminatory performance in distinguishing between cirrhotic, fibrotic, and non-fibrotic patients. Our results suggested that the expression of TGIF1, TGF-ß1, and TGF-ß2 genes in PBMCs may provide a valuable tool for identifying patients with advanced cirrhosis and that TGF-ß and TGIF1 may be potential biomarkers for cirrhosis. These findings may have implications for the diagnosis and treatment of cirrhosis in HCV patients.
Asunto(s)
Biomarcadores , Leucocitos Mononucleares , Cirrosis Hepática , Humanos , Cirrosis Hepática/genética , Cirrosis Hepática/diagnóstico , Cirrosis Hepática/sangre , Cirrosis Hepática/virología , Masculino , Femenino , Biomarcadores/sangre , Biomarcadores/metabolismo , Persona de Mediana Edad , Leucocitos Mononucleares/metabolismo , Estudios Transversales , Hepatitis C/genética , Hepatitis C/complicaciones , Hepatitis C/diagnóstico , Hepacivirus , Adulto , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta1/genética , Factor de Crecimiento Transformador beta1/sangre , Citocinas/sangre , Citocinas/genética , Regulación de la Expresión GénicaRESUMEN
Gemcitabine resistance is a major obstacle to the effectiveness of chemotherapy in pancreatic ductal adenocarcinoma (PDAC). Therefore, new strategies are needed to sensitize cancer cells to gemcitabine. Here, we constructed gemcitabine-resistant PDAC cells and analyzed them with RNA-sequence. Employing an integrated approach involving bioinformatic analyses from multiple databases, TGFB2 is identified as a crucial gene in gemcitabine-resistant PDAC and is significantly associated with poor gemcitabine therapeutic response. The patient-derived xenograft (PDX) model further substantiates the gradual upregulation of TGFB2 expression during gemcitabine-induced resistance. Silencing TGFB2 expression can enhance the chemosensitivity of gemcitabine against PDAC. Mechanistically, TGFB2, post-transcriptionally stabilized by METTL14-mediated m6A modification, can promote lipid accumulation and the enhanced triglyceride accumulation drives gemcitabine resistance by lipidomic profiling. TGFB2 upregulates the lipogenesis regulator sterol regulatory element binding factor 1 (SREBF1) and its downstream lipogenic enzymes via PI3K-AKT signaling. Moreover, SREBF1 is responsible for TGFB2-mediated lipogenesis to promote gemcitabine resistance in PDAC. Importantly, TGFB2 inhibitor imperatorin combined with gemcitabine shows synergistic effects in gemcitabine-resistant PDAC PDX model. This study sheds new light on an avenue to mitigate PDAC gemcitabine resistance by targeting TGFB2 and lipid metabolism and develops the potential of imperatorin as a promising chemosensitizer in clinical translation.
Asunto(s)
Adenosina , Carcinoma Ductal Pancreático , Desoxicitidina , Resistencia a Antineoplásicos , Gemcitabina , Metabolismo de los Lípidos , Neoplasias Pancreáticas , Factor de Crecimiento Transformador beta2 , Humanos , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacología , Carcinoma Ductal Pancreático/tratamiento farmacológico , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patología , Factor de Crecimiento Transformador beta2/metabolismo , Factor de Crecimiento Transformador beta2/genética , Resistencia a Antineoplásicos/genética , Animales , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Metabolismo de los Lípidos/efectos de los fármacos , Metabolismo de los Lípidos/genética , Ratones , Adenosina/análogos & derivados , Adenosina/farmacología , Adenosina/metabolismo , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto , Transducción de Señal/efectos de los fármacos , Reprogramación Metabólica , Proteína 1 de Unión a los Elementos Reguladores de EsterolesRESUMEN
This study aimed to assess the expression profile of messenger RNA (mRNA) and microRNA (miRNA) related to the dopaminergic system in five types of breast cancer in Polish women. Patients with five breast cancer subtypes were included in the study: luminal A (n = 130), luminal B (n = 196, including HER2-, n = 100; HER2+, n = 96), HER2+ (n = 36), and TNBC (n = 43); they underwent surgery, during which tumor tissue was removed along with a margin of healthy tissue (control material). The molecular analysis included a microarray profile of mRNAs and miRNAs associated with the dopaminergic system, a real-time polymerase chain reaction preceded by reverse transcription for selected genes, and determinations of their concentration using enzyme-linked immunosorbent assay (ELISA). The conducted statistical analysis showed that five mRNAs statistically significantly differentiated breast cancer sections regardless of subtype compared to control samples; these were dopamine receptor 2 (DRD2), dopamine receptor 3 (DRD3), dopamine receptor 25 (DRD5), transforming growth factor beta 2 (TGF-ß-2), and caveolin 2 (CAV2). The predicted analysis showed that hsa-miR-141-3p can regulate the expression of DRD2 and TGF-ß-2, whereas hsa-miR-4441 is potentially engaged in the expression regulation of DRD3 and DRD5. In addition, the expression pattern of DRD5 mRNA can also be regulated by has-miR-16-5p. The overexpression of DRD2 and DRD3, with concomitant silencing of DRD5 expression, confirms the presence of dopaminergic abnormalities in breast cancer patients. Moreover, these abnormalities may be the result of miR-141-3P, miR-16-5p, and miR-4441 activity, regulating proliferation or metastasis.
Asunto(s)
Neoplasias de la Mama , Dopamina , Regulación Neoplásica de la Expresión Génica , MicroARNs , Humanos , Femenino , MicroARNs/genética , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Neoplasias de la Mama/metabolismo , Persona de Mediana Edad , Dopamina/metabolismo , Adulto , Perfilación de la Expresión Génica/métodos , Anciano , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores de Dopamina D3/genética , Receptores de Dopamina D3/metabolismo , Receptores de Dopamina D2/genética , Receptores de Dopamina D2/metabolismo , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta2/metabolismoRESUMEN
Primary open-angle glaucoma (POAG) is a progressive optic neuropathy with a complex, multifactorial aetiology. Raised intraocular pressure (IOP) is the most important clinically modifiable risk factor for POAG. All current pharmacological agents target aqueous humour dynamics to lower IOP. Newer therapeutic agents are required as some patients with POAG show a limited therapeutic response or develop ocular and systemic side effects to topical medication. Elevated IOP in POAG results from cellular and molecular changes in the trabecular meshwork driven by increased levels of transforming growth factor ß (TGFß) in the anterior segment of the eye. Understanding how TGFß affects both the structural and functional changes in the outflow pathway and IOP is required to develop new glaucoma therapies that target the molecular pathology in the trabecular meshwork. In this study, we evaluated the effects of TGF-ß1 and -ß2 treatment on miRNA expression in cultured human primary trabecular meshwork cells. Our findings are presented in terms of specific miRNAs (miRNA-centric), but given miRNAs work in networks to control cellular pathways and processes, a pathway-centric view of miRNA action is also reported. Evaluating TGFß-responsive miRNA expression in trabecular meshwork cells will further our understanding of the important pathways and changes involved in the pathogenesis of glaucoma and could lead to the development of miRNAs as new therapeutic modalities in glaucoma.
Asunto(s)
MicroARNs , Malla Trabecular , Malla Trabecular/metabolismo , Malla Trabecular/efectos de los fármacos , Malla Trabecular/patología , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Glaucoma de Ángulo Abierto/genética , Glaucoma de Ángulo Abierto/metabolismo , Glaucoma de Ángulo Abierto/patología , Factor de Crecimiento Transformador beta2/metabolismo , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta/metabolismo , Factor de Crecimiento Transformador beta1/metabolismo , Células Cultivadas , Regulación de la Expresión Génica/efectos de los fármacos , Presión Intraocular/efectos de los fármacosRESUMEN
BACKGROUND: Herein, we report results from a genome-wide study conducted to identify protein quantitative trait loci (pQTL) for circulating angiogenic and inflammatory protein markers in patients with metastatic colorectal cancer (mCRC). The study was conducted using genotype, protein marker, and baseline clinical and demographic data from CALGB/SWOG 80405 (Alliance), a randomized phase III study designed to assess outcomes of adding VEGF or EGFR inhibitors to systemic chemotherapy in mCRC patients. Germline DNA derived from blood was genotyped on whole-genome array platforms. The abundance of protein markers was quantified using a multiplex enzyme-linked immunosorbent assay from plasma derived from peripheral venous blood collected at baseline. A robust rank-based method was used to assess the statistical significance of each variant and protein pair against a strict genome-wide level. A given pQTL was tested for validation in two external datasets of prostate (CALGB 90401) and pancreatic cancer (CALGB 80303) patients. Bioinformatics analyses were conducted to further establish biological bases for these findings. RESULTS: The final analysis was carried out based on data from 540,021 common typed genetic variants and 23 protein markers from 869 genetically estimated European patients with mCRC. Correcting for multiple testing, the analysis discovered a novel cis-pQTL in LINC02869, a long non-coding RNA gene, for circulating TGF-ß2 levels (rs11118119; AAF = 0.11; P-value < 1.4e-14). This finding was validated in a cohort of 538 prostate cancer patients from CALGB 90401 (AAF = 0.10, P-value < 3.3e-25). The analysis also validated a cis-pQTL we had previously reported for VEGF-A in advanced pancreatic cancer, and additionally identified trans-pQTLs for VEGF-R3, and cis-pQTLs for CD73. CONCLUSIONS: This study has provided evidence of a novel cis germline genetic variant that regulates circulating TGF-ß2 levels in plasma of patients with advanced mCRC and prostate cancer. Moreover, the validation of previously identified pQTLs for VEGF-A, CD73, and VEGF-R3, potentiates the validity of these associations.
Asunto(s)
Neoplasias Colorrectales , ARN Largo no Codificante , Factor de Crecimiento Transformador beta2 , Humanos , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/sangre , Neoplasias Colorrectales/patología , Masculino , Femenino , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta2/sangre , ARN Largo no Codificante/sangre , ARN Largo no Codificante/genética , Sitios de Carácter Cuantitativo , Persona de Mediana Edad , Metástasis de la Neoplasia , Anciano , Polimorfismo de Nucleótido Simple , Biomarcadores de Tumor/sangre , Biomarcadores de Tumor/genética , Estudio de Asociación del Genoma CompletoRESUMEN
The study investigates molecular changes in the lumbosacral (L/S) spine's yellow ligamentum flavum during degenerative stenosis, focusing on the role of transforming growth factor beta 1-3 (TGF-ß-1-3). Sixty patients with degenerative stenosis and sixty control participants underwent molecular analysis using real-time quantitative reverse transcription reaction technique (RTqPCR), enzyme-linked immunosorbent assay (ELISA), Western blot, and immunohistochemical analysis (IHC). At the mRNA level, study samples showed reduced expression of TGF-ß-1 and TGF-ß-3, while TGF-ß-2 increased by only 4%. Conversely, at the protein level, the study group exhibited significantly higher concentrations of TGF-ß-1, TGF-ß-2, and TGF-ß-3 compared to controls. On the other hand, at the protein level, a statistically significant higher concentration of TGF-ß-1 was observed (2139.33 pg/mL ± 2593.72 pg/mL vs. 252.45 pg/mL ± 83.89 pg/mL; p < 0.0001), TGF-ß-2 (3104.34 pg/mL ± 1192.74 pg/mL vs. 258.86 pg/mL ± 82.98 pg/mL; p < 0.0001), TGF-ß-3 (512.75 pg/mL ± 107.36 pg/mL vs. 55.06 pg/mL ± 9.83 pg/mL, p < 0.0001) in yellow ligaments obtained from patients of the study group compared to control samples. The study did not establish a significant correlation between TGF-ß-1-3 concentrations and pain severity. The findings suggest that molecular therapy aimed at restoring the normal expression pattern of TGF-ß-1-3 could be a promising strategy for treating degenerative stenosis of the L/S spine. The study underscores the potential therapeutic significance of addressing molecular changes at the TGF-ß isoforms level for better understanding and managing degenerative spinal conditions.
Asunto(s)
Isoformas de Proteínas , Estenosis Espinal , Humanos , Femenino , Masculino , Persona de Mediana Edad , Isoformas de Proteínas/metabolismo , Isoformas de Proteínas/genética , Estenosis Espinal/metabolismo , Estenosis Espinal/patología , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Crecimiento Transformador beta1/genética , Anciano , Factor de Crecimiento Transformador beta2/metabolismo , Factor de Crecimiento Transformador beta2/genética , Ligamento Amarillo/metabolismo , Ligamento Amarillo/patología , Factor de Crecimiento Transformador beta/metabolismo , Factor de Crecimiento Transformador beta/genética , ARN Mensajero/metabolismo , ARN Mensajero/genética , Factor de Crecimiento Transformador beta3/metabolismo , Factor de Crecimiento Transformador beta3/genética , Adulto , Vértebras Lumbares/metabolismo , Vértebras Lumbares/patología , Región Lumbosacra/patología , Estudios de Casos y ControlesRESUMEN
BACKGROUND: Posterior capsular opacification (PCO) is the main reason affecting the long-term postoperative result of cataract patient, and it is well accepted that fibrotic PCO is driven by transforming growth factor beta (TGFß) signaling. Ferroptosis, closely related to various ocular diseases, but has not been explored in PCO. METHODS: RNA sequencing (RNA-seq) was performed on both TGF-ß2 treated and untreated primary lens epithelial cells (pLECs). Differentially expressed genes (DEGs) associated with ferroptosis were analyzed using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) to investigate their biological function. Additionally, protein-to-protein interactions among selected ferroptosis-related genes by PPI network and the top 10 genes with the highest score (MCC algorithm) were selected as the hub genes. The top 20 genes with significant fold change values were validated using quantitative real-time polymerase chain reaction (qRT-PCR). RESULTS: Our analysis revealed 1253 DEGs between TGF-ß2 treated and untreated pLECs, uncovering 38 ferroptosis-related genes between two groups. Among these 38 ferroptosis-related genes,the most prominent GO enrichment analysis process involved in the response to oxidative stress (BPs), apical part of cell (CCs),antioxidant activity (MFs). KEGG were mainly concentrated in fluid shear stress and atherosclerosis, IL-17 and TNF signaling pathways, and validation of top 20 genes with significant fold change value were consistent with RNA-seq. CONCLUSIONS: Our RNA-Seq data identified 38 ferroptosis-related genes in TGF-ß2 treated and untreated pLECs, which is the first observation of ferroptosis related genes in primary human lens epithelial cells under TGF-ß2 stimulation.
Asunto(s)
Opacificación Capsular , Ferroptosis , Humanos , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta2/metabolismo , Factor de Crecimiento Transformador beta2/farmacología , Transcriptoma , Transición Epitelial-Mesenquimal/genética , Ferroptosis/genética , Western Blotting , Opacificación Capsular/genética , Opacificación Capsular/metabolismo , Células Epiteliales/metabolismoRESUMEN
The mesenchymal subtype of glioblastoma (GBM) cells characterized by aggressive invasion and therapeutic resistance is thought to be dependent on cell-intrinsic alteration and extrinsic cellular crosstalk. Tumor-associated macrophages (TAMs) are pivotal in tumor progression, chemo-resistance, angiogenesis, and stemness maintenance. However, the impact of TAMs on the shifts in glioma stem cells (GSCs) states remains largely uncovered. Herein, we showed that the triggering receptor expressed on myeloid cells-1 (TREM1) preferentially expressed by M2-like TAMs and induced GSCs into mesenchymal-like states by modulating the secretion of TGFß2, which activated the TGFßR/SMAD2/3 signaling in GSCs. Furthermore, we demonstrated that TREM1 was transcriptionally regulated by HIF1a under the hypoxic environment and thus promoted an immunosuppressive type of TAMs via activating the TLR2/AKT/mTOR/c-MYC axis. Collectively, this study reveals that cellular communication between TAMs and GSCs through the TREM1-mediated TGFß2/TGFßR axis is involved in the mesenchymal-like transitions of GSCs. Our study provides valuable insights into the regulatory mechanisms between the tumor immune microenvironment and the malignant characteristics of GBM, which can lead to potential novel strategies targeting TAMs for tumor control.
Asunto(s)
Células Madre Neoplásicas , Receptor Activador Expresado en Células Mieloides 1 , Microambiente Tumoral , Macrófagos Asociados a Tumores , Humanos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Células Madre Neoplásicas/inmunología , Receptor Activador Expresado en Células Mieloides 1/metabolismo , Receptor Activador Expresado en Células Mieloides 1/genética , Macrófagos Asociados a Tumores/metabolismo , Macrófagos Asociados a Tumores/inmunología , Animales , Línea Celular Tumoral , Transducción de Señal , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/inmunología , Glioma/patología , Glioma/genética , Glioma/metabolismo , Glioma/inmunología , Ratones , Glioblastoma/patología , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/inmunología , Factor de Crecimiento Transformador beta2/metabolismo , Factor de Crecimiento Transformador beta2/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Regulación Neoplásica de la Expresión Génica , Proteína Smad2/metabolismo , Proteína Smad2/genéticaRESUMEN
Clear cell renal cell carcinoma (ccRCC) is a malignant tumor of kidney epithelial cells, one of the most common tumors in the world. Transforming growth factor beta (TGFß)1 is a crucial factor that induces epithelial-mesenchymal transition (EMT) in cancer cells. microRNA-141-3p (miR-141-3p) is a microRNA that is considered a tumor suppressor. However, the role and mechanism of miR-141-3p in TGFß1-induced ccRCC cells are not fully understood. This study investigated the roles of miR-141-3p and its target gene in regulating EMT in ccRCC development. 786-0 and Caki-1cells were treated with TGFß1 to induce EMT. The levels of miR-141-3p and TGFß2 were determined by quantitative real-time polymerase chain reaction and Western blotting. The progression of EMT was evaluated by E-cadherin detection by immunofluorescence, and E-cadherin, N-cadherin, and vimentin detection by Western blotting. Furthermore, migration and invasion capacities were assessed using a Transwell system. The direct binding of miR-141-3p with the target gene TGFß2 was confirmed by dual luciferase reporter gene assay. Results indicated that TGFß1 treatment decreased the protein abundance of E-cadherin while increasing the protein expression of N-cadherin and vimentin, indicating TGFß1-induced EMT was constructed successfully. Moreover, TGFß1 treatment repressed the expression of miR-141-3p. miR-141-3p mimics reversed the effect of TGFß1 on the migration, invasion, and expression of E-cadherin, N-cadherin, and vimentin. The miR-141-3p directly binds with the 3' untranslated region of TGFß2 mRNA and suppresses its expression. Furthermore, TGFß2 overexpression abrogated the above changes regulated by miR-141-3p mimics. Taken together, miR-141-3p inhibited TGFß1-induced EMT by suppressing the migration and invasion of ccRCC cells via directly targeting TGFß2 gene expression.
Asunto(s)
Carcinoma de Células Renales , Movimiento Celular , Transición Epitelial-Mesenquimal , Regulación Neoplásica de la Expresión Génica , Neoplasias Renales , MicroARNs , Factor de Crecimiento Transformador beta2 , MicroARNs/genética , MicroARNs/metabolismo , Humanos , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/metabolismo , Carcinoma de Células Renales/patología , Transición Epitelial-Mesenquimal/genética , Transición Epitelial-Mesenquimal/efectos de los fármacos , Neoplasias Renales/genética , Neoplasias Renales/metabolismo , Neoplasias Renales/patología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta2/metabolismo , Factor de Crecimiento Transformador beta2/farmacología , Línea Celular Tumoral , Movimiento Celular/genética , Movimiento Celular/efectos de los fármacos , Factor de Crecimiento Transformador beta1/farmacología , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Crecimiento Transformador beta1/genética , Progresión de la EnfermedadRESUMEN
OBJECTIVE: The study objectives were to 1) determine the mesenchymal stem cell (MSC) surface expression of major histocompatibility complex (MHC) class I and transcriptome-wide gene expression changes following IL-1ß + TGF-ß2 dual licensing and 2) evaluate if IL-1ß + TGF-ß2 dual-licensed MSCs had a greater ability to positively modulate tenocyte function compared to naive MSCs. SAMPLE: Equine bone marrow-derived MSCs from 6 donors and equine superficial digital flexor tenocytes from 3 donors. METHODS: Experiments were performed in vitro. Flow cytometry and bulk RNA sequencing were utilized to determine naive and dual-licensed MSC phenotype and transcriptome-wide changes in gene expression. Conditioned media were generated from MSCs and utilized in tenocyte cell culture assays as a method to determine the effect of MSC paracrine factors on tenocyte function. RESULTS: Dual-licensed MSCs have a reduced expression of MHC class I and exhibit enrichment in functional pathways associated with the extracellular matrix, cell signaling, and tissue development. Additionally, dual-licensed MSC-conditioned media significantly improved in vitro tenocyte migration and metabolism to a greater degree than naive MSC-conditioned media. In tenocytes exposed to IL-1ß, dual-licensed conditioned media also positively modulated tenocyte gene expression. CLINICAL RELEVANCE: Our data indicate that conditioned media containing paracrine factors secreted from dual-licensed MSCs significantly modulates in vitro tenocyte function, which may confer benefits in vivo to healing tendons following injury. Additionally, due to reduced MHC class I expression in dual-licensed MSCs, this technique may also provide an avenue to provide an effective "off-the-shelf" allogenic source of MSCs.
Asunto(s)
Interleucina-1beta , Células Madre Mesenquimatosas , Tenocitos , Factor de Crecimiento Transformador beta2 , Animales , Células Madre Mesenquimatosas/metabolismo , Caballos , Interleucina-1beta/metabolismo , Interleucina-1beta/genética , Factor de Crecimiento Transformador beta2/metabolismo , Factor de Crecimiento Transformador beta2/genética , Tenocitos/metabolismo , Movimiento Celular/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Antígenos de Histocompatibilidad Clase I/metabolismo , Antígenos de Histocompatibilidad Clase I/genética , Células CultivadasRESUMEN
BACKGROUND: Leber hereditary optic neuropathy (LHON) is mainly the degeneration of retinal ganglion cells (RGCs) associated with high apoptosis and reactive oxygen species (ROS) levels, which is accepted to be caused by the mutations in the subunits of complex I of the mitochondrial electron transport chain. The treatment is still infant while efforts of correcting genes or using antioxidants do not bring good and consistent results. Unaffected carrier carries LHON mutation but shows normal phenotype, suggesting that the disease's pathogenesis is complex, in which secondary factors exist and cooperate with the primary complex I dysfunction. METHODS: Using LHON patient-specific induced pluripotent stem cells (iPSCs) as the in vitro disease model, we previously demonstrated that circRNA_0087207 had the most significantly higher expression level in the LHON patient-iPSC-derived RGCs compared with the unaffected carrier-iPSC-derived RGCs. To elaborate the underlying pathologies regulated by circRNA_008720 mechanistically, bioinformatics analysis was conducted and elucidated that circRNA_0087207 could act as a sponge of miR-548c-3p and modulate PLSCR1/TGFB2 levels in ND4 mutation-carrying LHON patient-iPSC-derived RGCs. RESULTS: Using LHON iPSC-derived RGCs as the disease-based platform, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis on targeted mRNA of miR-548c-3p showed the connection with apoptosis, suggesting downregulation of miR548c-3p contributes to the apoptosis of LHON patient RGCs. CONCLUSION: We showed that the downregulation of miR548c-3p plays a critical role in modulating cellular dysfunction and the apoptotic program of RGCs in LHON.
Asunto(s)
MicroARNs , Atrofia Óptica Hereditaria de Leber , Humanos , Atrofia Óptica Hereditaria de Leber/genética , Atrofia Óptica Hereditaria de Leber/patología , ARN Circular/genética , Mitocondrias , Apoptosis , Mutación , MicroARNs/genética , MicroARNs/metabolismo , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta2/metabolismoRESUMEN
N6-methyladenosine (m6A) is a major type of RNA modification implicated in various pathophysiological processes. Transforming growth factor ß2 (TGF-ß2) induces epithelial-mesenchymal transition (EMT) in retinal pigmental epithelial (RPE) cells and promotes the progression of proliferative vitreoretinopathy (PVR). However, the role of m6A methylation in the EMT of human telomerase reverse transcriptase (hTERT) retinal pigmental epithelium (RPE)-1 cells has not been clarified. Here, we extracted RNA from RPE cells subjected to 0 or 20 ng/mL TGF-ß2 for 72 h and identified differentially methylated genes (DMGs) by m6A-Seq and differentially expressed genes (DEGs) by RNA-Seq. We selected the genes related to EMT by conjoint m6A-Seq/RNA-Seq analysis and verified them by qRT-PCR. We then confirmed the function of m6A methylation in the EMT of RPE cells by knocking down the methyltransferase METTL3 and the m6A reading protein YTHDF1. Sequencing yielded 5814 DMGs and 1607 DEGs. Conjoint analysis selected 467 genes altered at the m6A and RNA levels that are closely associated with the EMT-related TGF-ß, AGE-RAGE, PI3K-Akt, P53, and Wnt signaling pathways. We also identified ten core EMT genes ACTG2, BMP6, CDH2, LOXL2, SNAIL1, SPARC, BMP4, EMP3, FOXM1, and MYC. Their RNA levels were evaluated by qRT-PCR and were consistent with the sequencing results. We observed that METTL3 knockdown enhanced RPE cell migration and significantly upregulated the EMT markers N-cadherin (encoded by CDH2), fibronectin (FN), Snail family transcription repressor (SLUG), and vimentin. However, YTHDF1 knockdown had the opposite effects and decreased both cell migration and the N-cadherin, FN, and SLUG expression levels. The present study clarified TGF-ß2-induced m6A- and RNA-level differences in RPE cells, indicated that m6A methylation might regulate EMT marker expression, and showed that m6A could regulate TGF-ß2-induced EMT.
Asunto(s)
Adenina/análogos & derivados , Factor de Crecimiento Transformador beta2 , Vitreorretinopatía Proliferativa , Humanos , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta2/farmacología , Factor de Crecimiento Transformador beta2/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Epitelio Pigmentado de la Retina/metabolismo , Vitreorretinopatía Proliferativa/genética , Vitreorretinopatía Proliferativa/metabolismo , Transición Epitelial-Mesenquimal , Metilación , Cadherinas/genética , Cadherinas/metabolismo , ARN/genética , ARN/metabolismo , Metiltransferasas/metabolismo , Glicoproteínas de Membrana/metabolismoRESUMEN
BACKGROUND: Age-related macular degeneration (AMD) is a prevalent source of visual impairment among the elderly population, and its incidence has risen in tandem with the increasing longevity of humans. Despite the progress made with anti-VEGF therapy, clinical outcomes have proven to be unsatisfactory. METHOD: We obtained differentially expressed genes (DEGs) of AMD patients and healthy controls from the GEO database. GO and KEGG analyses were used to enrich the DEGs. Weighted gene coexpression network analysis (WGCNA) was used to identify modules related to AMD. SVM, random forest, and least absolute shrinkage and selection operator (LASSO) were employed to screen hub genes. Gene set enrichment analysis (GSEA) was used to explore the pathways in which these hub genes were enriched. CIBERSORT was utilized to analyze the relationship between the hub genes and immune cell infiltration. Finally, Western blotting and RTâPCR were used to explore the expression of hub genes in AMD mice. RESULTS: We screened 1084 DEGs in GSE29801, of which 496 genes were upregulated. These 1084 DEGs were introduced into the WGCNA, and 94 genes related to AMD were obtained. Seventy-nine overlapping genes were obtained by the Venn plot. These 79 genes were introduced into three machine-learning methods to screen the hub genes, and the genes identified by the three methods were TNC, FAP, SREBF1, and TGF-ß2. We verified their diagnostic function in the GSE29801 and GSE103060 datasets. Then, the hub gene co-enrichment pathways were obtained by GO and KEGG analyses. CIBERSORT analysis showed that these hub genes were associated with immune cell infiltration. Finally, we found increased expression of TNC, FAP, SREBF1, and TGF-ß2 mRNA and protein in the retinas of AMD mice. CONCLUSION: We found that four hub genes, namely, FAP, TGF-ß2, SREBF1, and TNC, have diagnostic significance in patients with AMD and are related to immune cell infiltration. Finally, we determined that the mRNA and protein expression of these hub genes was upregulated in the retinas of AMD mice.
Asunto(s)
Degeneración Macular , Factor de Crecimiento Transformador beta2 , Humanos , Anciano , Animales , Ratones , Factor de Crecimiento Transformador beta2/genética , Degeneración Macular/genética , Retina , Western Blotting , ARN MensajeroRESUMEN
Ovarian hyperstimulation syndrome (OHSS) is a life-threatening and potentially fatal complication during in vitro fertilization treatment. The levels of transforming growth factor-ß1 (TGF-ß1) are upregulated in human follicular fluid and granulosa-lutein cells (hGL) of OHSS patients and could contribute to the development of OHSS by downregulating steroidogenic acute regulatory protein (StAR) expression. However, whether the same is true for the other two members of the TGF-ß family, TGF-ß2 and -ß3, remains unknown. We showed that all three TGF-ß isoforms were expressed in human follicular fluid. In comparison, TGF-ß1 was expressed at the highest level, followed by TGF-ß2 and TGF-ß3. Compared to non-OHSS patients, follicular fluid levels of TGF-ß1 and TGF-ß3 were significantly upregulated in OHSS patients. The same results were observed in mRNA levels of TGF-ß isoforms in hGL cells and ovaries of OHSS rats. In addition, StAR mRNA levels were upregulated in hGL cells of OHSS patients and the ovaries of OHSS rats. Treatment cells with TGF-ß isoforms downregulated the StAR expression with a comparable effect. Moreover, activations of SMAD3 signaling were required for TGF-ß isoforms-induced downregulation of StAR expression. This study indicates that follicular fluid TGF-ß1 and TGF-ß3 levels could be used as biomarkers and therapeutic targets for the OHSS.
Asunto(s)
Síndrome de Hiperestimulación Ovárica , Factor de Crecimiento Transformador beta1 , Femenino , Humanos , Ratas , Animales , Factor de Crecimiento Transformador beta1/genética , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Crecimiento Transformador beta3/genética , Factor de Crecimiento Transformador beta3/metabolismo , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta2/metabolismo , Síndrome de Hiperestimulación Ovárica/genética , ARN Mensajero/metabolismo , Isoformas de ProteínasRESUMEN
Oncolytic virotherapy is a promising therapeutic approach for glioblastoma (GBM) treatment, although the outcomes are partially satisfactory. Hence, more effective strategies are needed urgently to modify therapeutic viruses to enhance their efficiency and safety in killing tumor cells and improve the survival rate of GBM patients. This study generated a new-generation oncolytic adenovirus Ad5 KT-E1A-IL-15 (TS-2021) and evaluated its antitumor efficacy. Ex vivo analyses revealed Ki67 and TGF-ß2 co-localized in GBM cells. In addition, TS-2021 selectively replicated in GBM cells, which was dependent on the expression of Ki67 and TGF-ß2. The immunocompetent mice model of GBM demonstrated the in vivo efficacy of TS-2021 by inhibiting tumor growth and improving survival proficiently. Notably, TS-2021 effectively reduced MMP3 expression by inactivating the MKK4/JNK pathway, thereby reducing tumor invasiveness. Altogether, the findings of the present study highlight that TS-2021 can effectively target GBM cells expressing high levels of Ki67 and TGF-ß2, exerting potent antitumor effects. Additionally, it can improve efficacy and suppress tumor invasiveness by inhibiting the MKK4/JNK/MMP3 pathway. Thus our study demonstrates the efficiency of the novel TS-2021 in the mouse model and provides a potential therapeutic option for patients with GBM.
Asunto(s)
Infecciones por Adenoviridae , Glioblastoma , Animales , Ratones , Humanos , Adenoviridae/genética , Glioblastoma/terapia , Glioblastoma/genética , Glioblastoma/patología , Regiones no Traducidas 5' , Antígeno Ki-67/genética , Antígeno Ki-67/metabolismo , Metaloproteinasa 3 de la Matriz/metabolismo , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta2/metabolismo , Interleucina-15/metabolismo , Línea Celular TumoralRESUMEN
Transforming growth factor-beta 2 (TGF-ß2), an important member of the TGF-ß family, is a secreted protein that is involved in many biological processes, such as cell growth, proliferation, migration, and differentiation. TGF-ß2 had been thought to be functionally identical to TGF-ß1; however, an increasing number of recent studies uncovered the distinctive features of TGF-ß2 in terms of its expression, activation, and biological functions. Mice deficient in TGF-ß2 showed remarkable developmental abnormalities in multiple organs, especially the cardiovascular system. Dysregulation of TGF-ß2 signalling was associated with tumorigenesis, eye diseases, cardiovascular diseases, immune disorders, as well as motor system diseases. Here, we provide a comprehensive review of the research progress in TGF-ß2 to support further research on TGF-ß2.
Asunto(s)
Factor de Crecimiento Transformador beta2 , Factores de Crecimiento Transformadores , Ratones , Animales , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta2/metabolismo , Diferenciación Celular , Ciclo Celular , Proliferación CelularRESUMEN
The aim of this study was to investigate the mechanism underlying the role of a recently identified hsa_circ_0004805/hsa_miR-149-5p/transforming growth factor beta 2 (TGFB2) axis in the progression of diabetic retinopathy (DR). Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis suggested that hsa_circ_0004805 was highly expressed in aqueous humor samples of patients with DR, whereas hsa_miR-149-5p showed the opposite trend. Meanwhile, the results of a dual-luciferase reporter assay indicated that hsa_miR-149-5p directly interacted with both hsa_circ_0004805 and TGFB2. Using a variety of assays (Cell Counting Kit-8, EdU-labeling, Transwell, flow cytometric, wound healing, tube formation assays), we found that the overexpression of hsa_circ_0004805 significantly downregulated the level of hsa_miR-149-5p and promoted DNA synthesis, proliferation, migration, and tube formation in human retinal microvascular epithelial cells (hRECs) cultivated in a high-glucose environment. In contrast, hsa_miR-149-5p mimics inhibited DNA synthesis, proliferation, migration, and tube formation in hRECs by reducing the expression of its downstream target TGFB2 as well as the levels of phosphorylated SMAD2; however, these effects were reversed by the overexpression of hsa_circ_0004805. In a streptozotocin-induced Sprague-Dawley rat model of DR, retinal vascular leakage, capillary decellularization, loss of pericytes, fibrosis, and gliosis were evident, which could be reversed by vitreous microinjection of rat miR-149-5p mimics (rno-miR-149-5p agomir). Combined, our findings indicated that, under hyperglycemia, the hsa_circ_0004805/hsa_miR-149-5p/TGFB2 axis plays a critical role in the retinal pathophysiology associated with the development of DR, and has potential as a therapeutic target in the treatment of this condition.
Asunto(s)
Diabetes Mellitus , Retinopatía Diabética , MicroARNs , Ratas , Animales , Humanos , Ratas Sprague-Dawley , Retinopatía Diabética/genética , MicroARNs/genética , Retina , ADN , Proliferación Celular/genética , Factor de Crecimiento Transformador beta2/genéticaRESUMEN
BACKGROUND: Primary Open-angle Glaucoma (POAG) is a functional disease that.leads to blindness globally. The aims of this study are estimation the importance.of transforming growth factor-beta 2 (TGF-ß2) in the pathogenicity of POAG and.to evaluate the effect of the C/A SNP of the TGF-ß2 gene (rs991967) on POAG development. METHODS: Blood samples and some topographic data were collected from POAG.patients and the controls. The serum level of TGF-ß2 was estimated by ELISA.and the C/A SNP of the TGF-ß2 gene (rs991967) was determined by RFLP-PCR. RESULTS: The males are more susceptible to having POAG (p = 0.0201). The serum.TGF-ß2 is higher in POAG patients as compared with the control (p < 0.0001). The.AA (reference) genotype was the most common in the patients (61.7%). While..CC genotype (45.0%, OR: 0.136, 95%CI: 0.05-0.36, P < 0.0001) and AC..genotypes (41.7%, OR: 0.051, 95%CI: 0.01-0.16, P < 0.001) were most common..in the control group. Moreover, the TGF-ß2 C allele is protective (OR: 0.25,..95%, CI: 0.15-0.44, P < 0.0001). Patients with AA, CC, and AC genotypes have..significantly high levels of TGF-ß2 (P < 0.001) than the control. CONCLUSIONS: The males were more susceptible to acquiring POAG than females,.. especially the elderly. The TGF-ß2 plays important role in the pathogenesis of POAG. The CC and AC genotypes are common in the control and the C allele is a protective factor.
Asunto(s)
Glaucoma de Ángulo Abierto , Masculino , Femenino , Humanos , Anciano , Glaucoma de Ángulo Abierto/genética , Factor de Crecimiento Transformador beta2/genética , Polimorfismo Genético , Genotipo , Polimorfismo de Longitud del Fragmento de Restricción , Polimorfismo de Nucleótido Simple/genéticaRESUMEN
AIMS: Transforming growth factor-beta (TGF-ß) exists in three isoforms TGF-ß1, -ß2, and -ß3. TGF-ß1 has been suggested to be important for maintaining plaque stability, yet the role of TGF-ß2 and -ß3 in atherosclerosis remains to be investigated.This study explores the association of the three isoforms of TGF-ß with plaque stability in the human atherosclerotic disease. METHODS AND RESULTS: TGF-ß1, -ß2, and -ß3 proteins were quantified in 223 human carotid plaques by immunoassays. Indications for the endarterectomy were: symptomatic carotid plaque with stenosis >70% or without symptoms and >80% stenosis. Plaque mRNA levels were assessed by RNA sequencing. Plaque components and extracellular matrix were measured histologically and biochemically. Matrix metalloproteinases and monocyte chemoattractant protein-1 (MCP-1) was measured with immunoassays. The effect of TGF-ß2 on inflammation and protease activity was investigated in vitro using THP-1 and RAW264.7 macrophages. Patients were followed longitudinally for cardiovascular (CV) events.TGF-ß2 was the most abundant isoform and was increased at both protein and mRNA levels in asymptomatic plaques. TGF-ß2 was the main determinant separating asymptomatic plaques in an Orthogonal Projections to Latent Structures Discriminant Analysis. TGF-ß2 correlated positively to features of plaque stability and inversely to markers of plaque vulnerability. TGF-ß2 was the only isoform inversely correlated to the matrix-degrading matrix metalloproteinase-9 and inflammation in the plaque tissue. In vitro, TGF-ß2 pre-treatment reduced MCP-1 gene and protein levels as well as matrix metalloproteinase-9 gene levels and activity. Patients with plaques with high TGF-ß2 levels had a lower risk to suffer from future CV events. CONCLUSIONS: TGF-ß2 is the most abundant TGF-ß isoform in human plaques and may maintain plaque stability by decreasing inflammation and matrix degradation.
Asunto(s)
Enfermedades Cardiovasculares , Placa Aterosclerótica , Humanos , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta1 , Metaloproteinasa 9 de la Matriz/genética , Constricción Patológica , Factor de Crecimiento Transformador beta/metabolismo , Isoformas de Proteínas , ARN Mensajero/genética , ARN Mensajero/metabolismo , Inflamación/genética , Factores de Crecimiento TransformadoresRESUMEN
To assess the contribution of individual TGF-ß isoforms to aortopathy in Marfan syndrome (MFS), we quantified the survival and phenotypes of mice with a combined fibrillin1 (the gene defective in MFS) hypomorphic mutation and a TGF-ß1, 2, or 3 heterozygous null mutation. The loss of TGF-ß2, and only TGF-ß2, resulted in 80% of the double mutant animals dying earlier, by postnatal day 20, than MFS only mice. Death was not from thoracic aortic rupture, as observed in MFS mice, but was associated with hyperplastic aortic valve leaflets, aortic regurgitation, enlarged aortic root, increased heart weight, and impaired lung alveolar septation. Thus, there appears to be a relationship between loss of fibrillin1 and TGF-ß2 in the postnatal development of the heart, aorta and lungs.