RESUMO
Dysregulation of the deubiquitinating protease, UBP43, has been implicated in many human diseases, including cancer. Here, we evaluated the functional significance and mechanism of action of UBP43 in epithelial ovarian cancer. We found that UBP43 was significantly upregulated in the tumor tissues of patients with epithelial ovarian cancer. Similar results were observed in OVCAR-3, Caov-3, TOV-112D, A2780, and SK-OV-3 cells. Furthermore, in vitro functional assays of A2780 and TOV-112D cells demonstrated that UBP43 overexpression promoted cell proliferation, migration, and invasion. Upregulation of UBP43 might result in epithelial-mesenchymal transition by inducing the nuclear transport of ß-catenin, which was accompanied by enhanced N-cadherin but decreased E-cadherin expression. These malignant phenotypes were reversed by UBP43 silencing. Further investigation revealed that the knockdown of UBP43 inhibited cell proliferation by inducing a cell cycle arrest at the G2/M phase. The oncogenic characteristics of UBP43 were validated in a subcutaneous xenograft mouse model. In vivo, tumor growth was delayed in the UBP43-silenced group but accelerated after UBP43 overexpression. Finally, we demonstrated that ß-catenin is a key protein in the UBP43-mediated malignant development of epithelial ovarian cancer. Specifically, overexpression of UBP43 decreased the ubiquitination degradation of ß-catenin and enhanced its protein stability. Also, we observed that the downstream genes of beta-catenin such as cyclin D1, MMP2, and MMP9 were upregulated due to UBP43 overexpression. Thus, we concluded that UBP43 promoted epithelial ovarian cancer tumorigenesis and metastasis through activation of the ß-catenin pathway, suggesting that UBP43 may be a potential therapeutic target for this intractable disease.
Assuntos
Neoplasias Ovarianas , beta Catenina , Animais , Feminino , Humanos , Camundongos , Apoptose , beta Catenina/metabolismo , Carcinogênese/genética , Carcinoma Epitelial do Ovário/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Ovarianas/genética , Via de Sinalização WntRESUMO
BACKGROUND: LncRNA FGF14-AS2 is a critical suppressor in breast cancer (BCa) metastasis. However, whether FGF14-AS2 plays a role in the bone metastasis of BCa remains unknown. METHODS: TRAP assay and intratibial injection were carried out to evaluate the role of FGF14-AS2 in BCa bone metastasis in vitro and in vivo. Polyribosome profiling was done to examine the translation level. RNA pulldown combined with LC/MS was performed to identify the lncRNA-binding partner, RIP, dual-luciferase assay, and Co-IP assays as well to testify these physical interactions. The prognostic value of FGF14-AS2 expression level in BCa patients was analysed using Kaplan-Meier Plotter. RESULTS: We found that FGF14-AS2 suppresses osteoclast differentiation and osteolytic metastasis of BCa. Mechanistically, FGF14-AS2 suppresses the translation of RUNX2 by inhibiting the assembly of eIF4E/eIF4G complex and the phosphorylation of eIF4E, thereby reducing the transcription of RANKL, an essential regulator of osteoclast differentiation. Moreover, FGF14-AS2 is downregulated by YTHDF2-mediated RNA degradation in an m6A-dependent manner. Clinically, patients with high YTHDF2 and low FGF14-AS2 expression levels showed worse distant metastasis-free survival (DMFS). CONCLUSIONS: FGF14-AS2 plays a crucial role in osteolytic metastasis, and may serve as a promising prognostic biomarker and therapeutic target for BCa bone metastasis.
Assuntos
Neoplasias da Mama , RNA Longo não Codificante , Humanos , Feminino , Neoplasias da Mama/genética , RNA Longo não Codificante/genética , Biossíntese de Proteínas , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Proteínas de Ligação a RNA/genéticaRESUMO
BACKGROUND AND PURPOSE: Variants in the glucocerebrosidase (GBA) gene are recognized as a common and important genetic risk factor for Parkinson disease (PD). However, the impact of variant severity on the clinical phenotype of PD in the Chinese population remains unclear. Thus, the present study aimed to determine the frequency of GBA-related PD (GBA-PD) and the relationship of GBA variant severity with clinical characteristics in a large Chinese cohort. METHODS: Long-range polymerase chain reaction and next generation sequencing were performed for the entire GBA gene. GBA variant severity was classified into five classes: mild, severe, risk, complex, and unknown. RESULTS: Among the total 737 PD patients, 47 GBA variants were detected in 79 (10.72%) patients, and the most common GBA variants were R163Q, L444P, and R120W. Complete demographic and clinical data were obtained for 673 patients, which revealed that 18.50% of early onset PD patients had GBA variants. Compared with patients without GBA variants, GBA-PD patients experienced PD onset an average of 4 years earlier and had more severe motor and nonmotor symptoms. Patients carrying severe and complex variants had a higher burden of nonmotor symptoms, especially depression, and more mood/cognitive and gastrointestinal symptoms than patients carrying mild variants. CONCLUSIONS: GBA-PD is highly prevalent in the Chinese population. The severity of GBA variants underlies distinct phenotypic spectrums, with PD patients carrying severe and complex variants seeming to have similar phenotypes. PD patient stratification by GBA variant severity should become a prerequisite for selecting specific treatments.
Assuntos
Glucosilceramidase , Doença de Parkinson , China/epidemiologia , Estudos de Associação Genética , Predisposição Genética para Doença , Glucosilceramidase/genética , Humanos , Mutação/genética , Doença de Parkinson/epidemiologia , Doença de Parkinson/genética , Doença de Parkinson/psicologia , PrevalênciaRESUMO
Osteoporosis is a metabolic disorder characterized by low bone mass and deteriorated microarchitecture, with an increased risk of fracture. Some miRNAs have been confirmed as potential modulators of osteoblast differentiation to maintain bone mass. Our miRNA sequencing results showed that miR-664-3p was significantly down-regulated during the osteogenic differentiation of the preosteoblast MC3T3-E1 cells. However, whether miR-664-3p has an impact on bone homeostasis remains unknown. In this study, we identified overexpression of miR-664-3p inhibited the osteoblast activity and matrix mineralization in vitro. Osteoblastic miR-664-3p transgenic mice exhibited reduced bone mass due to suppressed osteoblast function. Target prediction analysis and experimental validation confirmed Smad4 and Osterix (Osx) are the direct targets of miR-664-3p. Furthermore, specific inhibition of miR-664-3p by subperiosteal injection with miR-664-3p antagomir protected against ovariectomy-induced bone loss. In addition, miR-664-3p expression was markedly higher in the serum from patients with osteoporosis compared to that from normal subjects. Taken together, this study revealed that miR-664-3p suppressed osteogenesis and bone formation via targeting Smad4 and Osx. It also highlights the potential of miR-664-3p as a novel diagnostic and therapeutic target for osteoporotic patients.
Assuntos
Diferenciação Celular , MicroRNAs/genética , Osteoblastos/patologia , Osteogênese , Osteoporose/patologia , Proteína Smad4/antagonistas & inibidores , Fator de Transcrição Sp7/antagonistas & inibidores , Animais , Densidade Óssea , Proliferação de Células , Células Cultivadas , Feminino , Regulação da Expressão Gênica , Humanos , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/patologia , Camundongos , Camundongos Endogâmicos C57BL , Osteoblastos/metabolismo , Osteoporose/etiologia , Osteoporose/metabolismo , Proteína Smad4/genética , Proteína Smad4/metabolismo , Fator de Transcrição Sp7/genética , Fator de Transcrição Sp7/metabolismoRESUMO
Purpose: Human jaw bone marrow mesenchymal stem cells (h-JBMMSCs) are multipotent progenitor cells with osteogenic differentiation potential. MicroRNAs (miRNAs) have emerged as crucial modulators of osteoblast differentiation. In this study, we focus on the role of miR-145 and its target protein in osteoblast differentiation of h-JBMMSCs. Materials and Methods: h-JBMMSCs were isolated and cultured in osteogenic medium. miR-145 mimics and inhibitors were used to elevate and inhibit miR-145 expression, respectively. Osteogenic differentiation was determined by Alkaline phosphatase (ALP) and Alizarin red S (ARS) staining, and osteogenic marker detection using quantitative real-time reverse transcription PCR (qRT-PCR) assay. Bioinformatic analysis and luciferase reporter assay were used to identify the target gene of miR-145. Results: MiR-145 was down-regulated during osteogenesis of h-JBMMSCs. Inhibition of miR-145 promoted osteogenic differentiation of h-JBMMSCs, revealed by enhanced activity of alkaline phosphatase (ALP), greater mineralisation, and increased expression levels of the osteogenic markers, such as Runt-related transcription factor 2 (RUNX2), Osterix (OSX), ALP and COL1A1. MiR-145 could negatively regulate semaphorin3A (SEMA3A), which acts as a positive regulator of osteogenesis. MiR-145 inhibitor induced osteogenesis could be partially attenuated by SEMA3A siRNA treatment in h-JBMMSCs. Conclusions: Our data show that miR-145 directly targets SEMA3A, and also suggest miR-145 as a suppressor, plays an important role in the osteogenic differentiation of h-JBMMSCs.
Assuntos
Células da Medula Óssea/citologia , Diferenciação Celular/genética , Arcada Osseodentária/citologia , Células-Tronco Mesenquimais/citologia , MicroRNAs/metabolismo , Osteogênese/genética , Semaforina-3A/metabolismo , Sequência de Bases , Regulação para Baixo/genética , Células HEK293 , Humanos , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/genéticaRESUMO
Osteoarthritis (OA) is a chronic joint disease and hard to cure at present. Alpha B-crystallin (CRYAB) has been identified as a downregulated gene in OA cartilage. However, the precise roles and underlying molecular mechanisms of CRYAB in OA progression have not been elucidated. In the present study, we found that the expression of CRYAB in cartilages from patients with OA was significantly lower than that in the cartilages from patients with no prior medical history of OA. We established mouse models with OA by destabilization of the medial meniscus (DMM) surgery and found that the expression of CRYAB in OA cartilage was lower than that in the normal cartilages, too. Moreover, we demonstrated that the expression of CRYAB was increased during chondrogenic differentiation and cartilage development. Functional assays revealed that overexpression of CRYAB promoted the proliferation of chondrocytes and inhibited apoptosis, while knockdown of CRYAB presented opposite results. In addition, overexpression of CRYAB upregulated the expression of anabolic markers, Col2a1 and ACAN, and reduced the expression of catabolic markers, MMP13 and ADAMTS5. Conversely, knockdown of CRYAB blocked the expression of the anabolic markers and increased the expression of catabolic markers. Collectively, the results suggest that CRYAB promoted the proliferation and extracellular matrix production of chondrocytes, and inhibited chondrocytes apoptosis and cartilage degradation simultaneously. Thus, CRYAB might be a potential therapeutic target for OA treatment.
Assuntos
Apoptose , Condrócitos/patologia , Matriz Extracelular/metabolismo , Osteoartrite/patologia , Cadeia B de alfa-Cristalina/metabolismo , Idoso , Idoso de 80 Anos ou mais , Animais , Cartilagem Articular/patologia , Proliferação de Células , Células Cultivadas , Condrogênese , Modelos Animais de Doenças , Humanos , Menisco/cirurgia , Camundongos , Pessoa de Meia-Idade , Regulação para CimaRESUMO
OBJECTIVES: Following a specific number of mitotic divisions, primary chondrocytes undergo proliferative senescence, thwarting efforts to expand sufficient populations in vitro suitable to meet the needs of scientific research or medical therapies. Therefore, the human telomerase reverse transcriptase (TERT) was used to immortalize human chondrocyte and establish a cell line that escape from cellular senescence. RESULTS: The human chondrocytes were successfully immortalized by ectopic stable expression of TERT. The established TERT-Chondrocyte cell line showed robust proliferation capacity, even in late passages up to P20, and displayed little cellular senescence. Moreover, TERT-Chondrocyte cells at 20th passage showed similar chondrocyte properties to normal chondrocytes at early passages. CONCLUSIONS: Ectopic stable expression of TERT is an effective way to immortalized human chondrocyte. The immortalized chondrocytes displayed little cellular senescence, showed promise as an in vitro model to investigate osteoarthritis, and may be a promising resource for cell-based therapy for damaged cartilage.
Assuntos
Técnicas de Cultura de Células/métodos , Condrócitos/citologia , Osteoartrite/patologia , Telomerase/genética , Linhagem Celular , Proliferação de Células , Células Cultivadas , Senescência Celular , Condrócitos/metabolismo , Humanos , Telomerase/metabolismo , TransfecçãoRESUMO
As a key transcription factor required for bone formation, osterix (OSX) has been reported to be overexpressed in various cancers, however, its roles in breast cancer progression remain poorly understood. In this study, we demonstrated that OSX was highly expressed in metastatic breast cancer cells. Moreover, it could upregulate the expression of S100 calcium binding protein A4 (S100A4) and potentiate breast cancer cell migration and tumor angiogenesis in vitro and in vivo. Importantly, inhibition of S100A4 impaired OSX-induced cell migration and capillary-like tube formation. Restored S100A4 expression rescued OSX-short hairpin RNA-suppressed cell migration and capillary-like tube formation. Moreover, the expression levels of OSX and S100A4 correlated significantly in human breast tumors. Our study suggested that OSX acts as an oncogenic driver in cell migration and tumor angiogenesis, and may serve as a potential therapeutic target for human breast cancer treatment.
Assuntos
Neoplasias da Mama/genética , Movimento Celular/genética , Neovascularização Patológica/genética , Proteína A4 de Ligação a Cálcio da Família S100/genética , Fator de Transcrição Sp7/genética , Regulação para Cima/genética , Neoplasias da Mama/patologia , Linhagem Celular , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Células Endoteliais da Veia Umbilical Humana , Humanos , Células MCF-7 , Neovascularização Patológica/patologia , RNA Interferente Pequeno/genética , Ativação Transcricional/genéticaRESUMO
Mesenchymal stem cells (MSCs) have been widely studied in the field of regenerative medicine with the potential to solve osteoporosis. Paired box 2 (Pax2), as a transcription factor, is the master regulator of embryogenesis and oncogenesis. However, the function of Pax2 in osteogenesis is unknown. Here, we reported for the first time that the expression of Pax2 gradually increased during osteogenic differentiation of mouse MSCs, and osteoprogenitor cells. However, detected in osteoblastic cells of mouse tibia, the expression of Pax2 in the embryonic stage was higher than that in adulthood. In C3H/10/T1/2 cells and compact bone-derived mouse MSCs (mMSCs), Pax2 knock-down inhibited the proliferation of these cells, down-regulated the expression of osteogenic marker genes, as well as repressed the ALP activity and mineralization. In addition, Pax2 enhanced the transcriptional activity of Runx2, and activated the MAPK pathway genes (ERK, JNK and p38). Furthermore, knock-down of Pax2 repressed the mMSCs-mediated bone regeneration in an ectopic bone formation model. In conclusion, Pax2 promotes osteogenesis of mouse MSCs, suggesting that Pax2 has a role in the pathophysiology of bone related diseases, and has potential application in bone tissue regeneration.
Assuntos
Envelhecimento/genética , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Células-Tronco Mesenquimais/metabolismo , Osteoblastos/metabolismo , Osteogênese/genética , Fator de Transcrição PAX2/genética , Envelhecimento/metabolismo , Fosfatase Alcalina/genética , Fosfatase Alcalina/metabolismo , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Osso e Ossos/citologia , Osso e Ossos/metabolismo , Diferenciação Celular , Coristoma/genética , Coristoma/metabolismo , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Dexametasona/farmacologia , Embrião de Mamíferos , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , MAP Quinase Quinase 4/genética , MAP Quinase Quinase 4/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Nus , Osteoblastos/citologia , Osteoblastos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Fator de Transcrição PAX2/antagonistas & inibidores , Fator de Transcrição PAX2/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Transfecção , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
BACKGROUND/AIMS: Osterix (Osx), a key regulator of osteoblast differentiation and bone formation, has been recently reported to be associated with the progression of breast cancer. However, the precise roles of Osx in breast cancer remain unclear. METHODS: Drug sensitivity of the cancer cells was assessed using an 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay. Target genes were obtained by high-throughput Illumina sequencing and were confirmed in vitro and in vivo. Apoptosis was analysed by Hoechst staining and western blotting. A tissue microarray including 129 samples from breast cancer patients was used for immunohistochemistry (IHC) assays. RESULTS: Overexpression of Osx decreased the chemosensitivity of breast cancer cells, while knockdown of Osx increased the chemosensitivity of breast cancer cells. In particular, we found that the decreased chemosensitivity effect was significantly associated with elevated expression of the polypeptide N-acetylgalactosaminyltransferase 14 (GALNT14). Silencing of GALNT14 in Osx-overexpressed cells restored the decreased chemosensitivity. Conversely, overexpression of GALNT14 in Osx-knockdown cells abrogated the increased chemosensitivity in breast cancer cells. In addition, we revealed that Osx decreased GALNT14-dependent chemosensitivity by enhancing anti-apoptosis. GALNT14 expression exhibited a significant association with breast cancer stages as well as the disease-free survival (DFS) rate. CONCLUSION: Osx plays an important role in the chemosensitivity and inhibition of Osx expression may represent a therapeutic strategy to enhance the chemosensitivity of breast cancer.
Assuntos
Neoplasias da Mama/patologia , N-Acetilgalactosaminiltransferases/metabolismo , Fator de Transcrição Sp7/metabolismo , Animais , Antineoplásicos/uso terapêutico , Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/mortalidade , Linhagem Celular Tumoral , Intervalo Livre de Doença , Feminino , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Nus , N-Acetilgalactosaminiltransferases/antagonistas & inibidores , N-Acetilgalactosaminiltransferases/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Fator de Transcrição Sp7/antagonistas & inibidores , Fator de Transcrição Sp7/genética , Taxa de Sobrevida , Transplante Heterólogo , Proteína X Associada a bcl-2/metabolismoRESUMO
Runx2 and Sp7 transcription factors are essential for skeletogenesis. Targeted deletion of either gene results in failure of osteoblast differentiation and bone formation. Loss of bone-matrix gene expression is surprisingly similar in Sp7 and Runx2 null mice. The molecular mechanisms responsible for similar transcriptional regulation of target genes remain largely unknown. Here, we demonstrate that Runx2 and Sp7 interact physically and functionally. Both proteins are co-expressed in osteoblastic cells. We first characterized a panel of Sp7 antibodies and demonstrate that majority of the published antibodies do not recognize Sp7 protein. Co-immunoprecipitation studies revealed that endogenous Runx2 protein physically interacts with Sp7 protein. We identified that runt homology domain (RHD) of Runx2 protein is involved in physical association with Sp7. Functional consequences of Runx2-Sp7 physical interaction was then assessed by promoter-reporter assays. We selected promoters of osteocalcin (OC), a marker of mature osteoblast and fibroblast growth factor 3 (FGF3), a signaling molecule that determine the fate of embryonic ecto-mesenchyme. Runx2 and Sp7 stimulate OC-promoter activity by 3-folds in epithelial cells. However, when both proteins were co-expressed, a dose-dependent synergistic activation of 22-folds was noted. Similar pattern of synergistic activation of OC-promoter was noted in mesenchymal cell. FGF3 promoter was activated by 25 - and 30-folds with Runx2 and Sp7 respectively. Again a dose-dependent synergistic activation of 130-folds was evident when Runx2 and Sp7 were co-expressed in epithelial cells. Synergistic activation of FGF3 promoter was also noted in mesenchymal cells. Together, our data demonstrated that Runx2-Sp7 molecular complex functionally cooperate for maximal induction of cell-phenotype-restricted genes.
Assuntos
Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Regulação da Expressão Gênica , Osteoblastos/citologia , Osteocalcina/metabolismo , Osteogênese/fisiologia , Fatores de Transcrição/metabolismo , Diferenciação Celular/fisiologia , Linhagem Celular , Regulação da Expressão Gênica/fisiologia , Humanos , Ligação Proteica , Fator de Transcrição Sp7RESUMO
In this study, we demonstrate that infection of HSB-2 cells with human herpesvirus 6 (HHV-6) resulted in the accumulation of infected cells in the G2/M phase of the cell cycle. Analysis of various cell-cycle-regulatory proteins indicated that the levels of cyclins A2, B1, and E1 were increased in HHV-6-infected cells, but there was no difference in cyclin D1 levels between mock-infected and HHV-6-infected cells. Our data also showed that inducing G2/M phase arrest in cells infected by HHV-6 provided favorable conditions for viral replication.
Assuntos
Ciclo Celular , Herpesvirus Humano 6/fisiologia , Interações Hospedeiro-Patógeno , Linfócitos T/fisiologia , Linfócitos T/virologia , Replicação Viral , Proteínas de Ciclo Celular/análise , Linhagem Celular , HumanosRESUMO
Mono-methylation of histone H3 on Lys 4 (H3K4me1), which is catalyzed by histone-lysine N-methyltransferase 2D (KMT2D), serves as an important epigenetic regulator in transcriptional control. In this study, the authors identify early B-cell factor 2 (EBF2) as a binding protein of H3K4me1. Combining analyses of RNA-seq and ChIP-seq data, the authors further identify killin (KLLN) as a transcriptional target of KMT2D and EBF2 in pancreatic ductal adenocarcinoma (PDAC) cells. KMT2D-dependent H3K4me1 and EBF2 are predominantly over-lapped proximal to the transcription start site (TSS) of KLLN gene. Comprehensive functional assays show that KMT2D and EBF2 cooperatively inhibit PDAC cells proliferation, migration, and invasion through upregulating KLLN. Such inhibition on PDAC progression is also achieved through increasing H3K4me1 level by GSK-LSD1, a selective inhibitor of lysine-specific demethylase 1 (LSD1). Taken together, these findings reveal a new mechanism underlying PDAC progression and provide potential therapeutic targets for PDAC treatment.
Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Carcinoma Ductal Pancreático/genética , Regulação da Expressão Gênica , Histona Desmetilases/genética , Histonas/genética , Neoplasias Pancreáticas/genéticaRESUMO
BACKGROUND: Lysine methyltransferase 2D (KMT2D) mediates mono-methylation of histone H3 lysine 4 (H3K4me1) in mammals. H3K4me1 mark is involved in establishing an active chromatin structure to promote gene transcription. However, the precise molecular mechanism underlying the KMT2D-mediated H3K4me1 mark modulates gene expression in triple-negative breast cancer (TNBC) progression is unresolved. METHODS AND RESULTS: We recognized Y-box-binding protein 1 (YBX1) as a "reader" of the H3K4me1 mark, and a point mutation of YBX1 (E121A) disrupted this interaction. We found that KMT2D and YBX1 cooperatively promoted cell growth and metastasis of TNBC cells in vitro and in vivo. The expression levels of KMT2D and YBX1 were both upregulated in tumour tissues and correlated with poor prognosis for breast cancer patients. Combined analyses of ChIP-seq and RNA-seq data indicated that YBX1 was co-localized with KMT2D-mediated H3K4me1 in the promoter regions of c-Myc and SENP1, thereby activating their expressions in TNBC cells. Moreover, we demonstrated that YBX1 activated the expressions of c-Myc and SENP1 in a KMT2D-dependent manner. CONCLUSION: Our results suggest that KMT2D-mediated H3K4me1 recruits YBX1 to facilitate TNBC progression through epigenetic activation of c-Myc and SENP1. These results together unveil a crucial interplay between histone mark and gene regulation in TNBC progression, thus providing novel insights into targeting the KMT2D-H3K4me1-YBX1 axis for TNBC treatment. HIGHLIGHTS: YBX1 is a KMT2D-mediated H3K4me1-binding effector protein and mutation of YBX1 (E121A) disrupts its binding to H3K4me1. KMT2D and YBX1 cooperatively promote TNBC proliferation and metastasis by activating c-Myc and SENP1 expression in vitro and in vivo. YBX1 is colocalized with H3K4me1 in the c-Myc and SENP1 promoter regions in TNBC cells and increased YBX1 expression predicts a poor prognosis in breast cancer patients.
Assuntos
Epigênese Genética , Neoplasias de Mama Triplo Negativas , Proteína 1 de Ligação a Y-Box , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Neoplasias de Mama Triplo Negativas/metabolismo , Humanos , Proteína 1 de Ligação a Y-Box/metabolismo , Proteína 1 de Ligação a Y-Box/genética , Feminino , Epigênese Genética/genética , Animais , Progressão da Doença , Camundongos , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Histonas/metabolismo , Histonas/genética , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Lisina/análogos & derivadosRESUMO
Targeted drug delivery and the reduction of off-target effects are crucial for the promising clinical application of nucleic acid drugs. To address this challenge, a new approach for treating osteoarthritis (OA) that accurately delivers antisense oligonucleotides (ASO) targeting matrix metalloproteinase-13 (ASO-MMP13) to chondrocytes, is developed. Small extracellular vesicles (exos) are ligated with chondrocyte affinity peptide (CAP) using Sortase A and subsequently incubated with cholesterol-modified ASO-MMP13 to construct a chondrocyte-targeted drug delivery exo (CAP-exoASO). Compared with exos without CAP (ExoASO), CAP-exoASOs attenuate IL-1ß-induced chondrocyte damage and prolong the retention time of ASO-MMP13 in the joint without distribution in major organs following intra-articular injection. Notably, CAP-exoASOs decrease MMP13 expression (P < 0.001) and upregulate COL2A1 expression (P = 0.006), resulting in reorganization of the cartilage matrix and alleviation of progression in the OA model. Furthermore, the Osteoarthritis Research Society International (OARSI) score of articular cartilage tissues treated with CAP-exoASO is comparable with that of healthy rats (P = 0.148). A mechanistic study demonstrates that CAP-exoASO may reduce inflammation by suppressing the IL-17 and TNF signaling pathways. Based on the targeted delivery effect, CAP-exoASOs successfully accomplish cartilage repair and have considerable potential for development as a promising therapeutic modality for satisfactory OA therapy.
Assuntos
Aminoaciltransferases , Proteínas de Bactérias , Condrócitos , Cisteína Endopeptidases , Vesículas Extracelulares , Metaloproteinase 13 da Matriz , Osteoartrite , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 13 da Matriz/genética , Animais , Osteoartrite/terapia , Osteoartrite/metabolismo , Osteoartrite/patologia , Osteoartrite/genética , Condrócitos/metabolismo , Ratos , Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/química , Aminoaciltransferases/metabolismo , Aminoaciltransferases/genética , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Cisteína Endopeptidases/metabolismo , Cisteína Endopeptidases/genética , Masculino , Sistemas de Liberação de Medicamentos/métodos , Ratos Sprague-Dawley , Oligonucleotídeos Antissenso/farmacologia , Oligonucleotídeos Antissenso/química , Oligonucleotídeos Antissenso/administração & dosagem , Cartilagem Articular/metabolismo , Cartilagem Articular/patologiaRESUMO
BACKGROUND: The aim of this study was to investigate the effect of isopsoralen on osteogenic differentiation of human jawbone marrow mesenchymal cells and its possible mechanism. METHOD: The cytotoxicity and proliferation of cells were measured by a cell counting kit 8. Alkaline phosphatase activity analysis was then used to determine the optimal concentration of isopsoralen to promote the differentiation. Western blot, qRT-PCR and Alizarin Red S staining were used to evaluate the role of Notch signaling pathway in isopsoralen-induced osteogenic differentiation. This study also investigated the anti-osteoporotic effects of ISO using in vivo osteoporosis models. RESULTS: Our results showed that 1 × 10-6 mol / L isopsoralen can effectively promote the proliferation and osteogenic differentiation of cells. Moreover, we found that activation of notch signaling pathway inhibited isopsoralen-induced osteogenesis and inhibition of Notch signal promoted the differentiation of osteoblasts induced by isopsoralen. In vivo experiments revealed that ISO significantly inhibited OVX-induced bone mineral density loss and restored the impaired bone structural properties in osteoporosis model mice. CONCLUSION: Our findings demonstrated that isopsoralen induced osteogenic differentiation by inhibiting Notch signaling and it might be a potential therapeutic agent for treating or preventing osteoporosis.
Assuntos
Células-Tronco Mesenquimais , Osteoporose , Humanos , Camundongos , Animais , Osteogênese , Medula Óssea/metabolismo , Células Cultivadas , Diferenciação Celular , Transdução de Sinais , Osteoporose/tratamento farmacológico , Células da Medula Óssea/metabolismoRESUMO
Osteoporosis is a complicated multifactorial disorder characterized by low bone mass and deteriorated bone microarchitecture with an elevated fracture risk. MicroRNAs play important roles in osteoblastic differentiation. In the present study, we found that miR-224-5p was markedly downregulated during the osteogenic differentiation of C2C12 cells. Overexpression of miR-224-5p in C2C12 cells inhibited osteoblast activity, as indicated by reduced ALP activity, matrix mineralization and the expression of osteogenic marker genes. Moreover, we demonstrated that Runx2 and Sp7 were direct targets of miR-224-5p. Furthermore, the specific inhibition of miR-224-5p by femoral bone marrow cavity injection with miR-224-5p antagomir prevented ovariectomy-induced bone loss. Finally, we found that the levels of miR-224-5p were markedly elevated in the sera of patients with osteoporosis. Collectively, this study revealed that miR-224-5p negatively regulates osteogenic differentiation by targeting Runx2 and Sp7. It also highlights the potential use of miR-224-5p as a therapeutic target and diagnostic biomarker for osteoporosis. Supplementary information: The online version contains supplementary material available at 10.1007/s10616-023-00593-z.
RESUMO
Objective: Variants in the glucocerebrosidase (GBA) gene are the most common and significant risk factor for Parkinson's disease (PD). However, the impact of GBA variants on PD disease progression in the Chinese population remains unclear. This study aimed to explore the significance of GBA status on motor and cognitive impairment in a longitudinal cohort of Chinese patients with PD. Methods: The entire GBA gene was screened by long-range polymerase chain reaction (LR-PCR) and next generation sequencing (NGS). A total of 43 GBA-related PD (GBA-PD) and 246 non-GBA-mutated PD (NM-PD) patients with complete clinical data at baseline and at least one follow-up were recruited for this study. The associations of GBA genotype with rate of motor and cognitive decline, as measured by Unified PD Rating Scale (UPDRS) motor and Montreal Cognitive Assessment (MoCA), were assessed by linear mixed-effect models. Results: The estimated (standard error, SE) UPDRS motor [2.25 (0.38) points/year] and MoCA [-0.53 (0.11) points/year] progression rates in the GBA-PD group were significantly faster than those in the NM-PD group [1.35 (0.19); -0.29 (0.04) points/year; respectively]. In addition, the GBA-PD group showed significantly faster estimated (SE) bradykinesia [1.04 (0.18) points/year], axial impairment [0.38 (0.07) points/year], and visuospatial/executive [-0.15 (0.03) points/year] progression rates than the NM-PD group [0.62 (0.10); 0.17 (0.04); -0.07 (0.01) points/year; respectively]. Conclusion: GBA-PD is associated with faster motor and cognitive decline, specifically greater disability in terms of bradykinesia, axial impairment, and visuospatial/executive function. Better understanding of GBA-PD progression may help predict prognosis and improve clinical trial design.
RESUMO
Osteoarthritis (OA) is the most common degenerative disorder, affecting approximately half of the elderly population. In this study, we find that the expressions of long noncoding RNA (lncRNA) IGFBP7-OT and its maternal gene, IGFBP7, are upregulated and positively correlated in osteoarthritic cartilage. Overexpression of IGFBP7-OT significantly inhibits chondrocyte viability, promotes chondrocyte apoptosis, and reduces extracellular matrix components, whereas IGFBP7-OT knockdown has the opposite effects. IGFBP7-OT overexpression promotes cartilage degeneration and markedly aggravates the monosodium iodoacetate-induced OA phenotype in vivo. Further mechanistic research reveals that IGFBP7-OT promotes OA progression by upregulating IGFBP7 expression. Specifically, IGFBP7-OT suppresses the occupancy of DNMT1 and DNMT3a on the IGFBP7 promoter, thereby inhibiting methylation of the IGFBP7 promoter. The upregulation of IGFBP7-OT in OA is partially controlled by METTL3-mediated N6-methyladenosine (m6A) modification. Collectively, our findings reveal that m6A modification of IGFBP7-OT promotes OA progression by regulating the DNMT1/DNMT3a-IGFBP7 axis and provide a potential therapeutical target for OA treatment.
Assuntos
DNA Metiltransferase 3A , Metilases de Modificação do DNA , Osteoartrite , RNA Longo não Codificante , Idoso , Humanos , Apoptose , Cartilagem/metabolismo , Condrócitos , Metilases de Modificação do DNA/metabolismo , Metiltransferases/metabolismo , Osteoartrite/genética , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Regulação para Cima/genética , DNA Metiltransferase 3A/metabolismo , Animais , CamundongosRESUMO
HS3ST3B1-IT1 was identified as a downregulated long noncoding RNA in osteoarthritic cartilage. However, its roles and mechanisms in the pathogenesis of osteoarthritis (OA) are unclear. In this study, we demonstrated that the expressions of HS3ST3B1-IT1 and its maternal gene HS3ST3B1 were downregulated and positively correlated in osteoarthritic cartilage. Overexpression of HS3ST3B1-IT1 significantly increased chondrocyte viability, inhibited chondrocyte apoptosis, and upregulated extracellular matrix (ECM) proteins, whereas HS3ST3B1-IT1 knockdown had the opposite effects. In addition, HS3ST3B1-IT1 significantly ameliorated monosodium-iodoacetate-induced OA in vivo. Mechanistically, HS3ST3B1-IT1 upregulated HS3ST3B1 expression by blocking its ubiquitination-mediated degradation. Knockdown of HS3ST3B1 reversed the effects of HS3ST3B1-IT1 on chondrocyte viability, apoptosis, and ECM metabolism. AlkB homolog 5 (ALKBH5)-mediated N6-methyladenosine (m6A) demethylation stabilized HS3ST3B1-IT1 RNA. Together, our data revealed that ALKBH5-mediated upregulation of HS3ST3B1-IT1 suppressed OA progression by elevating HS3ST3B1 expression, suggesting that HS3ST3B1-IT1/HS3ST3B1 may serve as potential therapeutic targets for OA treatment.