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1.
Anticancer Res ; 40(2): 733-741, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32014915

RESUMO

BACKGROUND/AIM: GPR87 is a member of the cell surface molecular G protein-coupled receptors (GPCR) family and suggested to contribute to the viability of human tumor cells. Its tumor-specific expression and cell surface location make it a potential molecule for targeted therapy. In the present study, we aimed to examine the effect of silencing GPR87 expression and explore the possibility of establishing gene therapy against GPR87-overexpressing lung cancer. MATERIALS AND METHODS: Twenty malignant cell lines were investigated and GPR87-overexpressing H358 and PC9 lung cancer cells were subjected to inhibiting experiments. A short hairpin siRNA targeting the GPR87 gene was transformed into an adenoviral vector (Ad-shGPR87). Real-time RT-PCR and western blot analyses were performed to evaluate gene and protein expression. Tumors derived from human H358 cells were subcutaneously implanted in nude mice for in vivo experiments. RESULTS AND CONCLUSION: About 50% (10/20) malignant cells showed GPR87-overexpression, especially for lung cancer cells (70%, 7/10). Ad-shGPR87 effectively down-regulated the GPR87 expression, and significantly inhibited the cell proliferation in GPR87-overexpressing H358 and PC9 cells. Treatment with Ad-shGPR87 exerted a significant antitumor effect against the GPR87-expressing H358 xenografts. In addition, the gene expression of H3.3, a recently proved activator for GPR87 transcription, was positively correlated with GPR87 gene expression. Furthermore, a significant decrease of KRAS and c-Myc expression was observed in both cell lines after Ad-shGPR87 infection. In conclusion, GPR87 may play a critical role in cancer cell proliferation, and indicate its potential as a novel target for lung cancer treatment.


Assuntos
Terapia Genética/métodos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/terapia , RNA Interferente Pequeno/administração & dosagem , Receptores de Ácidos Lisofosfatídicos/antagonistas & inibidores , Adenoviridae/genética , Animais , Linhagem Celular Tumoral , Proliferação de Células/genética , Expressão Gênica , Vetores Genéticos/administração & dosagem , Vetores Genéticos/genética , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Nus , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , RNA Interferente Pequeno/genética , Receptores de Ácidos Lisofosfatídicos/biossíntese , Receptores de Ácidos Lisofosfatídicos/genética , Transdução de Sinais , Ensaios Antitumorais Modelo de Xenoenxerto
2.
Genes (Basel) ; 10(9)2019 09 11.
Artigo em Inglês | MEDLINE | ID: mdl-31514484

RESUMO

Existing methods often fail to recognize the conversions for the biological roles of the pairs of genes and microRNAs (miRNAs) between the tumor and normal samples. We have developed a novel cluster scoring method to identify messenger RNA (mRNA) and miRNA interaction pairs and clusters while considering tumor and normal samples jointly. Our method has identified 54 significant clusters for 15 cancer types selected from The Cancer Genome Atlas project. We also determined the shared clusters across tumor types and/or subtypes. In addition, we compared gene and miRNA overlap between lists identified in our liver hepatocellular carcinoma (LIHC) study and regulatory relationships reported from human and rat nonalcoholic fatty liver disease studies (NAFLD). Finally, we analyzed biological functions for the single significant cluster in LIHC and uncovered a significantly enriched pathway (phospholipase D signaling pathway) with six genes represented in the cluster, symbols: DGKQ, LPAR2, PDGFRB, PIK3R3, PTGFR and RAPGEF3.


Assuntos
Regulação Neoplásica da Expressão Gênica , Redes Reguladoras de Genes , MicroRNAs/genética , RNA Mensageiro/genética , Algoritmos , Carcinoma Hepatocelular/genética , Genoma Humano , Genômica/métodos , Fatores de Troca do Nucleotídeo Guanina/genética , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Humanos , Neoplasias Hepáticas/genética , MicroRNAs/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , RNA Mensageiro/metabolismo , Receptor beta de Fator de Crescimento Derivado de Plaquetas/genética , Receptor beta de Fator de Crescimento Derivado de Plaquetas/metabolismo , Receptores de Ácidos Lisofosfatídicos/genética , Receptores de Ácidos Lisofosfatídicos/metabolismo , Receptores de Prostaglandina/genética , Receptores de Prostaglandina/metabolismo
3.
Med Sci Monit ; 25: 4705-4715, 2019 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-31235682

RESUMO

BACKGROUND The aim of this study was to evaluate lysophosphatidic acid receptor-2 (LPA2) and Krüppel-like factor 5 (KLF5) protein expression in gastric adenocarcinoma and their correlation with patient clinicopathological characteristics and prognosis. MATERIAL AND METHODS Fifty-one gastric adenocarcinoma tissue samples, 21 gastric intraepithelial neoplasia (GIN) samples, and 13 normal gastric tissue samples were collected to test for LPA2 and KLF5 expression by tissue microarray and immunohistochemistry assay. LPA2 and KLF5 positive expression rate between gastric adenocarcinoma, GIN, and normal gastric tissue were compared. The relationship between LPA2 expression, KLF5 expression, and patients' clinicopathological characteristics and prognosis were evaluated. RESULTS The positive expression rate of LPA2 and KLF5 were statistical different in gastric adenocarcinoma, GIN, and normal gastric tissue (P<0.05). LPA2 positive expression was associated with tumor invasion depth, Lauren type, vascular invasion, local lymph node metastasis, and clinical stage (P<0.05). There was no correlation between LPA2 expression (hazard ratio [HR]=1.84, 95% confidence interval [CI]: 0.89-3.80, P>0.05), KLF5 expression (HR=1.13, 95% CI: 0.53-2.36, P>0.05), and gastric cancer patients' overall survival. CONCLUSIONS LPA2 and KLF5 protein expressions were differently expressed in gastric adenocarcinoma, GIN, and normal gastric tissue, and differences were correlated with patients' clinical characteristic. However, LPA2 and KLF5 expressions were not correlated with the patients' prognosis.


Assuntos
Fatores de Transcrição Kruppel-Like/metabolismo , Receptores de Ácidos Lisofosfatídicos/metabolismo , Neoplasias Gástricas/metabolismo , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Adulto , Idoso , Feminino , Humanos , Imuno-Histoquímica , Fatores de Transcrição Kruppel-Like/biossíntese , Fatores de Transcrição Kruppel-Like/genética , Masculino , Pessoa de Meia-Idade , Prognóstico , Modelos de Riscos Proporcionais , Receptores de Ácidos Lisofosfatídicos/biossíntese , Receptores de Ácidos Lisofosfatídicos/genética , Neoplasias Gástricas/patologia
4.
Biomed Res Int ; 2019: 3065818, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31236404

RESUMO

Hepatocellular carcinoma (HCC) is a primary cause of cancer-related death in the world. Despite the fact that there are many methods to treat HCC, the 5-year survival rate of HCC is still at a low level. Emodin can inhibit the growth of HCC cells in vitro and in vivo. However, the gene regulation of emodin in HCC has not been well studied. In our research, RNA sequencing technology was used to identify the differentially expressed genes (DEGs) in HepG2 cells induced by emodin. A total of 859 DEGs were identified, including 712 downregulated genes and 147 upregulated genes in HepG2 cells treated with emodin. We used DAVID for function and pathway enrichment analysis. The protein-protein interaction (PPI) network was constructed using STRING, and Cytoscape was used for module analysis. The enriched functions and pathways of the DEGs include positive regulation of apoptotic process, structural molecule activity and lipopolysaccharide binding, protein digestion and absorption, ECM-receptor interaction, complement and coagulation cascades, and MAPK signaling pathway. 25 hub genes were identified and pathway analysis revealed that these genes were mainly enriched in neuropeptide signaling pathway, inflammatory response, and positive regulation of cytosolic calcium ion concentration. Survival analysis showed that LPAR6, C5, SSTR5, GPR68, and P2RY4 may be involved in the molecular mechanisms of emodin therapy for HCC. A quantitative real-time PCR (qRT-PCR) assay showed that the mRNA levels of LPAR6, C5, SSTR5, GPR68, and P2RY4 were significantly decreased in HepG2 cells treated with emodin. In conclusion, the identified DEGs and hub genes in the present study provide new clues for further researches on the molecular mechanisms of emodin.


Assuntos
Carcinoma Hepatocelular/tratamento farmacológico , Emodina/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Transcriptoma/genética , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Biologia Computacional , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Mapeamento de Interação de Proteínas , Receptores Acoplados a Proteínas-G/genética , Receptores de Ácidos Lisofosfatídicos/genética , Receptores Purinérgicos P2/genética , Receptores de Somatostatina/genética , Transdução de Sinais/efeitos dos fármacos , Software
5.
Int J Dermatol ; 58(8): 946-952, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31077348

RESUMO

BACKGROUND: Autosomal recessive wooly hair/hypotrichosis is an inherited disorder of hair characterized by less dense, short, and tightly curled hair on the scalp and sometimes less dense to complete absence of eyebrows and eyelashes. Autosomal recessive wooly hair/hypotrichosis phenotypes are mostly associated with pathogenic sequence variants in LIPH and LPAR6 genes. METHODS: To find out the molecular basis of the disease, five families with autosomal recessive wooly hair/hypotrichosis were recruited for genetic analysis. Direct Sanger sequencing of LIPH and LPAR6 genes was carried out using BigDye chain termination chemistry. P2RY5 protein homology models were developed to study the effect of mutation on protein structure in a family having novel mutation. RESULTS: Sanger sequencing revealed a novel homozygous missense mutation (c.47A>T) in the LPAR6 gene in family A, while recurrent mutation (c.436G>A) was detected in the rest of the four families (B-E). Protein homology models for both native and mutant P2RY5 protein were developed to study the difference in subtle structural features because of Lys16Met (K16M) mutation. We observed that P2RY5K16M mutation results decrease in the number of ionic interactions detrimental to the protein stability. Protein modeling studies revealed that the novel mutation identified here decreased the number of ionic interactions by affecting physicochemical parameters of the protein, leading to an overall decrease in protein stability with no major secondary structural changes. CONCLUSION: The molecular analysis further confirms the frequent involvement of LPAR6 in autosomal recessive wooly hair/hypotrichosis, while the bioinformatic study revealed that the missense mutation destabilizes the overall structure of P2RY5 protein.


Assuntos
Genes Recessivos/genética , Doenças do Cabelo/genética , Cabelo/anormalidades , Hipotricose/genética , Receptores de Ácidos Lisofosfatídicos/genética , Biologia Computacional , Consanguinidade , Feminino , Humanos , Masculino , Mutação de Sentido Incorreto , Paquistão , Linhagem , Fenótipo , Estrutura Secundária de Proteína/genética , Receptores de Ácidos Lisofosfatídicos/química , Receptores Purinérgicos P2/química , Receptores Purinérgicos P2/genética , Homologia de Sequência de Aminoácidos
6.
Nat Neurosci ; 22(5): 809-819, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30988526

RESUMO

Microscopic features (that is, microstructure) of axons affect neural circuit activity through characteristics such as conduction speed. To what extent axonal microstructure in white matter relates to functional connectivity (synchrony) between brain regions is largely unknown. Using MRI data in 11,354 subjects, we constructed multivariate models that predict functional connectivity of pairs of brain regions from the microstructural signature of white matter pathways that connect them. Microstructure-derived models provided predictions of functional connectivity that explained 3.5% of cross-subject variance on average (ranging from 1-13%, or r = 0.1-0.36) and reached statistical significance in 90% of the brain regions considered. The microstructure-function relationships were associated with genetic variants, co-located with genes DAAM1 and LPAR1, that have previously been linked to neural development. Our results demonstrate that variation in white matter microstructure predicts a fraction of functional connectivity across individuals, and that this relationship is underpinned by genetic variability in certain brain areas.


Assuntos
Encéfalo/anatomia & histologia , Encéfalo/fisiologia , Fenótipo , Substância Branca/anatomia & histologia , Substância Branca/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Idoso , Encéfalo/crescimento & desenvolvimento , Mapeamento Encefálico , Feminino , Estudo de Associação Genômica Ampla , Humanos , Imagem por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Modelos Neurológicos , Análise Multivariada , Vias Neurais/anatomia & histologia , Vias Neurais/fisiologia , Receptores de Ácidos Lisofosfatídicos/genética
7.
Dev Dyn ; 248(5): 375-395, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30847983

RESUMO

BACKGROUND: LPA is a small bioactive phospholipid that acts as an extracellular signaling molecule and is involved in cellular processes, including cell proliferation, migration, and differentiation. LPA acts by binding and activating at least six known G protein-coupled receptors: LPA1-6 . In recent years, LPA has been suggested to play an important role both in normal neuronal development and under pathological conditions in the nervous system. RESULTS: We show the expression pattern of LPA receptors during mouse brain development by using qRT-PCR, in situ hybridization, and immunocytochemistry. Only LPA 1 , LPA 2, LPA 4, and LPA 6 mRNA transcripts were detected throughout development stages from embryonic day 16 until postnatal day 30 of hippocampus, neocortex, cerebellum, and bulbus olfactorius in our experiments, while expression of LPA 3 and LPA 5 genes was below detection level. In addition to our qRT-PCR results, we also analyzed the cellular protein expression of endogenous LPA receptors, with focus on LPA1 and LPA2 within postnatal brain slices and primary neuron differentiation with and without cytoskeleton stabilization and destabilization. CONCLUSIONS: The expression of LPA receptors changes depends on the developmental stage in mouse brain and in cultured hippocampal primary neurons. Interestingly, we found that commercially available antibodies for LPA receptors are largely unspecific.


Assuntos
Encéfalo/crescimento & desenvolvimento , Receptores de Ácidos Lisofosfatídicos/metabolismo , Animais , Encéfalo/metabolismo , Células Cultivadas , Hipocampo/citologia , Camundongos , Neurônios/citologia , RNA Mensageiro/análise , Receptores de Ácidos Lisofosfatídicos/genética
8.
Biochim Biophys Acta Mol Basis Dis ; 1865(6): 1332-1340, 2019 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-30763641

RESUMO

Diabetic nephropathy (DN) is one of the major long-term complications of diabetes. Lysophosphatidic acid (LPA) signaling has been implicated in renal fibrosis. In our previous study, we found that the LPA receptor 1/3 (LPAR1/3) antagonist, ki16425, protected against DN in diabetic db/db mice. Here, we investigated the effects of a specific pharmacological inhibitor of LPA receptor 1 (LPA1), AM095, on DN in streptozotocin (STZ)-induced diabetic mice to exclude a possible contribution of LPAR3 inhibition. AM095 treatment significantly reduced albuminuria and the albumin to creatinine ratio and significantly decreased the glomerular volume and tuft area in the treated group compared with the STZ-vehicle group. In the kidney of STZ-induced diabetic mice, the expression of LPAR1 mRNA and protein was positively correlated with oxidative stress. AM095 treatment inhibited LPA-induced reactive oxygen species production and NADPH oxidase expression as well as LPA-induced toll like receptor 4 (TLR4) expression in mesangial cells and in the kidney of STZ-induced diabetic mice. In addition, AM095 treatment suppressed LPA-induced pro-inflammatory cytokines and fibrotic factors expression through downregulation of phosphorylated NFκBp65 and c-Jun N-terminal kinases (JNK) in vitro and in the kidney of STZ-induced diabetic mice. Pharmacological or siRNA inhibition of TLR4 and NADPH oxidase mimicked the effects of AM095 in vitro. In conclusion, AM095 is effective in preventing the pathogenesis of DN by inhibiting TLR4/NF-κB and the NADPH oxidase system, consequently inhibiting the inflammatory signaling cascade in renal tissue of diabetic mice, suggesting that LPAR1 antagonism might provide a potential therapeutic target for DN.


Assuntos
Antioxidantes/farmacologia , Nefropatias Diabéticas/tratamento farmacológico , Hipoglicemiantes/farmacologia , Fenilacetatos/farmacologia , Receptores de Ácidos Lisofosfatídicos/genética , Receptor 4 Toll-Like/genética , Fator de Transcrição RelA/genética , Albuminúria/induzido quimicamente , Albuminúria/tratamento farmacológico , Albuminúria/genética , Albuminúria/metabolismo , Animais , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Nefropatias Diabéticas/induzido quimicamente , Nefropatias Diabéticas/genética , Nefropatias Diabéticas/metabolismo , Regulação da Expressão Gênica , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Glomérulos Renais/efeitos dos fármacos , Glomérulos Renais/metabolismo , Glomérulos Renais/patologia , Lisofosfolipídeos/antagonistas & inibidores , Lisofosfolipídeos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NADPH Oxidases/antagonistas & inibidores , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Estresse Oxidativo/efeitos dos fármacos , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Receptores de Ácidos Lisofosfatídicos/antagonistas & inibidores , Receptores de Ácidos Lisofosfatídicos/metabolismo , Transdução de Sinais , Estreptozocina , Receptor 4 Toll-Like/antagonistas & inibidores , Receptor 4 Toll-Like/metabolismo , Fator de Transcrição RelA/antagonistas & inibidores , Fator de Transcrição RelA/metabolismo
9.
Glia ; 67(5): 999-1012, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30637823

RESUMO

Pruritus is a common and disabling symptom in patients with hepatobiliary disorders, particularly in those with cholestatic features. Serum levels of lysophosphatidic acid (LPA) and its forming enzyme autotaxin were increased in patients suffering from hepatic pruritus, correlated with itch severity and response to treatment. Here we show that in a culture of dorsal root ganglia LPA 18:1 surprisingly activated a large fraction of satellite glia cells, and responses to LPA 18:1 correlated inversely with responses to neuronal expressed transient receptor potential channels. LPA 18:1 caused only a marginal activation of heterologously expressed TRPV1, and responses in dorsal root ganglion cultures from TRPV1-deficient mice were similar to controls. LPA 18:1 desensitized subsequent responsiveness to chloroquine and TGR5 agonist INT-777. The LPA 18:1-induced increase in cytoplasmatic calcium stems from the endoplasmatic reticulum. LPA receptor expression in dorsal root ganglia and Schwann cells, LPAR1 immunohistochemistry, and pharmacological results indicate a signaling pathway through LPA receptor 1. Peripheral rat Schwann cells, which are of glial lineage as the satellite glia cells, were also responsive to LPA 18:1. Summarizing, LPA 18:1 primarily activates rather glial cells than neurons, which may subsequently modulate neuronal responsiveness and sensory sensations such as itch and pain.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Lisofosfolipídeos/farmacologia , Neuroglia/efeitos dos fármacos , Células Satélites Perineuronais/efeitos dos fármacos , Células de Schwann/efeitos dos fármacos , Animais , Cálcio/metabolismo , Células Cultivadas , Feminino , Gânglios Espinais/citologia , Regulação da Expressão Gênica/genética , Células HEK293 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , RNA Mensageiro/metabolismo , Receptores de Ácidos Lisofosfatídicos/genética , Receptores de Ácidos Lisofosfatídicos/metabolismo , Nervo Isquiático/citologia , Células Receptoras Sensoriais/efeitos dos fármacos , Células Receptoras Sensoriais/metabolismo , Canal de Cátion TRPA1/deficiência , Canal de Cátion TRPA1/genética , Canais de Cátion TRPV/deficiência , Canais de Cátion TRPV/genética
10.
Mol Diagn Ther ; 23(1): 127-138, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30694446

RESUMO

BACKGROUND AND OBJECTIVE: Lysophosphatidic acid (LPA) has widely been reported to participate in the numerous biological behaviors of tumors through its receptors. LPA receptor 6 (LPAR6) is a newly identified G protein-coupled receptor of LPA, and few studies have explored the role of LPAR6 in cancer. In breast cancer (BC), LPAR6 has not, as yet, been studied. This study aimed to evaluate LPAR6 expression in BC patients and to explore its possible role in BC. METHODS: A total of 98 pairs of clinical BC and para-cancer tissues were collected, and LPAR6 expression was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). Kaplan-Meier plots were employed for survival analysis. Human BC cell lines were cultured to study decitabine (5-aza-2'-deoxycytidine [5-Aza]) intervention. Bioinformatic analyses were carried out to support the study conclusions and predictions. RESULTS: LPAR6 expression was significantly reduced in BC tissues (p < 0.001). In the analysis of clinical parameters, LPAR6 expression was related to BC molecular classification (p < 0.05). Furthermore, patients with higher LPAR6 expression had better prognoses (p < 0.001). The CpG islands of LPAR6 were hypermethylated in BC tissues relative to those in para-cancer tissues (p < 0.01). 5-Aza significantly upregulated LPAR6 expression in BC cell lines. Additionally, LPAR6 knockdown significantly promoted cell migration and proliferation in the ZR-75-1 cell line (p < 0.001). Finally, through Gene Set Enrichment Analysis (GSEA), LPAR6 was found to be negatively correlated with cancer-promoting factors and positively correlated with tumor-suppressing factors. CONCLUSION: LPAR6 was downregulated in BC, and low LPAR6 expression was related to poor prognosis. The anti-tumor drug 5-Aza significantly upregulated LPAR6 expression in vitro, and LPAR6 might act as a tumor suppressor in BC.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Proliferação de Células/genética , Prognóstico , Receptores de Ácidos Lisofosfatídicos/genética , Adulto , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Decitabina/administração & dosagem , Intervalo Livre de Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Pessoa de Meia-Idade , Estudos Prospectivos , Transdução de Sinais
11.
Mol Cells ; 42(2): 123-134, 2019 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-30622227

RESUMO

Lysophosphatidic acid (LPA) is an endogenous lysophospholipid with signaling properties outside of the cell and it signals through specific G protein-coupled receptors, known as LPA1-6. For one of its receptors, LPA1 (gene name Lpar1), details on the cis-acting elements for transcriptional control have not been defined. Using 5'RACE analysis, we report the identification of an alternative transcription start site of mouse Lpar1 and characterize approximately 3,500 bp of non-coding flanking sequence 5' of mouse Lpar1 gene for promoter activity. Transient transfection of cells derived from mouse neocortical neuroblasts with constructs from the 5' regions of mouse Lpar1 gene revealed the region between -248 to +225 serving as the basal promoter for Lpar1. This region also lacks a TATA box. For the region between -761 to -248, a negative regulatory element affected the basal expression of Lpar1. This region has three E-box sequences and mutagenesis of these E-boxes, followed by transient expression, demonstrated that two of the E-boxes act as negative modulators of Lpar1. One of these E-box sequences bound the HeLa E-box binding protein (HEB), and modulation of HEB levels in the transfected cells regulated the transcription of the reporter gene. Based on our data, we propose that HEB may be required for a proper regulation of Lpar1 expression in the embryonic neocortical neuroblast cells and to affect its function in both normal brain development and disease settings.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Elementos E-Box/genética , Neurônios/metabolismo , Regiões Promotoras Genéticas , Receptores de Ácidos Lisofosfatídicos/genética , Região 5'-Flanqueadora/genética , Animais , Sequência de Bases , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Regulação da Expressão Gênica , Células HeLa , Humanos , Camundongos , Neocórtex , Ligação Proteica/genética , Receptores de Ácidos Lisofosfatídicos/metabolismo , Deleção de Sequência/genética , Sítio de Iniciação de Transcrição
12.
Animal ; 13(7): 1394-1402, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30378518

RESUMO

Sufficient amino acid (AA) transport is essential to ensure the normal physiological function and growth of growing animals. The processes of AA sensing and transport in humans and murine animals, but rarely in goats, have been arousing great interest recently. This study was conducted to investigate the messenger RNA expression patterns of lysophosphatidic acid receptor 5 (LPAR5), guanine nucleotide-binding protein α-transducing 3 (GNAT3) and important partial AA transporters in digestive tracts, metabolic organs and muscles of growing goats. The results showed that these genes were widely expressed in goats, and had different expression patterns. LPAR5, GNAT3, solute carrier (SLC38A2), SLC7A7, SLC7A1 and SLC3A1 were rarely expressed in the rumen, but were highly expressed in the abomasum and intestine which are the main sites of AA absorption. GNAT3, SLC38A1, SLC38A2, SLC6A19, SLC7A7 and SLC7A1 showed comparatively high expression in the pancreas and the vital digestive glands, and the relatively high expression of these nine genes were noted in the tibialis posterior, the active muscle in energy metabolism. The correlation analysis showed that there were certain positive correlation among most genes. The current results indicate that the AA sensing and transport occur extensively in the abomasum and small intestine, metabolic organs and muscle tissues of ruminants, and that related genes have tissue specificity.


Assuntos
Sistemas de Transporte de Aminoácidos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Cabras/crescimento & desenvolvimento , Cabras/metabolismo , RNA Mensageiro/metabolismo , Abomaso/metabolismo , Sistemas de Transporte de Aminoácidos/genética , Animais , Intestino Delgado/metabolismo , Músculos/metabolismo , RNA Mensageiro/genética , Receptores de Ácidos Lisofosfatídicos/genética , Receptores de Ácidos Lisofosfatídicos/metabolismo , Rúmen/metabolismo , Transducina/genética , Transducina/metabolismo
13.
J Gastroenterol Hepatol ; 34(5): 880-889, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30395690

RESUMO

BACKGROUND AND AIM: Differentially expressed (DE) genes detected at the population-level through case-control comparison provide no information on the dysregulation frequencies of DE genes in a cancer. In this work, we aimed to identify the genes with universally upregulated or downregulated expressions in colorectal cancer (CRC). METHODS: We firstly clarified that DE genes in an individual cancer tissue should be the disease-relevant genes, which are dysregulated in the cancer tissue in comparison with its own previously normal state. Then, we identified DE genes at the individual level for 2233 CRC samples collected from multiple data sources using the RankComp algorithm. RESULTS: We found 26 genes that were upregulated or downregulated in almost all the 2233 CRC samples and validated the results using 124 CRC tissues with paired adjacent normal tissues. Especially, we found that two genes (AJUBA and EGFL6), upregulated universally in CRC tissues, were extremely lowly expressed in normal colorectal tissues, which were considered to be oncogenes in CRC oncogenesis and development. Oppositely, we found that one gene (LPAR1), downregulated universally in CRC tissues, was silenced in CRC tissues but highly expressed in normal colorectal tissues, which were considered to be tumor suppressor genes in CRC. Functional evidences revealed that these three genes may induce CRC through deregulating pathways for ribosome biogenesis, cell proliferation, and cell cycle. CONCLUSIONS: In conclusion, the individual-level DE genes analysis can help us find genes dysregulated universally in CRC tissues, which may be important diagnostic biomarkers and therapy targets.


Assuntos
Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/genética , Regulação para Baixo/genética , Regulação Neoplásica da Expressão Gênica/genética , Proteínas com Domínio LIM/genética , Glicoproteínas de Membrana/genética , Receptores de Ácidos Lisofosfatídicos/genética , Regulação para Cima/genética , Idoso , Neoplasias Colorretais/terapia , Feminino , Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Terapia de Alvo Molecular
14.
Biosci Rep ; 39(1)2019 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-30559147

RESUMO

Purpose: Melanosis coli (MC) is a disorder of pigmentation of the wall of the colon, often identified at the time of colonoscopy. The aim of the present study is to identify candidate biomarkers for MC. Methods: The transcriptome data for MC (GSE78933) with five MC tissues and five corresponding normal tissues is obtained from the NCBI Gene Expression Omnibus (GEO) database. R/Bioconductor package limma was used to screen differently expressed genes (DEGs). ClueGO of cytoscape was applied for Gene Ontology (GO) functional and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment. Based on STRING V10 database, protein-protein interaction (PPI) network was constructed. The pathological tissue and normal tissue from 23 MC patients and 23 controls were collected, respectively. The relative expression of hub nodes was detected by qRT-PCR and Western blot. For regulating the expression of these genes, overexpression vector was constructed or siRNA transfection was used. Finally, apoptosis was detected by flow cytometry. Results: Total 1342 DEGs were screened, including 786 up-regulated and 556 down-regulated genes. These genes were mainly enriched in stimulatory C-type lectin receptor signaling pathway, polysaccharide biosynthetic process, intracellular, and oxidative phosphorylation. PPI network was then constructed with 426 DEGs and 895 interactions. Thereinto, G-protein subunit γ 5 (GNG5), lysophosphatidic acid receptor 3 (LPAR3), mitogen-activated protein kinase 8 (MAPK8), NHP2L1, proteasome 26S subunit, ATPase 6 (PSMC6), and phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit ß (PIK3CB) were hub nodes with higher degree. RT-PCR and Western blot results showed that GNG5, LPAR3, MAPK8, and PSMC6 were differently expressed with significance. The expression of these screened genes is also related with cell apoptosis. Conclusion: GNG5, LPAR3, MAPK8, and PSMC6 might be candidate biomarkers associated with apoptosis in MC.


Assuntos
ATPases Associadas a Diversas Atividades Celulares/genética , Doenças do Colo/genética , Subunidades gama da Proteína de Ligação ao GTP/genética , Melanose/genética , Proteína Quinase 8 Ativada por Mitógeno/genética , Complexo de Endopeptidases do Proteassoma/genética , Receptores de Ácidos Lisofosfatídicos/genética , ATPases Associadas a Diversas Atividades Celulares/metabolismo , Apoptose/fisiologia , Biomarcadores , Estudos de Casos e Controles , Subunidades gama da Proteína de Ligação ao GTP/metabolismo , Humanos , Proteína Quinase 8 Ativada por Mitógeno/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Mapas de Interação de Proteínas , Receptores de Ácidos Lisofosfatídicos/metabolismo , Transcriptoma
15.
BMC Cancer ; 18(1): 1273, 2018 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-30567518

RESUMO

BACKGROUND: Breast cancer is the most common malignancy in women affecting one out of eight females throughout their lives. Autotaxin (ATX) is upregulated in breast cancer which results in increased lysophosphatidic acid (LPA) formation within the tumor. This study's aim was to identify the role of different mammary cell populations within the ATX-LPA axis. METHODS: Epithelial-cell-adhesion-molecule-positive (EpCAM) and -negative cells from breast tumors, adipose-derived stem cells (ADSCs) of tumor-adjacent and tumor-distant mammary fat were isolated and compared to healthy ADSCs, mammary epithelial cells (HMECs), and mesenchymal cells (MES) of healthy mammary tissue (n = 4 each) and further to well-established breast (cancer) cell lines. RESULTS: mRNA expression analyses revealed that ADSCs and MES largely expressed LPA receptor 1 (LPAR1) while epithelial cells mainly expressed LPAR6. LPA 18:1 activated all the cell populations and cell lines by rise in cytosolic free calcium concentrations. MES and ADSCs expressed ATX whereas epithelial cells did not. ADSCs revealed the highest expression in ATX with a significant decline after adipogenic differentiation in healthy ADSCs, whereas ATX expression increased in ADSCs from tumor patients. Breast (cancer) cell lines did not express ATX. Transmigration of MES was stimulated by LPA whereas an inhibitory effect was observed in epithelial cells with no differences between tumors and healthy cells. Triple-negative breast cancer (TNBC) cell lines were also stimulated and the transmigration partly inhibited using the LPA receptor antagonist Ki16425. CONCLUSIONS: We here show that each mammary cell population plays a different role in the ATX-LPA axis with ADSCs and adipocytes being the main source of ATX in tumor patients in our experimental setting. Inhibitors of this axis may therefore present a valuable target for pharmacological therapies.


Assuntos
Lisofosfolipídeos/genética , Diester Fosfórico Hidrolases/genética , Receptores de Ácidos Lisofosfatídicos/genética , Neoplasias de Mama Triplo Negativas/genética , Adipócitos/metabolismo , Adipócitos/patologia , Diferenciação Celular/genética , Linhagem Celular Tumoral , Linhagem da Célula/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Glândulas Mamárias Humanas/metabolismo , Células-Tronco Mesenquimais/metabolismo , RNA Mensageiro/genética , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia
16.
Pol J Vet Sci ; 21(2): 419-421, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30450886

RESUMO

Effect of single nucleotide polymorphism (SNP) in splicing site of the LPAR1 (lysophosphatidic acid receptor 1) gene on selected quality traits was investigated in frozen-thawed semen of Holstein-Friesian bulls. Splicing mutation A/G in the LPAR1 gene (rs43581860) was identified in 120 Holstein-Friesian bulls using PCR-RFLP technique (Hph I). Heterozygotes AG were the most frequent (37.5%) compared with AA (30.8%) and GG (31.7%) homozygotes. Observed differences in total motility (TM), sperm membrane integrity (SYBR-14/PI) and ATP content were significant between homozygotes AA or GG and heterozygotes AG. For all three traits disadvantageous effect of heterozygotes AG was detected. This means that LPAR1 splicing mutation has significant effect on semen quality and should be considered as a new marker of semen quality in Holstein-Friesian bulls.


Assuntos
Mutação , Receptores de Ácidos Lisofosfatídicos , Análise do Sêmen , Animais , Bovinos , Masculino , Receptores de Ácidos Lisofosfatídicos/genética , Sêmen , Preservação do Sêmen , Motilidade Espermática , Espermatozoides
17.
J Recept Signal Transduct Res ; 38(4): 367-371, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30396320

RESUMO

Lysophosphatidic acid (LPA) is a simple biological lipid and mediates several biological functions with LPA receptors (LPA1 to LPA6). In the present study, to assess whether LPA receptors promote cell-invasive activity of pancreatic cancer cells, highly invasion PANC-R9 cells were established from PANC-1 cells, using Matrigel-coated Cell Culture Insert. The cell-invasive activity of PANC-R9 cells was shown to be approximately 15 times higher than that of PANC-1 cells. LPAR1 expression level was markedly elevated in PANC-R9 cells in comparison with PANC-1 cells, while LPAR3 expression level was reduced. The cell-invasive activity of PANC-R9 cells was enhanced by LPA, but LPA had no impact on PANC-1 cell invasion. Before initiation of the cell invasion assay, PANC-R9 cells were pretreated with dioctanoylglycerol pyrophosphate (DGPP), an antagonist of LPA1/LPA3. The invasive activity of PANC-R9 cells was markedly suppressed by DGPP. Autotaxin (ATX) is a key enzyme that catalyzes the conversion of lysophosphatidylcholine (LPC) to LPA. ATX expression level was elevated in PANC-R9 cells compared with PANC-1 cells. In the presence of LPC, the cell motile activity of PANC-R9 cells was markedly stimulated. In contrast, LPC did not affect the cell motile activity of PANC-1 cells. PANC-R9 cell motility was inhibited by an ATX inhibitor, PF-8380. These results suggest that LPA signaling via LPA1 is a potent molecular target for the regulation of tumor progression in PANC-1 cells.


Assuntos
Lisofosfatidilcolinas/metabolismo , Neoplasias Pancreáticas/tratamento farmacológico , Diester Fosfórico Hidrolases/genética , Receptores de Ácidos Lisofosfatídicos/genética , Benzoxazóis/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Humanos , Lisofosfatidilcolinas/genética , Invasividade Neoplásica/genética , Neoplasias Pancreáticas/genética , Ácidos Fosfatídicos/metabolismo , Piperazinas/farmacologia , Receptores de Ácidos Lisofosfatídicos/metabolismo , Transdução de Sinais/efeitos dos fármacos
18.
Cell Physiol Biochem ; 50(2): 597-611, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30317243

RESUMO

BACKGROUND/AIMS: Hyperglycemia has been shown to increase the incidence and metastasis in various types of cancers. However, the correlation between hyperglycemia and lymphatic metastasis in prostate cancer (PCa) remains unclear. Our previous study demonstrated that lysophosphatidic acid (LPA) enhances vascular endothelial growth factor-C (VEGF-C) expression, a lymphangiogenic factor, through activating it receptors LPA1/3 in prostate cancer (PCa) cells. Moreover, hyperglycemia up-regulates autotaxin (ATX) expression, a LPA-generating enzyme. Therefore, we propose that high glucose promotes VEGF-C expression through LPA signaling in PCa cells. METHODS: Pharmacological inhibitors and siRNAs were utilized to investigate the molecular mechanism of high glucose-induced VEGF-C expression. Real-time PCR and Western blot were used to determine the mRNA and protein expressions, respectively. Cellular bioenergetics analysis was performed to determine the glycolysis levels. RESULTS: We demonstrated that the expressions of VEGF-C, ATX, and calreticulin were increased upon high glucose treatments in PC-3 cells. Moreover, high glucose-induced VEGF-C expression was mediated through the LPA1/3, PLC, Akt, ROS and LEDGF-dependent pathways. Additionally, high glucose enhanced the aerobic glycolysis via LPA1/3. CONCLUSION: These results indicated that hyperglycemia leads to LPA synthesis, and subsequent promoting pathological consequence of PCa. These novel findings could potentially provide new strategies for PCa treatments.


Assuntos
Glucose/farmacologia , Transdução de Sinais/efeitos dos fármacos , Fator C de Crescimento do Endotélio Vascular/metabolismo , Calreticulina/antagonistas & inibidores , Calreticulina/genética , Calreticulina/metabolismo , Linhagem Celular Tumoral , Glicólise , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Lisofosfolipídeos/metabolismo , Masculino , Consumo de Oxigênio/efeitos dos fármacos , Fosfoinositídeo Fosfolipase C/genética , Fosfoinositídeo Fosfolipase C/metabolismo , Diester Fosfórico Hidrolases/genética , Diester Fosfórico Hidrolases/metabolismo , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptores de Ácidos Lisofosfatídicos/antagonistas & inibidores , Receptores de Ácidos Lisofosfatídicos/genética , Receptores de Ácidos Lisofosfatídicos/metabolismo , Regulação para Cima/efeitos dos fármacos , Fator C de Crescimento do Endotélio Vascular/genética
19.
J Biol Chem ; 293(44): 17229-17239, 2018 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-30217824

RESUMO

Mammalian target of rapamycin complex 2 (mTORC2) has been shown to regulate mTORC1/4E-BP1/eIF4E signaling and collagen I expression in mesenchymal cells (MCs) during fibrotic activation. Here we investigated the regulation of the mTORC2 binding partner mammalian stress-activated protein kinase-interacting protein 1 (mSin1) in MCs derived from human lung allografts and identified a novel role for mSin1 during fibrosis. mSin1 was identified as a common downstream target of key fibrotic pathways, and its expression was increased in MCs in response to pro-fibrotic mediators: lysophosphatidic acid (LPA), transforming growth factor ß, and interleukin 13. Fibrotic MCs had higher mSin1 protein levels than nonfibrotic MCs, and siRNA-mediated silencing of mSIN1 inhibited collagen I expression and mTORC1/2 activity in these cells. Autocrine LPA signaling contributed to constitutive up-regulation of mSin1 in fibrotic MCs, and mSin1 was decreased because of LPA receptor 1 siRNA treatment. We identified c-Jun N-terminal kinase (JNK) as a key intermediary in mSin1 up-regulation by the pro-fibrotic mediators, as pharmacological and siRNA-mediated inhibition of JNK prevented the LPA-induced mSin1 increase. Proteasomal inhibition rescued mSin1 levels after JNK inhibition in LPA-treated MCs, and the decrease in mSin1 ubiquitination in response to LPA was counteracted by JNK inhibitors. Constitutive JNK1 overexpression induced mSin1 expression and could drive mTORC2 and mTORC1 activation and collagen I expression in nonfibrotic MCs, effects that were reversed by siRNA-mediated mSIN1 silencing. These results indicate that LPA stabilizes mSin1 protein expression via JNK signaling by blocking its proteasomal degradation and identify the LPA/JNK/mSin1/mTORC/collagen I pathway as critical for fibrotic activation of MCs.


Assuntos
Proteínas de Transporte/metabolismo , Fibrose/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Mesoderma/metabolismo , Proteínas de Transporte/genética , Células Cultivadas , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Fibrose/genética , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Pulmão/citologia , Pulmão/metabolismo , Lisofosfolipídeos/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/genética , Mesoderma/citologia , Proteínas Monoméricas de Ligação ao GTP , Fosforilação , Receptores de Ácidos Lisofosfatídicos/genética , Receptores de Ácidos Lisofosfatídicos/metabolismo , Transdução de Sinais
20.
Dis Model Mech ; 11(9)2018 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-30061118

RESUMO

Animal models of psychopathology are particularly useful for studying the neurobiology of depression and characterising the subtypes. Recently, our group was the first to identify a possible relationship between the LPA1 receptor and a mixed anxiety-depression phenotype. Specifically, maLPA1-null mice exhibited a phenotype characterised by depressive and anxious features. However, the constitutive lack of the gene encoding the LPA1 receptor (Lpar1) can induce compensatory mechanisms that might have resulted in the observed deficits. Therefore, in the present study, we have compared the impact of permanent loss and acute pharmacological inhibition of the LPA1 receptor on despair-like behaviours and on the functional brain map associated with these behaviours, as well as on the degree of functional connectivity among structures. Although the antagonist (intracerebroventricularly administered Ki16425) mimicked some, but not all, effects of genetic deletion of the LPA1 receptor on the results of behavioural tests and engaged different brain circuits, both treatments induced depression-like behaviours with an agitation component that was linked to functional changes in key brain regions involved in the stress response and emotional regulation. In addition, both Ki16425 treatment and LPA1 receptor deletion modified the functional brain maps in a way similar to the changes observed in depressed patients. In summary, the pharmacological and genetic approaches could ultimately assist in dissecting the function of the LPA1 receptor in emotional regulation and brain responses, and a combination of those approaches might provide researchers with an opportunity to develop useful drugs that target the LPA1 receptor as treatments for depression, mainly the anxious subtype.This article has an associated First Person interview with the first author of the paper.


Assuntos
Comportamento Animal , Encéfalo/fisiopatologia , Depressão/fisiopatologia , Deleção de Genes , Receptores de Ácidos Lisofosfatídicos/antagonistas & inibidores , Receptores de Ácidos Lisofosfatídicos/genética , Animais , Camundongos Endogâmicos C57BL , Análise de Componente Principal
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