RESUMO
We investigated single nucleotide polymorphism (SNP) of the betaglycan gene (TGFBR3) encoding the TGFß co-receptor in endometrial cancer (EC) and its association with betaglycan expression. The study group included 153 women diagnosed with EC and 248 cancer-free controls. SNP genotyping and gene expression were analyzed using TaqMan probes. Three out of the eight SNPs tested, i.e., rs12566180 (CT; OR = 2.22; 95% CI = 1.15-4.30; p = 0.0177), rs6680463 (GC; OR = 2.34; 95% CI = 1.20-4.53; p = 0.0120) and rs2296621 (TT; OR = 6.40; 95% CI = 1.18-34.84; p = 0.0317) were found to be significantly associated with increased risk of EC (adjusted to age, body mass index, menarche and parity). Among the analyzed SNPs, only rs2296621 demonstrated the impact on the increased cancer aggressiveness evaluated by the WHO grading system (G3 vs. G1/2, GT-OR = 4.04; 95% CI = 1.56-10.51; p = 0.0026; T-OR = 2.38; 95% CI = 1.16-4.85; p = 0.0151). Linkage disequilibrium (LD) analysis revealed high LD (r2 ≥ 0.8) in two haploblocks, constructed by rs2770186/rs12141128 and rs12566180/rs6680463, respectively. In the case of C/C haplotype (OR = 4.82; 95% CI = 1.54-15.07; p = 0.0116-Bonferroni corrected) and T/G haplotype (OR = 3.25; 95% CI = 1.29-8.15; p = 0.0328-Bonferroni corrected) in haploblock rs12566180/rs6680463, significantly higher frequency was observed in patients with EC as compared to the control group. The genotype-phenotype studies showed that SNPs of the TGFBR3 gene associated with an increased risk of EC, i.e., rs12566180 and rs2296621 may affect betaglycan expression at the transcriptomic level (rs12566180-CC vs. TT, p < 0.01; rs2296621-GG vs. TT, p < 0.001, GT vs. TT, p < 0.05). Functional consequences of evaluated TGFBR3 gene SNPs were supported by RegulomeDB search. In conclusion, polymorphism of the TGFBR3 gene may be associated with an increased EC occurrence, as well as may be the molecular mechanism responsible for observed betaglycan down-regulation in EC patients.
RESUMO
Endogenous opioid system is involved in the maintenance of the intestinal homeostasis. Recently, we proved that stimulation of opioid receptors using P-317, a cyclic morphiceptin analog, resulted in the alleviation of acute colitis in mice. The aim of the current study was to assess the effect of P-317 during colitis and colitis-associated colorectal cancer in mice. Colitis was induced by addition of dextran sodium sulfate (DSS) into drinking water. Colitis-associated colorectal cancer was induced by a single intraperitoneal injection of azoxymethane (AOM) and subsequent addition of DSS into drinking water (week 2, 5, 8). During macroscopic damage evaluation the samples were collected and used for biochemical (MPO activity assay), molecular (qPCR and western blot) and histological studies. In experimental colitis, P-317 induced an anti-inflammatory response as indicated by macroscopic and microscopic scores. In the colitis-associated colorectal cancer model, a significant difference in colorectal tumor development was observed between vehicle- and P-317-treated mice. P-317 decreased the total number of colonic tumors and inhibited MPO activity. Hematoxylin and eosin staining confirmed anti-tumor activity of P-317. The expression of TNF-α was decreased in P-317-treated mice as compared to the vehicle-treated group. P-317 decreased proliferation as well as ß-catenin expression in tumors. P-317, a mixed MOP and KOP receptor agonist, induced an anti-inflammatory response in experimental colitis and decreased tumor development in colitis-associated colorectal cancer in mice.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Antineoplásicos/farmacologia , Colite/tratamento farmacológico , Neoplasias Colorretais/tratamento farmacológico , Endorfinas/farmacologia , Receptores Opioides kappa/agonistas , Receptores Opioides mu/agonistas , Animais , Anti-Inflamatórios não Esteroides/uso terapêutico , Antineoplásicos/uso terapêutico , Azoximetano , Carcinógenos , Proliferação de Células/efeitos dos fármacos , Colite/induzido quimicamente , Colite/complicações , Neoplasias Colorretais/etiologia , Sulfato de Dextrana , Endorfinas/uso terapêutico , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fator de Necrose Tumoral alfa/antagonistas & inibidoresRESUMO
The regulatory role of estrogens and nuclear estrogen receptors, i. e., estrogen receptor α and ß has been reported in gastrointestinal diseases. However, the contribution of G protein-coupled estrogen receptor, the membrane-bound estrogen receptor, is still poorly understood. Unlike nuclear estrogen receptors, which are responsible for the genomic activity of estrogens, the G protein-coupled estrogen receptor affects the "rapid" non-genomic activity of estrogens, leading to modulation of many signaling pathways and ultimately changing gene expression. Recently, the crucial role of G protein-coupled estrogen receptor in intestinal pathogenesis has been documented. It has been shown that the G protein-coupled estrogen receptor can modulate the progression of irritable bowel syndrome, inflammatory bowel diseases such as Crohn's disease and ulcerative colitis as well as colorectal cancer. The G protein-coupled estrogen receptor appears to be a potent factor regulating abdominal sensitivity and pain, intestinal peristalsis, colitis development, proliferation and migration potential of colorectal cancer cells and seems to be a useful target in gastrointestinal diseases. In this review, we present the current state of knowledge about the contribution of the G protein-coupled estrogen receptor to irritable bowel syndrome, inflammatory bowel diseases and colorectal cancer.
Assuntos
Neoplasias Colorretais/fisiopatologia , Doenças Inflamatórias Intestinais/fisiopatologia , Síndrome do Intestino Irritável/fisiopatologia , Receptores de Estrogênio/fisiologia , Receptores Acoplados a Proteínas G/fisiologia , Animais , Humanos , Transdução de SinaisRESUMO
Crohn's disease is the most diagnosed type of inflammatory bowel disease. Chronic inflammation developing in the intestine leads to peristalsis disorder and damage of intestinal mucosa and seems to be associated with an increased risk of colon neoplastic transformation. Accumulating evidence indicates that estrogens and estrogen receptors affect not only hormone-sensitive tissues, but also other tissues not directly related to estrogens, such as the lungs or colon. Here, we describe the protocol for the successful immunofluorescence staining of estrogen receptors in colon obtained from a murine model of TNBS-induced Crohn's disease. A detailed protocol for the induction of Crohn's disease in mice and intestine preparation is provided as well as a step-by-step immunohistochemical procedure using formalin-fixed paraffin-embedded intestine sections. The described methods are not only useful for estrogen receptor detection and estrogen signaling investigation in vivo but can also be applied to for other proteins which may be involved in the development of colitis.
Assuntos
Colo/metabolismo , Doença de Crohn/fisiopatologia , Imunofluorescência/métodos , Receptores de Estrogênio/metabolismo , Animais , Colo/citologia , Humanos , CamundongosRESUMO
Estrogens play important roles in the development and progression of multiple tumor types. Accumulating evidence points to the significance of estrogen action not only in tumors of hormonally regulated tissues such as the breast, endometrium and ovary, but also in the development of colorectal cancer (CRC). The effects of estrogens in physiological and pathophysiological conditions are mediated by the nuclear estrogen receptors α and ß, as well as the membrane-bound G protein-coupled estrogen receptor (GPER). The roles of GPER in CRC development and progression, however, remain poorly understood. Studies on the functions of GPER in the colon have shown that this estrogen receptor regulates colonic motility as well as immune responses in CRC-associated diseases, such as Crohn's disease and ulcerative colitis. GPER is also involved in cell cycle regulation, endoplasmic reticulum stress, proliferation, apoptosis, vascularization, cell migration, and the regulation of fatty acid and estrogen metabolism in CRC cells. Thus, multiple lines of evidence suggest that GPER may play an important role in colorectal carcinogenesis. In this review, we present the current state of knowledge regarding the contribution of GPER to colon function and CRC.
Assuntos
Colite Ulcerativa/patologia , Colo/fisiopatologia , Neoplasias Colorretais/patologia , Doença de Crohn/patologia , Receptores de Estrogênio/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Animais , Carcinogênese/imunologia , Carcinogênese/patologia , Colite Ulcerativa/imunologia , Colite Ulcerativa/fisiopatologia , Colo/imunologia , Colo/patologia , Neoplasias Colorretais/imunologia , Doença de Crohn/imunologia , Doença de Crohn/fisiopatologia , Modelos Animais de Doenças , Progressão da Doença , Estrogênios/metabolismo , Motilidade Gastrointestinal/imunologia , Humanos , Mucosa Intestinal/imunologia , Mucosa Intestinal/patologia , Mucosa Intestinal/fisiopatologiaRESUMO
The pathogenesis of inflammatory bowel diseases (IBD) seems to be associated with alterations of immunoregulation. Several lines of evidence suggest that estrogens play a role in the modulation of immune responses and may be related to the etiology of IBD. The purpose of this work was to examine the involvement of G protein-coupled estrogen receptor (GPER), estrogen receptor α (ERα), estrogen receptor ß (ERß) and ERα spliced variants ERα36 and ERα46 in Crohn's disease (CD) and ulcerative colitis (UC). The studied group included 73 patients with IBD and 31 sex and age-related controls. No differences in serum levels of 17ß-estradiol nor of CYP1A1 and SULT1E1 enzymes involved in estrogen catabolism were stated. The expression pattern of estrogen receptors in tissue samples was quantified using real-time PCR and Western blotting. Statistically significant up-regulation of GPER and ERα in both CD and UC as well as down-regulation of ERß in CD patients was found. However, differences in the expression of estrogen receptors in CD and UC have been identified, depending on the sex and age of patients. In men, up-regulation of GPER, ERα and ERα46 expression was shown in CD and UC patients. In women under 50 years of age, GPER protein level increased in UC whereas ERß expression tended to decrease in CD and UC patients. In turn, in women over 50 the protein level of ERα increased in UC while ERß expression decreased in CD patients. Dysregulation of estrogen receptors in the intestinal mucosa of patients with CD and UC indicates that estrogen signaling may play a role in the local immune response and maintain epithelial homeostasis in a gender- and age-dependent manner.
Assuntos
Colite Ulcerativa/metabolismo , Doença de Crohn/metabolismo , Estrogênios/metabolismo , Receptores de Estrogênio/metabolismo , Transdução de Sinais , Fatores Etários , Idoso , Colite Ulcerativa/patologia , Doença de Crohn/patologia , Estradiol/sangue , Estradiol/metabolismo , Estrogênios/sangue , Feminino , Humanos , Mucosa Intestinal/metabolismo , Masculino , Pessoa de Meia-Idade , Receptores de Estrogênio/genética , Fatores SexuaisRESUMO
Genetic polymorphisms in DNA repair genes may affect DNA repair efficiency and may contribute to the risk of developing cancer. The aim of our study was to investigate single nucleotide polymorphisms (SNPs) in RAD51 (rs2619679, rs2928140, and rs5030789) and XRCC3 (rs1799796) involved in DNA double-strand break repair and their relationship to prostate cancer. The study group included 99 men diagnosed with prostate cancer and 205 cancer-free controls. SNP genotyping was performed using the PCR-RFLP method. A significant association was detected between RAD51 rs5030789 polymorphism and XRCC3 rs1799796 polymorphism and an increased risk of prostate cancer. Our results indicate that RAD51 and XRCC3 polymorphism may contribute to prostate cancer.
RESUMO
Estrogens exert immunomodulatory action in many autoimmune diseases. Accumulating evidence highlights the meaningful impact of estrogen receptors in physiology and pathophysiology of the colon. However, the significance of G protein-coupled estrogen receptor (GPER) on Crohn's disease (CD), one of the inflammatory bowel disease (IBD) types, is still elusive. Our study revealed GPER overexpression at the mRNA and protein levels in patients with CD. To evaluate the effects of GPER activation/inhibition on colitis development, a murine 2, 4, 6-trinitrobenzene sulfonic acid (TNBS)-induced model of CD was used. We showed that activation of GPER reduces mortality, improves macroscopic and microscopic scores and lowers C-reactive protein (CRP) level. The impact of estrogen signaling on the suppression of the intestinal inflammation was proved by immunohistochemistry. It was demonstrated that GPER activation is accompanied by modulation of extracellular-signal regulated kinase (ERK) signaling pathway and expression level of genes involved in signal transmission and immune response as well as the expression of some microRNAs (miR-145, miR-148-5p and miR-592). Our study revealed that the membrane-bound estrogen receptor GPER mediates anti-inflammatory action and seems to be a potent therapeutic target in maintaining remission in CD.
Assuntos
Doença de Crohn/etiologia , Doença de Crohn/metabolismo , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Animais , Biomarcadores , Biópsia , Doença de Crohn/patologia , Citocinas/metabolismo , Modelos Animais de Doenças , Suscetibilidade a Doenças , Estrogênios/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Expressão Gênica , Humanos , Imunomodulação/genética , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Camundongos , Transporte Proteico , Transdução de Sinais/efeitos dos fármacosRESUMO
BACKGROUND AND AIMS: Molecular basis of abdominal pain in IBD is not fully characterized. Serotonin (5-HT) increases visceral pain severity and contributes to exacerbation of inflammation. Moreover, it is well established that decreased anandamide (AEA) signaling in the gut increases visceral pain severity. The aim of this study was to investigate the interplay between 5-HT and endocannabinoid signaling in colitis. METHODS: We used 3% DSS in drinking water to induce colitis in mice. From day 3 5-HT was administered for 5â¯days and each day visceromotor response to colorectal distention (CRD) was measured. Expression of cannabinoid (CB) receptors as well as enzymes responsible of biosynthesis and degradation of endocannabinoids were investigated. Moreover, endocannabinoid levels were assessed by mass spectrometry. Additionally, we measured the expression of enzymes synthesizing 5-HT and AEA in the colon of IBD patients and healthy controls. RESULTS: Chronic exposure to 5-HT increased visceromotor response to CRD and worsened colitis, which was associated with decrease of AEA via 5-HT3 and 5-HT4 receptors. Moreover, exposure to 5-HT led to the downregulation of CB1 receptors. Colonic levels of N-acyl-phosphatidylethanolamine-specific phospholipase D (NAPE-PLD), which is responsible for synthesis of AEA, significantly declined after chronic treatment with 5-HT and this effect was reversed by the 5-HT3 and 5-HT4 receptor antagonists. NAPE-PLD was also downregulated in the colon of UC patients. CONCLUSIONS: Our study shows a link between 5-HT and endocannabinoid signaling pathways in IBD. Thus, pharmacological blockade of 5-HT signaling or supplementation with endocannabinoids in the gut might be of benefit in severe cases of abdominal pain in IBD.
Assuntos
Dor Abdominal/metabolismo , Colite/metabolismo , Endocanabinoides/metabolismo , Serotonina/metabolismo , Serotonina/toxicidade , Dor Abdominal/induzido quimicamente , Adulto , Idoso , Animais , Colite/induzido quimicamente , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , Serotonina/administração & dosagem , Adulto JovemRESUMO
The etiology and pathogenesis underlying irritable bowel syndrome (IBS), a common gastrointestinal disorder are still unclear. Cumulating data suggest dysregulation of inflammatory and immune response pathways and changes of epithelial barrier function. The role of estrogens albeit varied, in responses of immune system is well documented. The aim of this study was to investigate estrogen receptors engagment in IBS subtypes, i.e. constipation predominant (IBS-C) and diarrhea predominant (IBS-D). Furthermore, we analyzed whether estrogen signaling is accompanied by alteration in expression of pro- and anti-inflammatory cytokines and microRNAs that can regulate among others genes involved in immune responses. It was found that estrogen receptor α (ERα) and G protein-coupled estrogen receptor (GPER) expression is up-regulated in IBS while estrogen receptor ß (ERß) appears to be down-regulated at mRNA but up-regulated at the protein level. When gender and female age were included statistically significant overexpression of ERα in IBS-D women under the age of 50, while of GPER in IBS-D men was stated. In all studied IBS samples disturbances in expression of cytokines IL-6, IL-10 and TNF-α as well as miR-145, miR-148-5p and miR-592 were observed. This research reveals the association of estrogen receptors with IBS. Simultaneous alterations of studied immunomodulatory cytokines and microRNAs suggest that in IBS dysregulation of local immune response may involve estrogen-dependent way.
Assuntos
Estrogênios/metabolismo , Síndrome do Intestino Irritável/imunologia , Transdução de Sinais , Adulto , Idoso , Idoso de 80 Anos ou mais , Citocinas/genética , Citocinas/metabolismo , Estradiol/sangue , Feminino , Humanos , Imunomodulação/genética , Síndrome do Intestino Irritável/sangue , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismoRESUMO
AIM: To investigate the levels of G protein-coupled receptor 55 (GPR55) expression in colonic tissue of inflammatory bowel disease (IBD) patients and healthy controls, and its potential implication in IBD treatment. METHODS: Fifty patients were enrolled in our prospective study: n = 21 with Crohn's disease (CD) and n = 16 with ulcerative colitis (UC); 19 women and 18 men. Control consisted of 13 non-IBD patients. In each subject, two biopsies were taken from different colonic locations. In IBD patients, biopsies both from endoscopically inflamed and non-inflamed areas were drawn and the development of inflammation confirmed in histopathological examination. GPR55 mRNA and protein expression were measured using real-time PCR and Western blot, respectively. RESULTS: GPR55 expression at mRNA and protein level was detected in all samples tested. The level of GPR55 mRNA expression in non-inflamed colonic areas was comparable in all analyzed groups (p = .2438). However, in the inflamed tissues GPR55 mRNA expression was statistically significantly (p < .0001) higher (6.9 fold) in CD patients compared to UC. Moreover, CD patients manifested higher (12.5 fold) GPR55 mRNA expression in inflamed compared with non-inflamed colonic tissues (p < .0001). Although no significant differences were stated, GPR55 protein level tends to decrease in IBD as compared to control. CONCLUSIONS: Different patterns of GPR55 expression at mRNA level were observed in IBD patients. We speculate that GPR55 is crucial for the mucosal inflammatory processes in IBD, particularly in CD and its expression may affect disease severity, and response to treatment. The GPR55 receptors may become an attractive target for novel therapeutic strategies in IBD.
Assuntos
Colite Ulcerativa/genética , Colo/patologia , Doença de Crohn/genética , Receptores Acoplados a Proteínas G/metabolismo , Adulto , Western Blotting , Estudos de Casos e Controles , Colo/metabolismo , Feminino , Humanos , Masculino , Polônia , Estudos Prospectivos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Receptores de Canabinoides , Receptores Acoplados a Proteínas G/genéticaRESUMO
BACKGROUND AND AIMS: G protein-coupled receptor 30 (GPR30) is a recently de-orphanized estrogen receptor that mediates the effects of estrogens on different cells. It has been postulated that in inflammatory bowel diseases (IBD) activation of GPR30 blocks the pathways dependent on pro-inflammatory cytokines. The aim of our study was to investigate GPR30 expression in patients with IBD and its potential implication in future therapies. METHODS: Fifty-seven patients were enrolled in our study: 20 subjects with Crohn's disease (CD), 22 with ulcerative colitis (UC) and 15 controls. In each subject, biopsies were taken from various left-colonic locations. Gene and protein expression of GPR30 was quantified using real time RT-PCR or Western blot. RESULTS: GPR30 mRNA and protein expression were detected in all tested colonic tissues. No significant differences in GPR30 gene expression were observed. In non-inflamed areas, GPR30 protein was strongly increased in CD patients, but moderately in UC patients (p= 0.014 and p=0.143, respectively, vs. controls). In CD patients, a significantly lower GPR30 protein content in inflamed than in non-inflamed tissue was observed (p=0.039). The change was independent of patient gender. CONCLUSION: Our observations indicate that GPR30 may play a role in the development and progression of inflammatory lesions in IBD, thus affecting disease severity, and consequently IBD treatment. Therefore, GPR30 may become an attractive target for novel anti-IBD drugs, particularly in CD.
Assuntos
Mediadores da Inflamação/metabolismo , Doenças Inflamatórias Intestinais/metabolismo , Receptores de Estrogênio/biossíntese , Receptores Acoplados a Proteínas G/biossíntese , Adulto , Estudos de Casos e Controles , Colite Ulcerativa/genética , Colite Ulcerativa/metabolismo , Colo/metabolismo , Doença de Crohn/genética , Doença de Crohn/metabolismo , Progressão da Doença , Feminino , Regulação da Expressão Gênica , Humanos , Doenças Inflamatórias Intestinais/genética , Masculino , Projetos Piloto , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Receptores de Estrogênio/genética , Receptores Acoplados a Proteínas G/genética , Adulto JovemRESUMO
Estrogens play a crucial role in the regulation of physiological and pathophysiological processes. These hormones act through specific receptors, most notably the canonical estrogen receptors α and ß (ERα and ERß) and their truncated forms as well as the G protein-coupled estrogen receptor (GPER). Several studies have shown that GPER is expressed in many normal and cancer cells, including those of the breast, endometrium, ovary, testis and lung. Hormonal imbalance is one possible cause of cancer development. An accumulating body of evidence indicates that GPER is involved in the regulation of cancer cell proliferation, migration and invasion, it may act as a mediator of microRNA, and is believed to modulate the inflammation associated with neoplastic transformation. Furthermore, used in various treatment regimens anti-estrogens such as tamoxifen, raloxifen and fulvestrant (ICI 182.780), antagonists/modulators of canonical estrogen receptors, were found to be GPER agonists. This review presents the current knowledge about the potential role of GPER in neoplastic transformation.
Assuntos
Transformação Celular Neoplásica/metabolismo , Receptores de Estrogênio/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Animais , HumanosRESUMO
Downregulation of betaglycan (ß-glycan) [transforming growth factor ß receptor type III (TGFßR3)], which belongs to co-receptors of the TGFß pathway, occurs in a broad spectrum of primary human malignancies. However, in the case of endometrial cancer (EC), the mechanisms responsible for genetic alterations are still unknown. Therefore, we investigated allelic imbalance at the TGFBR3 locus (1p33p32) in the context of ß-glycan mRNA and protein expression, as a possible genetic event determining ß-glycan deregulation in EC patients. Study of ß-glycan allelic imbalance in 48 primary human ECs was performed with the use of three different microsatellite markers, spanned within or in direct proximity to the TGFBR3 locus. Realtime PCR and western blotting were used for ß-glycan mRNA and protein quantification methods, respectively. Altogether, 25 of 39 (64%) informative cases and 25 of 48 (52%) of all specimens showed allelic imbalance in at least one microsatellite marker, concomitantly with decrease at both the ß-glycan transcript and protein levels. Interestingly, 54% (15/28), 36% (8/22) and 35% (7/20) of informative ECs displayed allelic loss in D1S188, D1S435 and D1S1588 microsatellite markers, respectively. It is worth pointing out that 5 out of 39 (13%) informative cases showed loss of heterozygosity (LOH) at two microsatellite markers. Microsatellite instability (MSI) was found in two markers, but to a very strictly limited extent. None of the clinicoprognostic features was found to be of significance. Our results suggest that LOH in the TGFBR3 locus may be one of the mechanisms responsible for loss of ß-glycan expression. No correlation of LOH at the TGFBR3 locus with clinicopathological parameters suggests that allelic imbalance may be an early genetic event during neoplastic transformation of human endometrium.
Assuntos
Neoplasias do Endométrio/genética , Proteoglicanas/genética , Receptores de Fatores de Crescimento Transformadores beta/genética , Transdução de Sinais/fisiologia , Fator de Crescimento Transformador beta/metabolismo , Idoso , Western Blotting , Neoplasias do Endométrio/metabolismo , Feminino , Humanos , Perda de Heterozigosidade , Instabilidade de Microssatélites , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Although prostate cancer is one of the most common cancers in men, the genetic defects underlying its pathogenesis remain poorly understood. DNA damage repair mechanisms have been implicated in human cancer. Accumulating evidence indicates that the fidelity of the response to DNA double-strand breaks is critical for maintaining genome integrity. RAD51 is a central player in double-strand break repair via homologous recombination, and its alterations may confer and increase the risk of cancer. RAD51 functioning depends on the indirect or direct interactions with BRCA1 and BRCA2. To evaluate the contribution of RAD51 to sporadic prostate cancer, loss of heterozygosity (LOH) for chromosomal region 15q14-21.1 (RAD51 locus) was determined and compared to LOH in 17q21.31 (BRCA1 locus) and 13q12.3-13.1 (BRCA2 region). DNA was isolated from prostate biopsies and matched peripheral blood of 50 patients. The regions 15q14-21.1, 17q21.31, and 13q12.3-13.1 were examined using microsatellite markers on chromosome 15 (D15S118, D15S214, D15S1006), chromosome 17 (D17S855, D17S1323), and chromosome 13 (D13S260, D13S290), respectively. The LOH in tumors was analyzed by PCR with fluorescently labeled primers and an ABI PRISM 377 DNA Sequencer. Allele sizing was determined by GeneScan version 3.1.2 and Genotyper version 2.5 software (Applied Biosystems, USA). LOH was identified in 57.5, 23, and 40 % for chromosomal regions 15q14-21.1, 17q21.31, and 13q12.3-13.1, respectively. Twenty-six percent of studied cases manifested LOH for at least one marker in 15q14-21.1 exclusively. A significant correlation was found between LOH for studied region and PSAD (prostate-specific antigen density). The findings suggest that RAD51 may be considered as a prostate cancer susceptibility gene.
Assuntos
Cromossomos Humanos Par 13/genética , Cromossomos Humanos Par 15/genética , Cromossomos Humanos Par 17/genética , Perda de Heterozigosidade/genética , Neoplasias da Próstata/diagnóstico , Neoplasias da Próstata/genética , Idoso , Idoso de 80 Anos ou mais , Proteína BRCA1/genética , Proteína BRCA2/genética , Humanos , Masculino , Pessoa de Meia-Idade , Rad51 Recombinase/genéticaRESUMO
Genetic polymorphisms in DNA repair genes may induce individual variations in DNA repair capacity, which may in turn contribute to the risk of cancer developing. Homologous recombination repair (HRR) plays a critical role in maintaining chromosomal integrity and protecting against carcinogenic factors. The aim of the present study was to evaluate the relationship between prostate cancer risk and the presence of single nucleotide polymorphisms (SNPs) in the genes involved in HRR, that is, RAD51 (rs1801320 and rs1801321), RAD51B (rs10483813 and rs3784099), XRCC2 (rs3218536), and XRCC3 (rs861539). Polymorphisms were analyzed by PCR-RFLP and Real-Time PCR in 101 patients with prostate adenocarcinoma and 216 age- and sex-matched controls. A significant relationship was detected between the RAD51 gene rs1801320 polymorphism and increased prostate cancer risk. Our results indicate that the RAD51 gene rs1801320 polymorphism may contribute to prostate cancer susceptibility in Poland.
Assuntos
Proteínas de Ligação a DNA/genética , Predisposição Genética para Doença , Recombinação Homóloga/genética , Polimorfismo de Nucleotídeo Único/genética , Neoplasias da Próstata/genética , Rad51 Recombinase/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Frequência do Gene/genética , Loci Gênicos , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Fatores de RiscoRESUMO
Recent studies reveal that estrogens act on cells and tissues, not only through two canonical estrogen receptors ERα and ERß, but also through the receptor coupled with G proteins, named GPER, formerly GPR30, and member of seven transmembrane receptor superfamily. GPER was found to be implicated both in rapid non-genomic estrogen response and in transcriptional regulation. Effects of GPER include activation of MAPK and PI3K signaling pathways, stimulation of adenyl cyclase, and mobilization of calcium ions from intracellular stores, as well as upregulation of genes such as FOS and CTGF. Identification of agonists and antagonists of GPER allowed not only to shed light on the participation of estrogen signaling in the biology and pathobiology of hormone-dependent tissues, but also its importance in the estrogen and antiestrogen therapies. Antiestrogens tamoxifen, raloxifene, or fulvestrant proved,to be agonists of GPER, which undoubtedly is not without significance for the efficacy of the therapy.
Assuntos
Fosfatase 1 de Especificidade Dupla/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Receptores de Estrogênio/metabolismo , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/metabolismo , Transdução de Sinais/genética , Animais , Comunicação Autócrina/genética , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Estradiol/análogos & derivados , Estradiol/farmacologia , Feminino , Fulvestranto , Regulação da Expressão Gênica , Humanos , Proteínas Proto-Oncogênicas c-fos/metabolismo , Cloridrato de Raloxifeno/farmacologia , Receptores de Estrogênio/antagonistas & inibidores , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Tamoxifeno/farmacologia , Ativação TranscricionalRESUMO
OBJECTIVE: The aim of our study was to characterize the effect of P-317, a novel cyclic derivative of morphiceptin, on gastrointestinal (GI) motility and abdominal pain in mouse models mimicking symptoms of diarrhoea-predominant irritable bowel syndrome (IBS-D). METHODS: The effect of P-317 on mouse intestinal motility was characterized in vitro and in vivo in physiological and pathophysiological conditions. The antinociceptive action of P-317 was characterized in the mustard oil-induced abdominal pain model and the writhing test. Locomotor activity and grip-strength tests were used to evaluate the effect of P-317 in the central nervous system (CNS). To translate our study to clinical conditions, the semi-quantitative expression of µ-opioid receptors (MOP) and κ-opioid receptors (KOP) messenger RNA (mRNA) in human colonic samples from IBS-D patients was quantified. KEY FINDINGS: In vitro, P-317 (10(-10) -10(-6) M) inhibited colonic and ileal smooth muscle contractions in a concentration-dependent, ß-funaltrexamine and nor-binaltorphimine-reversible manner. In vivo, P-317 (0.1 mg/kg, i.p. and 1 mg/kg, p.o.) inhibited GI transit, displayed a potent antinociceptive action in abdominal pain tests and did not influence the CNS. CONCLUSION: P-317 produced a potent analgesic and antidiarrhoeal action in the mouse GI tract after oral administration. Given lower expression of MOP and KOP mRNA in IBS-D patients, P-317 is a promising peptide-based drug candidate for IBS-D therapy.
Assuntos
Dor Abdominal/tratamento farmacológico , Analgésicos/uso terapêutico , Antidiarreicos/uso terapêutico , Diarreia/tratamento farmacológico , Endorfinas/uso terapêutico , Trânsito Gastrointestinal/efeitos dos fármacos , Síndrome do Intestino Irritável/tratamento farmacológico , Dor Abdominal/induzido quimicamente , Administração Oral , Adulto , Idoso , Analgésicos/farmacologia , Animais , Antidiarreicos/farmacologia , Estudos de Casos e Controles , Colo/efeitos dos fármacos , Colo/metabolismo , Colo/fisiopatologia , Diarreia/etiologia , Modelos Animais de Doenças , Endorfinas/farmacologia , Humanos , Íleo/efeitos dos fármacos , Síndrome do Intestino Irritável/patologia , Masculino , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Mostardeira , Óleos de Plantas , Receptores Opioides/genética , Receptores Opioides/metabolismo , Adulto JovemRESUMO
BACKGROUND: Salvinorin A (SA) is a potent anti-inflammatory diterpene isolated from the Mexican plant S. divinorum. Recently we showed that the novel SA analog, PR-38 has an inhibitory effect on mouse gastrointestinal (GI) motility mediated by opioid and cannabinoid (CB) receptors. The aim of the study was to characterize possible anti-inflammatory and antinociceptive action of PR-38 in the mouse GI tract. METHODS: Macro- and microscopic colonic damage scores and myeloperoxidase activity were determined after intraperitoneal (i.p.), intracolonic (i.c.), and per os (p.o.) administration of PR-38 in the trinitrobenzene sulfonic acid (TNBS) and dextran sodium sulfate (DSS) models of colitis in mice. Additionally, MOP, KOP and CB1 protein expression was determined using Western blot analysis of mouse colon samples. The antinociceptive effect of PR-38 was examined based on the number of behavioral responses to i.c. instillation of mustard oil (MO). RESULTS: The i.p. (10 mg/kg, twice daily), i.c. (10 mg/kg, twice daily) and p.o. (20 mg/kg, once daily) administration of PR-38 significantly attenuated TNBS- and DSS-induced colitis in mice. The effect of PR-38 was partially blocked by the KOP antagonist nor-binaltorphimine and CB1 antagonist AM 251. Western blot analysis showed a significant increase of MOP, KOP and CB1 receptor expression during colonic inflammation, which was reversed to the control levels by the administration of PR-38. PR-38 significantly decreased the number of pain responses after i.c. instillation of MO in the TNBS-treated mice. CONCLUSIONS: Our results suggest that PR-38 has the potential to become a valuable anti-inflammatory and analgesic therapeutic for the treatment of GI inflammation.