RESUMO
BACKGROUND: Colorectal polyps that develop via the conventional adenoma-carcinoma sequence [e.g., tubular adenoma (TA)] often progress to malignancy and are closely associated with changes in the composition of the gut microbiome. There is limited research concerning the microbial functions and gut microbiomes associated with colorectal polyps that arise through the serrated polyp pathway, such as hyperplastic polyps (HP). Exploration of microbiome alterations associated with HP and TA would improve the understanding of mechanisms by which specific microbes and their metabolic pathways contribute to colorectal carcinogenesis. AIM: To investigate gut microbiome signatures, microbial associations, and microbial functions in HP and TA patients. METHODS: Full-length 16S rRNA sequencing was used to characterize the gut microbiome in stool samples from control participants without polyps [control group (CT), n = 40], patients with HP (n = 52), and patients with TA (n = 60). Significant differences in gut microbiome composition and functional mechanisms were identified between the CT group and patients with HP or TA. Analytical techniques in this study included differential abundance analysis, co-occurrence network analysis, and differential pathway analysis. RESULTS: Colorectal cancer (CRC)-associated bacteria, including Streptococcus gallolyticus (S. gallolyticus), Bacteroides fragilis, and Clostridium symbiosum, were identified as characteristic microbial species in TA patients. Mediterraneibacter gnavus, associated with dysbiosis and gastrointestinal diseases, was significantly differentially abundant in the HP and TA groups. Functional pathway analysis revealed that HP patients exhibited enrichment in the sulfur oxidation pathway exclusively, whereas TA patients showed dominance in pathways related to secondary metabolite biosynthesis (e.g., mevalonate); S. gallolyticus was a major contributor. Co-occurrence network and dynamic network analyses revealed co-occurrence of dysbiosis-associated bacteria in HP patients, whereas TA patients exhibited co-occurrence of CRC-associated bacteria. Furthermore, the co-occurrence of SCFA-producing bacteria was lower in TA patients than HP patients. CONCLUSION: This study revealed distinct gut microbiome signatures associated with pathways of colorectal polyp development, providing insights concerning the roles of microbial species, functional pathways, and microbial interactions in colorectal carcinogenesis.
Assuntos
Pólipos do Colo , Neoplasias Colorretais , Fezes , Microbioma Gastrointestinal , RNA Ribossômico 16S , Humanos , Feminino , Masculino , Pessoa de Meia-Idade , Pólipos do Colo/microbiologia , Pólipos do Colo/patologia , Neoplasias Colorretais/microbiologia , Neoplasias Colorretais/patologia , RNA Ribossômico 16S/genética , Idoso , Fezes/microbiologia , Tailândia/epidemiologia , Adulto , Adenoma/microbiologia , Bactérias/isolamento & purificação , Bactérias/genética , Bactérias/classificação , Hiperplasia/microbiologia , Estudos de Casos e Controles , Disbiose/microbiologia , População do Sudeste AsiáticoRESUMO
Novel compound heterozygous mutations, G701D, a recessive mutation, and A858D, a mild dominant mutation, of human solute carrier family 4, anion exchanger, member 1 (SLC4A1) were identified in two pediatric patients with distal renal tubular acidosis (dRTA). To examine the interaction, trafficking, and cellular localization of the wild-type and two mutant kidney AE1 (kAE1) proteins, we expressed the proteins alone or together in human embryonic kidney (HEK) 293T and Madin-Darby canine kidney (MDCK) epithelial cells. In individual expressions, wild-type kAE1 was localized at the cell surface of HEK 293T and the basolateral membrane of MDCK cells. In contrast, kAE1 G701D was mainly retained intracellularly, while kAE1 A858D was observed intracellularly and at the cell surface. In co-expression experiments, wild-type kAE1 formed heterodimers with kAE1 G701D and kAE1 A858D, and promoted the cell surface expression of the mutant proteins. The co-expressed kAE1 G701D and A858D could also form heterodimers but showed predominant intracellular retention in HEK 293T and MDCK cells. Thus impaired trafficking of the kAE1 G701D and A858D mutants would lead to a profound decrease in functional kAE1 at the basolateral membrane of alpha-intercalated cells in the distal nephron of the patients with dRTA.
Assuntos
Acidose Tubular Renal/metabolismo , Substituição de Aminoácidos/genética , Proteína 1 de Troca de Ânion do Eritrócito/metabolismo , Espaço Intracelular/metabolismo , Proteínas Mutantes/metabolismo , Animais , Western Blotting , Extratos Celulares , Linhagem Celular , Criança , Cães , Citometria de Fluxo , Imunofluorescência , Humanos , Multimerização Proteica , Transporte Proteico , Frações Subcelulares/metabolismoRESUMO
Osteogenesis imperfecta (OI) is commonly caused by monoallelic mutations in COL1A1 or COL1A2. Biallelic mutations are extremely rare. Only five previous reports have identified seven OI patients with homozygous mutations in COL1A2. OI is a genetically and phenotypically heterogeneous disorder which challenges an establishment of genotype-phenotype correlation. Notably, more than thirty patients with OI possess the heterozygous mutation, p.Gly337Ser, in COL1A2. Their clinical severity ranges from mild OI type I to severe types III and IV. Here, we report a 17-year-old Thai female with recurrent bone fractures, short stature, blue sclerae, triangular face, missing teeth, dentinogenesis imperfecta (DI), skeletal deformities, and scoliosis. She was diagnosed with OI type III. Her parents were second-cousin-once-removed. The father was a professional Thai boxer. Both had normal bone mineral density, no history of bone fractures, and only teeth problems. They were diagnosed with DI without OI. Whole exome sequencing identified that the proband harbored the homozygous mutation, c.1009G > A (p.Gly337Ser), in exon 19 of COL1A2 while her parents were heterozygous for this mutation. This study reports the eighth child with OI and the homozygous mutation in COL1A2; and the first two individuals with the heterozygous p.Gly337Ser mutation in COL1A2 causing an isolated DI without OI.
Assuntos
Colágeno Tipo I/genética , Mutação de Sentido Incorreto , Osteogênese Imperfeita/genética , Adolescente , Feminino , Heterozigoto , Homozigoto , Humanos , Osteogênese Imperfeita/patologia , LinhagemRESUMO
BACKGROUND: Resistance of cholangiocarcinoma to chemotherapy is a major problem in cancer treatment. The mechanism of resistance is believed to involve phosphoinositide-3- kinase (PI3K)/Akt activation. Although the platinum-containing compound oxaliplatin has been extensively used in the treatment of several solid tumors, recent data regarding its use to treat cholangiocarcinoma are ambiguous. Oxaliplatin resistance in this disease could potentially involve PI3K pathways. We, therefore, examined the effects of PI3K pathways in cholangiocarcinoma cells in modulating oxaliplatin resistance. RESULTS: After exposing the cholangiocarcinoma cell lines RMCCA1 and KKU100 to oxaliplatin, the levels of Akt and mTOR phosphorylation increased, as shown by western blot analysis. The WST-1 cell proliferation assay showed increased inhibition of cell growth under high concentrations of oxaliplatin. The combination of oxaliplatin with LY294002, an inhibitor of PI3K, resulted in a remarkable arrest of cell proliferation. Deactivation of mTOR by RAD001 was also synergistic with oxaliplatin, although to a lesser extent. The combination of oxaliplatin and a PI3K inhibitor also resulted in a significant induction of apoptosis, as demonstrated by the TUNEL assay. CONCLUSION: Activation of PI3K might protect cholangiocarcinoma cells from oxaliplatininduced cytotoxicity. Although the inhibition of PI3K and the inhibition of mTOR both enhance oxaliplatin-induced cytotoxicity, PI3K inhibition has a greater effect. Targeting the PI3K pathway may be a useful approach to improve the chemotherapeutic sensitivity of cholangiocarcinoma.
RESUMO
Colorectal adenomas are precursor lesions of colorectal adenocarcinoma. The transition from adenoma to carcinoma in patients with colorectal cancer (CRC) has been associated with an accumulation of genetic aberrations. However, criteria that can screen adenoma progression to adenocarcinoma are still lacking. This present study is the first attempt to identify genetic aberrations, such as the somatic mutations, copy number variations (CNVs), and high-frequency mutated genes, found in Thai patients. In this study, we identified the genomic abnormality of two sample groups. In the first group, five cases matched normal-colorectal adenoma-colorectal adenocarcinoma. In the second group, six cases matched normal-colorectal adenomas. For both groups, whole-exome sequencing was performed. We compared the genetic aberration of the two sample groups. In both normal tissues compared with colorectal adenoma and colorectal adenocarcinoma analyses, somatic mutations were observed in the tumor suppressor gene APC (Adenomatous polyposis coli) in eight out of ten patients. In the group of normal tissue comparison with colorectal adenoma tissue, somatic mutations were also detected in Catenin Beta 1 (CTNNB1), Family With Sequence Similarity 123B (FAM123B), F-Box And WD Repeat Domain Containing 7 (FBXW7), Sex-Determining Region Y-Box 9 (SOX9), Low-Density Lipoprotein Receptor-Related Protein 5 (LRP5), Frizzled Class Receptor 10 (FZD10), and AT-Rich Interaction Domain 1A (ARID1A) genes, which are involved in the Wingless-related integration site (Wnt) signaling pathway. In the normal tissue comparison with colorectal adenocarcinoma tissue, Kirsten retrovirus-associated DNA sequences (KRAS), Tumor Protein 53 (TP53), and Ataxia-Telangiectasia Mutated (ATM) genes are found in the receptor tyrosine kinase-RAS (RTK-RAS) signaling pathway and p53 signaling pathway, respectively. These results suggest that APC and TP53 may act as a potential screening marker for colorectal adenoma and early-stage CRC. This preliminary study may help identify patients with adenoma and early-stage CRC and may aid in establishing prevention and surveillance strategies to reduce the incidence of CRC.
RESUMO
Due to the invasive procedure associated with Pap smears for diagnosing cervical cancer and the conservative culture of developing countries, identifying less invasive biomarkers is of great interest. Quantitative label-free mass spectrometry was performed to identify potential biomarkers in the urine samples of patients with cervical cancer. This technique was used to study the differential expression of urinary proteomes between normal individuals and cancer patients. The alterations in the levels of urinary proteomes in normal and cancer patients were analyzed by Progenesis label-free software and the results revealed that 60 proteins were upregulated while 73 proteins were downregulated in patients with cervical cancer. This method could enrich high molecular weight proteins from 100 kDa. The protein-protein interactions were obtained by Search Tool for the Retrieval of Interacting Genes/Proteins analysis and predicted the biological pathways involving various functions including cell-cell adhesion, blood coagulation, metabolic processes, stress response and the regulation of morphogenesis. Two notable upregulated urinary proteins were leucine-rich α-2-glycoprotein (LRG1) and isoform-1 of multimerin-1 (MMRN1), while the 3 notable downregulated proteins were S100 calcium-binding protein A8 (S100A8), serpin B3 (SERPINB3) and cluster of differentiation-44 antigen (CD44). The validation of these 5 proteins was performed by western blot analysis and the biomarker sensitivity of these proteins was analyzed individually and in combination with receiver operator characteristic curve (ROC) analysis. Quantitative mass spectrometry analysis may allow for the identification of urinary proteins of high molecular weight. The proteins MMRN1 and LRG1 were presented, for the first time, to be highly expressed urinary proteins in cervical cancer. ROC analysis revealed that LRG1 and SERPINB3 could be individually used, and these 5 proteins could also be combined, to detect the occurrence of cervical cancer.
RESUMO
Human telomerase consisting of telomerase RNA template (hTR) and telomerase reverse transcriptase (hTERT) provides a mechanism for synthesis of telomere repeats that prolongs life span of cells. Telomerase activity is present in germ-line and malignant tumor cells but not in most normal human somatic cells. This study determined hTERT mRNA level in tissue samples from patients with gastrointestinal tract (GI) cancers. Tissue samples were obtained from 22 GI cancer patients, 3 gastrointestinal stomal tumors (GIST) and 25 corresponding non-cancerous tissues. hTERT expression was determined by real-time reverse transcriptase-polymerase chain reaction (RT-PCR) using Taqman probe, hTERT mRNA was detected in 12 of 22 cancerous tissue samples. Six of 8 tissue samples obtained from patients with hepatocellular carcinoma and cholangiocarcinoma were positive for hTERT. However, hTERT mRNA was not detected in GIST and non-cancerous tissues. These results suggest that hTERT may be an effective target for cancer therapies to treat many type of GI cancers including cholangiocarcinoma and hepatocellular carcinoma.
Assuntos
Carcinoma Hepatocelular/genética , Colangiocarcinoma/genética , Neoplasias Gastrointestinais/genética , RNA/metabolismo , Telomerase/metabolismo , Carcinoma Hepatocelular/enzimologia , Colangiocarcinoma/enzimologia , Neoplasias Gastrointestinais/enzimologia , HumanosRESUMO
BACKGROUND: Cervical cancer is the major cause of morbidity and mortality in Thai women. Nevertheless, the preventive strategy such as HPV vaccination program has not been implemented at the national level. This study explored the HPV prevalence and genotypic distribution in a large cohort of Thai women. METHODS: A hospital-based cervical cancer screening program at Chulabhorn Hospital, Bangkok and a population-based screening program at a rural Pathum Thani Province were conducted using liquid-based cytology and HPV genotyping. RESULTS: Of 5906 women aged 20-70 years, Pap smear was abnormal in 4.9% and the overall HPV prevalence was 15.1%, with 6.4% high-risk (HR), 3.5% probable high-risk (PR), and 8.4% low-risk (LR) HPV. The prevalence and genotypic distribution were not significantly different between the two cohorts. Among HR-HPV genotypes, HPV52 was the most frequent (1.6%), followed by HPV16 (1.4%), HPV51 (0.9%), HPV58 (0.8%), HPV18 (0.6%), and HPV39 (0.6%). Among LR-HPV genotypes, HPV72 and HPV62 were the most frequent while HPV6 and HPV11 were rare. HPV infection was found to be proportionately high in young women, aged 20-30 years (25%) and decreasing with age (11% in women aged >50). The more severe abnormal cytology results, the higher positivity of HR-HPV infection was observed. CONCLUSIONS: In conclusion, HPV52, HPV16, and HPV51 were identified as the most common HR-HPV genotypes in Thai women. This study contributes genotypic evidence that should be essential for the development of appropriate HPV vaccination program as part of Thailand's cervical cancer prevention strategies.
RESUMO
AIM: To determine the role of circulating tumor cells (CTCs) in prediction of the overall survival of patients with advanced malignant biliary tract obstruction. METHODS: We investigated the prognostic value of CTCs by examining two markers, cytokeratin (CK) 19 and human telomerase reverse transcriptase (hTERT) mRNA, in 40 patients diagnosed with advanced malignant biliary tract diseases. Quantitative real-time reverse transcription polymerase chain reaction was used to detect CK19 and hTERT mRNA in the peripheral blood of these patients. Overall survival was analyzed using the Kaplan-Meier method and Cox regression modeling. RESULTS: Positive CK19 and hTERT mRNA expression was detected in 45% and 60%, respectively, of the 40 patients. Univariable analysis indicated that positive CK19 mRNA expression was significantly associated with worse overall survival (P = 0.009). Multivariable analysis determined that positive CK19 mRNA expression, patient's age and serum bilirubin were each independently associated with overall survival. CONCLUSION: CK19 mRNA expression levels in peripheral blood appear to provide a valuable marker to predict the overall survival of patients with advanced malignant biliary tract obstruction.
Assuntos
Doenças Biliares , Biomarcadores Tumorais , Queratina-19 , Células Neoplásicas Circulantes/metabolismo , Telomerase , Adulto , Idoso , Idoso de 80 Anos ou mais , Doenças Biliares/sangue , Doenças Biliares/genética , Doenças Biliares/patologia , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Feminino , Humanos , Estimativa de Kaplan-Meier , Queratina-19/sangue , Queratina-19/genética , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , Análise de Sobrevida , Telomerase/sangue , Telomerase/genéticaRESUMO
PURPOSE: Hilar cholangiocarcinoma (Klatskin tumor) is one of the most difficult cancers to treat. We demonstrate activation of phosphoinositide-3-kinase (PI3K)/Akt signaling, which is a critical pathway for cell survival, in hilar cholangiocarcinoma cells. However, inhibition of PI3K has little effect on hilar cholangiocarcinoma cell survival. In this study, we investigated the mechanism by which hilar cholangiocarcinoma cells resist PI3K inhibitors. METHODS: Human hilar cholangiocarcinoma cells KKU-100 were treated with PI3K inhibitors, and cell viability and apoptosis assays were performed. The expression of a MAPK phosphatase (MKP-1) that contributes to cancer cell survival in response to multiple stress stimuli was assayed by quantitative real-time RT-PCR and western blotting. In addition, the effects of the MKP-1 inhibitor were studied in KKU-100 cells treated with PI3K inhibitors. RESULTS: Incubation of KKU-100 cells with PI3K inhibitors resulted in increased expression of MKP-1. Furthermore, we found that inhibition of MKP-1 using siRNA silencing sensitized KKU-100 cells to PI3K inhibitor-induced apoptosis via increased phosphorylation of p38 MAPK. CONCLUSIONS: These results indicate that concurrent inhibition of PI3K and MKP-1 induces apoptosis in KKU-100 cells. Simultaneous targeting of the PI3K pathway and MKP-1 may be a useful approach to improve therapies directed against hilar cholangiocarcinoma.
Assuntos
Neoplasias dos Ductos Biliares/tratamento farmacológico , Ductos Biliares Intra-Hepáticos , Colangiocarcinoma/tratamento farmacológico , Fosfatase 1 de Especificidade Dupla/fisiologia , Inibidores de Fosfoinositídeo-3 Quinase , Neoplasias dos Ductos Biliares/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Colangiocarcinoma/patologia , Cromonas/farmacologia , Cicloeximida/farmacologia , Fosfatase 1 de Especificidade Dupla/genética , Humanos , Morfolinas/farmacologia , Fosfatidilinositol 3-Quinases/fisiologia , Fosforilação , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Compound heterozygous anion exchanger 1 (AE1) SAO/G701D mutations result in distal renal tubular acidosis with Southeast Asian ovalocytosis. Interaction, trafficking and localization of wild-type and mutant (SAO and G701D) kAE1 proteins fused with hemagglutinin, six-histidine, Myc, or green fluorescence protein (GFP) were examined in human embryonic kidney (HEK) 293 cells. When individually expressed, wild-type kAE1 was localized at cell surface while mutant kAE1 SAO and G701D were intracellularly retained. When co-expressed, wild-type kAE1 could form heterodimer with kAE1 SAO or kAE1 G701D and could rescue mutant kAE1 proteins to express on the cell surface. Co-expression of kAE1 SAO and kAE1 G701D also resulted in heterodimer formation but intracellular retention without cell surface expression, suggesting their trafficking defect and failure to rescue each other to the plasma membrane, most likely the molecular mechanism of the disease in the compound heterozygous condition.
Assuntos
Acidose Tubular Renal/metabolismo , Proteína 1 de Troca de Ânion do Eritrócito/metabolismo , Eliptocitose Hereditária/metabolismo , Rim/metabolismo , Acidose Tubular Renal/genética , Proteína 1 de Troca de Ânion do Eritrócito/genética , Sudeste Asiático , Eliptocitose Hereditária/genética , Humanos , Mutagênese Sítio-Dirigida , Mutação , Transporte Proteico/genética , Distribuição TecidualRESUMO
Human telomerase consisting of telomerase RNA template (hTR) and telomerase reverse transcriptase (hTERT) provides a mechanism for synthesis of telomere repeats that prolongs life span of cells. Telomerase activity is present in germ-line and malignant tumor cells but not in most normal human somatic cells. This study determined hTERT mRNA level in tissue samples from patients with gastrointestinal tract (GI) cancers. Tissue samples were obtained from 22 GI cancer patients, 3 gastrointestinal stomal tumors (GIST) and 25 corresponding non-cancerous tissues. hTERT expression was determined by real-time reverse transcriptase-polymerase chain reaction (RT-PCR) using Taqman probe, hTERT mRNA was detected in 12 of 22 cancerous tissue samples. Six of 8 tissue samples obtained from patients with hepatocellular carcinoma and cholangiocarcinoma were positive for hTERT. However, hTERT mRNA was not detected in GIST and non-cancerous tissues. These results suggest that hTERT may be an effective target for cancer therapies to treat many type of GI cancers including cholangiocarcinoma and hepatocellular carcinoma.