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Drug resistance is a major challenge in the treatment of advanced cholangiocarcinoma (CCA). Understanding the mechanisms of drug resistance can aid in identifying novel prognostic biomarkers and therapeutic targets to improve treatment efficacy. This study established 5-fluorouracil- (5-FU) and gemcitabine-resistant CCA cell lines, KKU-213FR and KKU-213GR, and utilized comparative proteomics to identify differentially expressed proteins in drug-resistant cells compared to parental cells. Additionally, bioinformatics analyses were conducted to explore the biological and clinical significance of key proteins. The drug-resistant phenotypes of KKU-213FR and KKU-213GR cell lines were confirmed. In addition, these cells demonstrated increased migration and invasion abilities. Proteomics analysis identified 81 differentially expressed proteins in drug-resistant cells, primarily related to binding functions, biological regulation, and metabolic processes. Protein–protein interaction analysis revealed a highly interconnected network involving MET, LAMB1, ITGA3, NOTCH2, CDH2, and NDRG1. siRNA-mediated knockdown of these genes in drug-resistant cell lines attenuated cell migration and cell invasion abilities and increased sensitivity to 5-FU and gemcitabine. The mRNA expression of these genes is upregulated in CCA patient samples and is associated with poor prognosis in gastrointestinal cancers. Furthermore, the functions of these proteins are closely related to the epithelial–mesenchymal transition (EMT) pathway. These findings elucidate the potential molecular mechanisms underlying drug resistance and tumor progression in CCA, providing insights into potential therapeutic targets. [ABSTRACT FROM AUTHOR]

Cholangiocarcinoma (CCA) is an aggressive cancer originating from bile duct epithelium, particularly prevalent in Asian countries with liver fluke infections. Current chemotherapy for CCA often fails due to drug resistance, necessitating novel anticancer agents. This study investigates the potential of 5'-deoxy-5'-methylthioadenosine (MTA), a naturally occurring nucleoside, against CCA. While MTA has shown promise against various cancers, its effects on CCA remain unexplored. We evaluated MTA's anticancer activity in CCA cell lines and drug-resistant sub-lines, assessing cell viability, migration, invasion, and apoptosis. The potential anticancer mechanisms of MTA were explored through proteomic analysis using LC-MS/MS and bioinformatic analysis. The results show a dose-dependent reduction in CCA cell viability, with enhanced effects on cancer cells compared to normal cells. Moreover, MTA inhibits growth, induces apoptosis, and suppresses cell migration and invasion. Additionally, MTA enhanced the anticancer effects of gemcitabine on drug-resistant CCA cells. Proteomics revealed the down-regulation of multiple proteins by MTA, affecting various molecular functions, biological processes, and cellular components. Network analysis highlighted MTA's role in inhibiting proteins related to mitochondrial function and energy derivation, crucial for cell growth and survival. Additionally, MTA suppressed proteins involved in cell morphology and cytoskeleton organization, important for cancer cell motility and metastasis. Six candidate genes, including ZNF860, KLC1, GRAMD1C, MAMSTR, TANC1, and TTC13, were selected from the top 10 most down-regulated proteins identified in the proteomics results and were subsequently verified through RT-qPCR. Further, KLC1 protein suppression by MTA treatment was confirmed through Western blotting. Additionally, based on TCGA data, KLC1 mRNA was found to be upregulated in the tissue of CCA patients compared to that of normal adjacent tissues. In summary, MTA shows promising anticancer potential against CCA by inhibiting growth, inducing apoptosis, and suppressing migration and invasion, while enhancing gemcitabine's effects. Proteomic analysis elucidates possible molecular mechanisms underlying MTA's anticancer activity, laying the groundwork for future research and development of MTA as a treatment for advanced CCA. [ABSTRACT FROM AUTHOR]

Cholangiocarcinoma (CCA) is an extremely aggressive malignancy arising from the epithelial cells lining the bile ducts. It presents a substantial global health issue, with the highest incidence rates, ranging from 40–100 cases/100,000 individuals, found in Southeast Asia, where liver fluke infection is endemic. In Europe and America, incidence rates range from 0.4–2 cases/100,000 individuals. Globally, mortality rates range from 0.2–2 deaths/100,000 person-years and are increasing in most countries. Chemotherapy is the primary treatment for advanced CCA due to limited options from late-stage diagnosis, but its efficacy is hindered by drug-resistant phenotypes. In a previous study, proteomics analysis of drug-resistant CCA cell lines (KKU-213A-FR and KKU-213A-GR) and the parental KKU-213A line identified cullin 3 (Cul3) as markedly overexpressed in drug-resistant cells. Cul3, a scaffold protein within CUL3-RING ubiquitin ligase complexes, is crucial for ubiquitination and proteasome degradation, yet its role in drug-resistant CCA remains to be elucidated. The present study aimed to elucidate the role of Cul3 in drug-resistant CCA cell lines. Reverse transcription-quantitative PCR and western blot analyses confirmed significantly elevated Cul3 mRNA and protein levels in drug-resistant cell lines compared with the parental control. Short interfering RNA-mediated Cul3 knockdown sensitized cells to 5-fluorouracil and gemcitabine and inhibited cell proliferation, colony formation, migration and invasion. In addition, Cul3 knockdown induced G0/G1 cell cycle arrest and suppressed key cell cycle regulatory proteins, cyclin D, cyclin-dependent kinase (CDK)4 and CDK6. Bioinformatics analysis of CCA patient samples using The Cancer Genome Atlas data revealed Cul3 upregulation in CCA tissues compared with normal bile duct tissues. STRING analysis of upregulated proteins in drug-resistant CCA cell lines identified a highly interactive Cul3 network, including COMM Domain Containing 3, Ariadne RBR E3 ubiquitin protein ligase 1, Egl nine homolog 1, Proteasome 26S Subunit Non-ATPase 13, DExH-box helicase 9 and small nuclear ribonucleoprotein polypeptide G, which showed a positive correlation with Cul3 in CCA tissues. Knocking down Cul3 significantly suppressed the mRNA expression of these genes, suggesting that Cul3 may act as an upstream regulator of them. Gene Ontology analysis revealed that the majority of these genes were categorized under binding function, metabolic process, cellular anatomical entity, protein-containing complex and protein-modifying enzyme. Taken together, these findings highlighted the biological and clinical significance of Cul3 in drug resistance and progression of CCA. [ABSTRACT FROM AUTHOR]

Flooding in rainfed lowlands greatly impairs the mutualistic relationship between indigenous arbuscular mycorrhizal fungi (AMF) and rice. In flooded soils, root colonization by AMF is arrested, but some AMF genera, defined as the core AMF, remain present. However, the core AMF in rainfed lowlands and their symbiotic roles remain unknown. Here, we showed that Acaulospora fungi were the core AMF in rice seedling roots of the Sangyod Muang Phatthalung (SMP) landrace rice variety grown in non-flooded and flooded paddy soils. Subsequently, indigenous Acaulospora spores were propagated by trap cultures using maize as the host plants. Therefore, to clarify the roles of cultured Acaulospora spores in a symbiotic partnership, the model japonica rice variety Nipponbare was grown in sterile soil inoculated with Acaulospora spores, and recolonized with a native microbial filtrate from the organic rice paddy soil. Our data demonstrated that the inoculation of Acaulospora spores in well-drained soil under a nutrient-sufficient condition for six weeks enabled 70 percent of the rice roots to be colonized by the fungi, leading to higher phosphate (Pi) accumulation in the mycorrhizal roots. Unexpectedly, the growth of rice seedlings was significantly suppressed by inoculation while photosynthetic parameters such as fractions of incoming light energy and relative chlorophyll content were unaltered. In the soil, the Acaulospora fungi increased soil phosphorus (P) availability by enhancing the secretion of acid phosphatase in the mycorrhizal roots. The findings of this work elucidate the symbiotic roles of the dominant Acaulospora fungi from lowland rice paddies. [ABSTRACT FROM AUTHOR]