TC2N inhibits distant metastasis and stemness of breast cancer via blocking fatty acid synthesis.

BACKGROUND: Tandem C2 domains, nuclear (TC2N) is a C2 domain-containing protein that belongs to the carboxyl-terminal type (C-type) tandem C2 protein family, and acts as an oncogenic driver in several cancers. Previously, we preliminarily reported that TC2N mediates the PI3K-Akt signaling pathway to inhibit tumor growth of breast cancer (BC) cells. Beyond that, its precise biological functions and detailed molecular mechanisms in BC development and progression are not fully understood. METHODS: Tumor tissues of 212 BC patients were subjected to tissue microarray and further assessed the associations of TC2N expression with pathological parameters and FASN expression. The protein levels of TC2N and FASN in cell lines and tumor specimens were monitored by qRT-PCR, WB, immunofluorescence and immunohistochemistry. In vitro cell assays, in vivo nude mice model was used to assess the effect of TC2N ectopic expression on tumor metastasis and stemness of breast cancer cells. The downstream signaling pathway or target molecule of TC2N was mined using a combination of transcriptomics, proteomics and lipidomics, and the underlying mechanism was explored by WB and co-IP assays. RESULTS: Here, we found that the expression of TC2N remarkedly silenced in metastatic and poorly differentiated tumors. Function-wide, TC2N strongly inhibits tumor metastasis and stem-like properties of BC via inhibition of fatty acid synthesis. Mechanism-wise, TC2N blocks neddylated PTEN-mediated FASN stabilization by a dual mechanism. The C2B domain is crucial for nuclear localization of TC2N, further consolidating the TRIM21-mediated ubiquitylation and degradation of FASN by competing with neddylated PTEN for binding to FASN in nucleus. On the other hand, cytoplasmic TC2N interacts with import proteins, thereby restraining nuclear import of PTEN to decrease neddylated PTEN level. CONCLUSIONS: Altogether, we demonstrate a previously unidentified role and mechanism of TC2N in regulation of lipid metabolism and PTEN neddylation, providing a potential therapeutic target for anti-cancer.

Development of a nomogram for prediction of central lymph node metastasis of papillary thyroid microcarcinoma.

BACKGROUND: Papillary thyroid carcinoma (PTC) is the most frequent malignant tumor in thyroid carcinoma. The aim of this study was to explore the risk factors associated with central lymph node metastasis in papillary thyroid microcarcinoma (PTMC) and establish a nomogram model that can assess the probability of central lymph node metastasis (CLNM). METHODS: The clinicopathological data of 377 patients with cN0 PTMC were collected and analyzed from The Second Affiliated Hospital of Fujian Medical University from July 1st, 2019 to December 30th, 2021. All patients were examined by underwent ultrasound (US), found without metastasis to central lymph nodes, and diagnosed with PTMC through pathologic examination. All patients received thyroid lobectomy or total thyroidectomy with therapeutic or prophylactic central lymph node dissection (CLND). R software (Version 4.1.0) was employed to conduct a series of statistical analyses and establish the nomogram. RESULTS: A total of 119 patients with PTMC had central lymph node metastases (31.56%). After that, age (P < 0.05), gender (P < 0.05), tumor size (P < 0.05), tumor multifocality (P < 0.05), and ultrasound imaging-suggested tumor boundaries (P < 0.05) were identified as the risk factors associated with CLNM. Subsequently, multivariate logistic regression analysis indicated that the area under the receiver operating characteristic (ROC) curve (AUC) of the training cohort was 0.703 and that of the validation cohort was 0.656, demonstrating that the prediction ability of this model is relatively good compared to existing models. The calibration curves indicated a good fit for the nomogram model. Finally, the decision curve analysis (DCA) showed that a probability threshold of 0.15-0.50 could benefit patients clinically. The probability threshold used in DCA captures the relative value the patient places on receiving treatment for the disease, if present, compared to the value of avoiding treatment if the disease is not present. CONCLUSION: CLNM is associated with many risk factors, including age, gender, tumor size, tumor multifocality, and ultrasound imaging-suggested tumor boundaries. The nomogram established in our study has moderate predictive ability for CLNM and can be applied to the clinical management of patients with PTMC. Our findings will provide a better preoperative assessment and treatment strategies for patients with PTMC whether to undergo central lymph node dissection.

Metabolic reprogramming-associated genes predict overall survival for rectal cancer.

Metabolic reprogramming has become a hot topic recently in the regulation of tumour biology. Although hundreds of altered metabolic genes have been reported to be associated with tumour development and progression, the important prognostic role of these metabolic genes remains unknown. We downloaded messenger RNA expression profiles and clinicopathological data from The Cancer Genome Atlas and the Gene Expression Omnibus database to uncover the prognostic role of these metabolic genes. Univariate Cox regression analysis and lasso Cox regression model were utilized in this study to screen prognostic associated metabolic genes. Patients with high-risk demonstrated significantly poorer survival outcomes than patients with low-risk in the TCGA database. Also, patients with high-risk still showed significantly poorer survival outcomes than patients with low-risk in the GEO database. What is more, gene set enrichment analyses were performed in this study to uncover significantly enriched GO terms and pathways in order to help identify potential underlying mechanisms. Our study identified some survival-related metabolic genes for rectal cancer prognosis prediction. These genes might play essential roles in the regulation of metabolic microenvironment and in providing significant potential biomarkers in metabolic treatment.

Rab7 Is Associated with Poor Prognosis of Gastric Cancer and Promotes Proliferation, Invasion, and Migration of Gastric Cancer Cells.

BACKGROUND Rab7 belongs to the Ras oncogene family. Many studies have shown that its dysfunction is associated with many types of malignant tumors, but its effect on the pathogenesis of gastric cancer (GC) is still unknown. Therefore, we investigated the effect and mechanism of Rab7 in GC. MATERIAL AND METHODS The expression of Rab7 in GC and adjacent tissues was detected by immunohistochemistry, Western blot analysis, and qRT-PCR. The relationship of Rab7 with clinicopathological parameters and prognosis was analyzed. The expressions of Rab7, PI3K, and AKT in GC cells were assessed by Western blot. Overexpressed and silenced GC cell lines were constructed and AGS cells were treated with LY294002. The proliferation capacity of GC cells was detected by CCK8 assay, cell cycle changes were detected by flow cytometry, and the invasion and migration abilities of GC cells were assessed by transwell assay. RESULTS The expression of Rab7 was upregulated in the samples and cells, and was positively correlated with lymph node metastasis but negatively correlated with histological differentiation and clinical prognosis. In cell function experiments, overexpression of Rab7 induced the transition from S phase to G2 phase and promoted the proliferation, invasion, and migration of GC cells. Our assessment of the molecular mechanism showed that Rab7 promoted the phosphorylation of PI3K and AKT in GC cells. Incubation with the PI3K inhibitor Ly294002 impaired the enhanced effect of Rab7 overexpression on proliferation, migration, and invasion abilities of GC cells. These results show that the Rab7 affects GC cell progression by modulating the PI3K/AKT pathway. CONCLUSIONS Rab7 could be a prognostic biomarker and therapeutic target of the PI3K/AKT pathway in GC.

Chronic Microcystin-LR Exposure Induces Hepatocarcinogenesis via Increased Gankyrin in Vitro and in Vivo.

BACKGROUND/AIMS: Our recent study indicated that the serum microcystin-LR (MC-LR) level is positively linked to the risk of human hepatocellular carcinoma (HCC). Gankyrin is over-expressed in cancers and mediates oncogenesis; however, whether MC-LR induces tumor formation and the role of gankyrin in this process is unclear. METHODS: We induced malignant transformation of L02 liver cells via 35 passages with exposure to 1, 10, or 100 nM MC-LR. Wound healing, plate and soft agar colony counts, and nude mice tumor formation were used to evaluate the tumorigenic phenotype of MC-LR-treated cells. Silencing gankyrin was used to confirm its function. We established a 35-week MC-LR exposure rat model by twice weekly intraperitoneal injection with 10 µg/kg body weight. In addition, 96 HCC patients were tested for tumor tissue gankyrin expression and serum MC-LR levels. RESULTS: Chronic low-dose MC-LR exposure increased proliferation, mobility, clone and tumor formation abilities of L02 cells as a result of gankyrin activation, while silencing gankyrin inhibited the carcinogenic phenotype of MC-LR-treated cells. MC-LR also induced neoplastic liver lesions in Sprague-Dawley rats due to up-regulated gankyrin. Furthermore, a trend of increased gankyrin was observed in humans exposed to MC-LR. CONCLUSION: These results suggest that MC-LR induces hepatocarcinogenesis in vitro and in vivo by increasing gankyrin levels, providing new insight into MC-LR carcinogenicity studies.

Intestinal T-cell and NK/T-cell lymphomas: A clinicopathological study of 27 Chinese patients.

BACKGROUND: Intestinal T-cell and NK/T- cell lymphomas are rare and aggressive. The diagnosis is quite difficult, especial in biopsy specimens. This study investigates the clinicopathological features of intestinal T-cell and NK/T-cell lymphomas to aid their differential diagnosis. METHODS: Clinical data of 27 cases were collected. Including extranodal NK/T-cell lymphoma, nasal type (ENKTCL-N), monomorphic epitheliotropic intestinal T-cell lymphoma (MEITL), peripheral T-cell lymphoma, not otherwise specified (PTCL, NOS), anaplastic large-cell lymphoma, ALK+ (ALCL, ALK+) and angioimmunoblastic T-cell lymphoma (AITL). The histologic features, immunohistochemical findings, T-cell receptor gene rearrangement results, and follow-up data were analyzed, with review of literature. RESULTS: The age of the patients (N = 27) was 15-85 years (mean, 47.5 years), and male:female ratio, 3.5:1. Abdominal pain and B symptoms were the most common symptoms. Although 85.2% of the patients were in clinical stage I-II, 59.3% died within 1 year. MEITL showed certain distinctive clinic opathological features from ENKTCL-N. Compared to lesions at other sites, there were no differences in the morphological features, immunophenotype and TCR gene rearrangement of intestinal ENKTCL-N, PTCL, NOS, ALCL, ALK+ and AITL. CONCLUSION: Intestinal T-cell and NK/T-cell lymphomas are a heterogeneous group of lymphomas. They could be classified to 5 histological subtypes in our study. ENKTCL-N and MEITL formed the majority of the tumor types. Each subtype has distinctive pathological features, but most of them have diamal prognosis.

The risk of new-onset cancer associated with HFE C282Y and H63D mutations: evidence from 87,028 participants.

To investigate the association between mutation of HFE (the principal pathogenic gene in hereditary haemochromatosis) and risk of cancer, we conducted a meta-analysis of all available case-control or cohort studies relating to two missense mutations, C282Y and H63D mutations. Eligible studies were identified by searching databases including PubMed, Embase and the ISI Web of Knowledge. Overall and subgroup analyses were performed and odds ratios (ORs) combined with 95% confidence intervals (CIs) were applied to evaluate the association between C282Y mutation, H63D mutation and cancer risk. Sensitivity and cumulative analyses were used to evaluate the stability of the results. A total of 36 eligible studies were included, comprising 13,680 cases and 73,348 controls. C282Y was significantly associated with elevated cancer risk in a recessive genetic model (OR: 1.991, 95% CI: 1.448-2.737). On subgroup analysis stratified by cancer type, statistically significantly increased cancer risks were found for breast cancer, colorectal cancer and hepatocellular carcinoma in a recessive model. When stratified by territory, a significantly increased risk of cancer was found in Oceanic populations in a recessive model and in Asian populations in an allele model and dominant model. H63D mutation did not significantly increase overall cancer risk in any genetic model. However, when, stratified by territory, an increased cancer risk was found in the Asian population in an allele and dominant. C282Y but not H63D mutation was related to elevated cancer risk. Further large-scale studies considering gene-environment interactions and functional research should be conducted to further investigate this association.

Preliminary screening of differentially expressed genes involved in methyl-CpG-binding protein 2 gene-mediated proliferation in human osteosarcoma cells.

Methyl-CpG-binding protein 2 (MeCP2) is essential in human brain development and has been linked to several cancer types and neuro-developmental disorders. This study aims to screen the MeCP2 related differentially expressed genes and discover the therapeutic targets for osteosarcoma. CCK8 assay was used to detect the proliferation and SaOS2 and U2OS cells. Apoptosis of cells was detected by flow cytometry analysis that monitored Annexin V-APC/7-DD binding and 7-ADD uptake simultaneously. Denaturing formaldehyde agarose gel electrophoresis was employed to examine the quality of total RNA 18S and 28S units. Gene chip technique was utilized to discover the differentially expressed genes correlated with MeCP2 gene. Differential gene screening criteria were used to screen the changed genes. The gene up-regulation or down-regulation more than 1.5 times was regarded as significant differential expression genes. The CCK8 results indicated that the cell proliferation of MeCP2 silencing cells (LV-MeCP2-RNAi) was significantly decreased compared to non-silenced cells (LV-MeCP2-RNAi-CN) (P < 0.05). MeCP2 silencing could also induce significant apoptosis compared to non-silenced cells (P < 0.05); 107 expression changed genes were screened from a total of 49,395 transcripts. Among the total 107 transcripts, 34 transcripts were up-regulated and 73 transcripts were down-regulated. There were five significant differentially expressed genes, including IGFBP4, HOXC8, LMO4, MDK, and CTGF, which correlated with the MeCP2 gene. The methylation frequency of CpG in IGFBP4 gene could achieve 55%. In conclusion, the differentially expressed IGFBP4, HOXC8, LMO4, MDK, and CTGF genes may be involved in MeCP2 gene-mediated proliferation and apoptosis in osteosarcoma cells.

Endothelial cells promote stem-like phenotype of glioma cells through activating the Hedgehog pathway.

Microenvironmental regulation of cancer stem cells (CSCs) strongly influences the onset and spread of cancer. The way in which glioma cells interact with their microenvironment and acquire the phenotypes of CSCs remains elusive. We investigated how communication between vascular endothelial cells and glioma cells promoted the properties of glioma stem cells (GSCs). We observed that CD133(+) GSCs were located closely to Shh(+) endothelial cells in specimens of human glioblastoma multiforme (GBM). In both in vitro and in vivo studies, we found that endothelial cells promoted the appearance of CSC-like glioma cells, as demonstrated by increases in tumourigenicity and expression of stemness genes such as Sox2, Olig2, Bmi1 and CD133 in glioma cells that were co-cultured with endothelial cells. Knockdown of Smo in glioma cells led to a significant reduction of their CSC-like phenotype formation in vitro and in vivo. Endothelial cells with Shh knockdown failed to promote Hedgehog (HH) pathway activation and CSC-like phenotype formation in co-cultured glioma cells. By examination of glioma tissue specimens from 65 patients, we found that the survival of glioma patients was closely correlated with the expression of both Shh by endothelial cells and Gli1 by perivascular glioma cells. Taken together, our study demonstrates that endothelial cells in the tumour microenvironment provide Shh to activate the HH signalling pathway in glioma cells, thereby promoting GSC properties and glioma propagation.

Epigenetic silencing of methyl-CpG-binding protein 2 gene affects proliferation, invasion, migration, and apoptosis of human osteosarcoma cells.

Methyl-CpG-binding protein 2 (MeCP2) is a DNA methylation-related gene of the methyl-CpG-binding protein family. Here, we investigated the epigenetic function of the MeCP2 in SaOS2 and U2OS cell lines, and explored the antitumor effects of the gene silencing for osteosarcoma. In this study, chromatin immunoprecipitation assay was used to detect MeCP2 binding activity with TSSC3 gene. RT-PCR and western blot assay were used to analyze the MeCP2 expression in osteosarcoma cell lines after transfection with LV-MECP2-RNAi. Transwell invasion and migration assays were used to detect the cell invasion and migration. The cell apoptosis was examined by using the flow cytometry assay. The tumor size was also assessed to determine the therapeutic effects of gene silencing. The results indicated that MeCP2 indicated the highest combining power with TSSC3 gene. LV-MECP2-RNAi could decrease MeCP2 level in tumor cells compared with the untreated cells (P < 0.05). LV-MECP2-RNAi inhibited the U2OS and SaOS2 cells invasion and migration compared with the control cells (P < 0.05). LV-MECP2-RNAi triggered the U2OS and SaOS2 cell apoptosis, and inhibited the cell proliferation significantly compared with the control cells (P < 0.05). The gene silencing of RNAi could also decreased the tumor size significantly compared with untreated cells (P < 0.05). In conclusion, silencing the MeCP2 gene could block the MeCP2 expression and inhibit the tumor cell migration, invasion, and proliferation, and decreases the tumor size by inducing the apoptosis of the tumor cells.

Integrated machine learning algorithms identify KIF15 as a potential prognostic biomarker and correlated with stemness in triple-negative breast cancer.

Cancer stem cells (CSCs) have the potential to self-renew and induce cancer, which may contribute to a poor prognosis by enabling metastasis, recurrence, and therapy resistance. Hence, this study was performed to identify the association between CSC-related genes and triple-negative breast cancer (TNBC) development. Stemness gene sets were downloaded from StemChecker. Based on the online databases, a consensus clustering algorithm was conducted for unsupervised classification of TNBC samples. The variations between subtypes were assessed with regard to prognosis, tumor immune microenvironment (TIME), and chemotherapeutic sensitivity. The stemness-related gene signature was established and random survival forest analysis was employed to identify the core gene for validation experiments and tumor sphere formation assays. 499 patients with TNBC were classified into three subgroups and the Cluster 1 had a better OS than others. After that, WGCNA study was performed to identify genes important for Cluster 1 subtype. Out of all 8 modules, the subtype of Cluster 1 and the yellow module with 103 genes demonstrated the largest positive association. After that, a four-gene stemness-related signature was established. Based on the yellow module, the 39 potential pivotal genes were subjected to the random forest survival analysis to find out the gene that was relatively important for OS. KIF15 was confirmed as the targeted gene by LASSO and random survival forest analyses. In vitro experiments, the downregulation of KIF15 promoted the stemness of TNBC cells. The expression levels of stem cell markers Nanog, SOX2, and OCT4 were found to be elevated in TNBC cell lines after KIF15 inhibition. A stemness-associated risk model was constructed to forecast the clinical outcomes of TNBC patients. The downregulation of KIF15 expression in a subpopulation of TNBC stem cells may promote stemness and possibly TNBC progression.

Mutations in the mitochondrial ATPase6 gene are frequent in human osteosarcoma.

To explore the polymorphisms and mutations of mitochondrial ATPase6 gene in Chinese patients with osteosarcoma and their possible association with carcinogenesis, direct DNA sequencing method was used to detect the variants of the mitochondrial ATPase6 gene in 39 patients with osteosarcoma. We found mutations of the mitochondrial ATPase6 gene in 24/39 (61.5%) of the tested osteosarcoma samples, and identified 27 variant sites in ATPase6 coding regions. We did not detect any new polymorphisms in osteosarcoma, nor was there any association between variants and the three histopathological subtypes. These data demonstrated that mtDNA mutations within the ATPase6 gene are a frequent event in Chinese patients with osteosarcoma.

Expressions of some neurotrophins and neurotrophic cytokines at site of spinal cord injury in mice after vaccination with dendritic cells pulsed with homogenate proteins.

OBJECTIVE: Immune cells are key mediators of secondary damage following spinal cord injury (SCI), and dendritic cell (DC)-based vaccines have received considerable interest for treatment of SCI. We previously showed that vaccination with DCs pulsed with homogenate proteins of the spinal cord (hpDCs) promotes functional recovery from SCI in mice. However, the underlying molecular mechanisms remain unclear. Here, changes of neurotrophins, cytokines and T cells at the site of SCI in mice after vaccination with hpDCs were investigated and correlated with recovery from SCI. METHODS: hpDCs, DCs (control) or PBS (control) were injected intraperitoneally into injured mouse spinal cords. Functional recovery of the spinal cord was measured weekly using the Basso Mouse Scale (BMS) and confirmed by histological and immunohistochemical analysis. Brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), interleukin-4 (IL-4) and interferon-γ (IFN-γ) levels in T cell culture supernatants and spinal cord tissues were determined by ELISA. RESULTS: Eighty-four days after immunization, the BMS score of the hpDCs group (6.92 ± 0.20) was significantly higher than those of the DCs and PBS groups (p < 0.01). Meanwhile, the injury area and number of cysts in the hpDCs group decreased significantly compared with control groups. BDNF, NT-3, IL-4 and IFN-γ levels at the injured site as well as BDNF and NT-3 levels in the supernatant of cultured T cells from the hpDCs group were significantly higher than in control groups (p < 0.05). CONCLUSION: These results reveal that vaccination with hpDCs can promote SCI repair potentially by upregulating BDNF, NT-3, IL-4 and IFN-γ at the injury site.

Construction and validation of a transient receptor potential-related long noncoding RNA signature for prognosis prediction in breast cancer patients.

Breast cancer (BC) is the most commonly diagnosed malignancy in women around the world. Accumulating evidence suggests that transient receptor potential (TRP) channels play a significant role in tumor progression and immune cell infiltration. Hence, we conducted the study to investigate the correlation between TRP-associated lncRNAs and the prognosis of breast carcinoma. In the current study, 33 TRP-associated genes were selected from a review published by Amrita Samanta et al, and the TRP-related lncRNAs were identified by Pearson analysis. Based on the sum of the expression levels of 12 lncRNAs provided by the Cancer Genome Atlas (TCGA), a TRP-associated lncRNA signature was established by using Cox regression analysis. According to the median value of the risk score in the training set, BC patients were separated into high- and low-risk groups. Subsequently, functional enrichment analysis was conducted on the differential expression genes (DEGs) between different risk groups. The Estimation of Stromal and Immune Cells in Malignant Tumor Tissues Using Expression (ESTIMATE) Score was calculated by ESTIMATE, and the immune cell infiltration was evaluated by ssGSEA. Finally, the immune checkpoint gene expression levels, microsatellite instability (MSI), and immunophenoscore (IPS) were further assessed. The high-risk groups exhibited lower survival rates, while the low-risk groups showed higher survival rates. As a result, the DEGs between different risk groups were highly enriched in immune cell activation and immunoregulation. Besides, the ESTIMATE scores of patients in low-risk groups were higher than those in high-risk groups. The infiltration levels of several immune cells were remarkably elevated in low-risk groups, and various immune signatures were activated with a decreased risk score. Eventually, the TRP-associated lncRNA signature was confirmed with a highly potential ability to evaluate the immunotherapy response in breast carcinoma patients. The outcomes of the current study indicated that the 12-TRP-associated-lncRNA risk model was an independent prognostic risk factor for BC patients. This risk model could be closely related to the tumor immune microenvironment in BC. Our findings will provide new insights for future immunotherapy for BC treatment.

Identification of an exosome-related signature associated with prognosis and immune infiltration in breast cancer.

Exosomes, nanosized vesicles, play a vital role in breast cancer (BC) occurrence, development, and drug resistance. Hence, we proceeded to study the potential prognostic value of exosome-related genes and their relationship to the immune microenvironment in BC. 121 exosome-related genes were provided by the ExoBCD database, and 7 final genes were selected to construct the prognostic signature. Besides, the expression levels of the 7 exosome-related genes were validated by the experiment in BC cell lines. Based on the signature, BC patients from the training and validation cohorts were separated into low- and high-risk groups. Subsequently, the R clusterProfiler package was applied to identify the distinct enrichment pathways between high-risk groups and low-risk groups. The relevance of the tumor immune microenvironment and exosome-related gene risk score were analyzed in BC. Eventually, the different expression levels of immune checkpoint-related genes were compared between the two risk groups. Based on the risk model, the low-risk groups were identified with a higher survival rate both in the training and validation cohorts. A better overall survival was revealed in patients with higher scores evaluated by the estimation of stromal and immune cells in malignant tumor tissues using expression (ESTIMATE) algorithm. Subsequently, BC patients with lower risk scores were indicated by higher expression levels of some immune checkpoint-related genes and immune cell infiltration. Exosomes are closely associated with the prognosis and immune cell infiltration of BC. These findings may contribute to improving immunotherapy and provide a new vision for BC treatment strategies.

Natural killer cell-related prognostic risk model predicts prognosis and treatment outcomes in triple-negative breast cancer.

Background: Natural killer (NK) cells are crucial to the emergence, identification, and prognosis of cancers. The roles of NK cell-related genes in the tumor immune microenvironment (TIME) and immunotherapy treatment are unclear. Triple-negative breast cancer (TNBC) is a highly aggressive malignant tumor. Hence, this study was conducted to develop a reliable risk model related to NK cells and provide a novel system for predicting the prognosis of TNBC. Methods: NK cell-related genes were collected from previous studies. Based on TCGA and GEO database, univariate and LASSO cox regression analysis were used to establish the NK cell-related gene signature. The patients with TNBC were separated to high-risk and low-risk groups. After that, survival analysis was conducted and the responses to immunotherapies were evaluated on the basis of the signature. Moreover, the drug sensitivity of some traditional chemotherapeutic drugs was assessed by using the "oncoPredict" R package. In addition, the expression levels of the genes involved in the signature were validated by using qRT-PCR in TNBC cell lines. Results: The patients with TNBC were divided into high- and low-risk groups according to the median risk score of the 5-NK cell-related gene signature. The low-risk group was associated with a better clinical outcome. Besides, the differentially expressed genes between the different risk groups were enriched in the biological activities associated with immunity. The tumor immune cells were found to be highly infiltrated in the low-risk groups. In accordance with the TIDE score and immune checkpoint-related gene expression analysis, TNBC patients in the low-risk groups were suggested to have better responses to immunotherapies. Eventually, some classical anti-tumor drugs were shown to be less effective in high-risk groups than in low-risk groups. Conclusion: The 5-NK cell-related gene signature exhibit outstanding predictive performance and provide fresh viewpoints for evaluating the success of immunotherapy. It will provide new insights to achieve precision and integrated treatment for TNBC in the future.

Long noncoding RNA XLOC_006786 inhibits the proliferation, invasion and metastasis of osteosarcoma cells through NOTCH3 signaling pathway by targeting miR-491-5p.

Recent research has indicated that Long noncoding RNAs (LncRNAs) are crucial in many disorders, especially tumors. However, the exact role of LncRNA XLOC_006786 (LncRNA-SPIDR-2:1) in malignancies, especially in human osteosarcoma, is unclear. The results of RT‒qPCR, western blotting, CCK-8 assays, and Transwell assays showed that LncRNA XLOC_006786 inhibited osteosarcoma cell proliferation, invasion, and migration, indicating that it may be a tumor suppressor gene in osteosarcoma. We found that LncRNA XLOC_006786 negatively regulated NOTCH3, which is an oncogenic gene in osteosarcoma, as we previously reported. Bioinformatics analysis showed that miR-491-5p may be a direct target of LncRNA XLOC_006786, while NOTCH3 is a key target of miR-491-5p. Then, we verified that LncRNA XLOC_006786 could prevent lung metastatic osteosarcoma in vivo. Taken together, our research showed that LncRNA XLOC_006786 suppresses osteosarcoma proliferation, invasion, and metastasis through the NOTCH3 signaling pathway by targeting miR-491-5p.

Rat tight junction proteins are disrupted after subchronic exposure to okadaic acid.

Okadaic acid (OA), a lipophilic phycotoxin distributed worldwide, causes diarrheic shellfish poisoning and even leads to tumor formation. Currently, the consumption of contaminated seafood is the most likely cause of chronic OA exposure, but there is a serious lack of relevant data. Here, the Sprague-Dawley rats were exposure to OA by oral administration at 100 µg/kg body weight, and the tissues were collected and analyzed to assess the effect of subchronic OA exposure. The results showed that subchronic OA administration disturbed colonic mucosal integrity and induced colitis. The colonic tight junction proteins were disrupted and the cell cycle of colonic epithelial cells was accelerated. It is inferred that disruption of the colonic tight junction proteins might be related to the development of chronic diarrhea by affecting water and ion transport. Moreover, the accelerated proliferation of colonic epithelial cells indicated that subchronic OA exposure might promote the restitution process of gut barrier or induce tumor promoter activity in rat colon.

TSSC3 overexpression associates with growth inhibition, apoptosis induction and enhances chemotherapeutic effects in human osteosarcoma.

Loss of expression of TSSC3, an apoptosis-related imprinted gene, has been reported in several cases of malignant tumors. However, the roles and mechanisms of TSSC3 in human osteosarcoma remain to be defined. In this study, we found TSSC3 to be downregulated during osteosarcoma transformation and progression in osteosarcoma cell lines and tissues. The SaOS2 cell line was used to further evaluate the precise role of TSSC3 in osteosarcoma development. Overexpression of TSSC3 markedly reduced cell vitality and growth, colony formation, Ki67 expression as well as cell cycle arrest in the G(0)/G(1) phase. Consistently, TSSC3 overexpression was associated with increased apoptosis assayed by annexin V/propidium iodide and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining. Subcutaneous injection of TSSC3 overexpressing SaOS2 cells into athymic nude mice showed that TSSC3 also inhibited tumorigenesis through growth inhibition and apoptosis induction in vivo. Further mechanistic studies revealed that the mitochondrial apoptosis pathway was required for TSSC3-mediated cell apoptosis. These findings support a suppressor role for TSSC3 in osteosarcoma development by regulating apoptosis. In addition, constitutive TSSC3 expression greatly enhanced the sensitivity of human osteosarcoma cells to the chemotherapeutic drugs cisplatin and epirubicin. Conversely, TSSC3 knockdown increased SaOS2 cell growth and decreased apoptosis in vitro and in vivo and reduced sensitivity of the cells to chemotherapy. This is the first study to demonstrate that TSSC3 has a potent tumor suppressor role in osteosarcoma, probably by inhibition of growth and induction of apoptosis via the mitochondrial apoptosis pathway.

TSSC3 overexpression reduces stemness and induces apoptosis of osteosarcoma tumor-initiating cells.

Osteosarcoma (OS) is the most common primary bone tumor in children and adolescents, typically presenting with poor prognosis. Recent studies suggested that tumor initiating cells (T-ICs) drive tumor formation and relapse or metastasis and are relatively resistant to cell death induced by conventional chemo- and radiotherapies. Therefore, the poor prognosis of OS appears to be associated with T-ICs. Here, we enriched T-ICs in OS cell lines and evaluated whether the imprinted gene TSSC3 (tumor-suppressing STF cDNA 3) associated with apoptosis could affect T-ICs in OS. Sarcosphere selection and serial clone-forming unit assays were successfully used to enrich T-ICs from OS cell lines. Enrichment of T-ICs from a malignantly transformed hFOB1.19 osteoblast cell line (MThFOB1.19) indicated that OS T-ICs could originate from differentiated cells, and most of these MThFOB1.19 cells showed stem-like features. TSSC3 was expressed at a low level in T-ICs, while overexpression of TSSC3 could efficiently downregulate the expression of stem cell markers Nanog, Oct4 and Sox2 in T-ICs and decrease the clone formation rate, as well as downregulate tumorigenesis in MThFOB1.19 cells, supporting a suppressive role for TSSC3 in OS T-ICs. Furthermore, overexpression of TSSC3 was found to induce apoptosis of OS T-ICs through increasing cleaved caspase-3 (active form), increasing the release of Cyt c and decreasing pro-caspase-9 (pro-enzyme form), as well as disruption of the mitochondrial membrane potential (ΔΨ). Taken together, our findings provide preliminary evidence that TSSC3 inhibits OS tumorigenicity through reducing stemness and promoting apoptosis of T-ICs. Thus, targeting TSSC3 may be a promising approach to suppressing tumorigenicity in OS.