Metastasis and basement membrane-related signature enhances hepatocellular carcinoma prognosis and diagnosis by integrating single-cell RNA sequencing analysis and immune microenvironment assessment.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer and second leading cause of cancer-related deaths worldwide. The heightened mortality associated with HCC is largely attributed to its propensity for metastasis, which cannot be achieved without remodeling or loss of the basement membrane (BM). Despite advancements in targeted therapies and immunotherapies, resistance and limited efficacy in late-stage HCC underscore the urgent need for better therapeutic options and early diagnostic biomarkers. Our study aimed to address these gaps by investigating and evaluating potential biomarkers to improve survival outcomes and treatment efficacy in patients with HCC. METHOD: In this study, we collected the transcriptome sequencing, clinical, and mutation data of 424 patients with HCC from The Cancer Genome Atlas (TCGA) and 240 from the International Cancer Genome Consortium (ICGC) databases. We then constructed and validated a prognostic model based on metastasis and basement membrane-related genes (MBRGs) using univariate and multivariate Cox regression analyses. Five immune-related algorithms (CIBERSORT, QUANTISEQ, MCP counter, ssGSEA, and TIMER) were then utilized to examine the immune landscape and activity across high- and low-risk groups. We also analyzed Tumor Mutation Burden (TMB) values, Tumor Immune Dysfunction and Exclusion (TIDE) scores, mutation frequency, and immune checkpoint gene expression to evaluate immune treatment sensitivity. We analyzed integrin subunit alpha 3 (ITGA3) expression in HCC by performing single-cell RNA sequencing (scRNA-seq) analysis using the TISCH 2.0 database. Lastly, wound healing and transwell assays were conducted to elucidate the role of ITGA3 in tumor metastasis. RESULTS: Patients with HCC were categorized into high- and low-risk groups based on the median values, with higher risk scores indicating worse overall survival. Five immune-related algorithms revealed that the abundance of immune cells, particularly T cells, was greater in the high-risk group than in the low-risk group. The high-risk group also exhibited a higher TMB value, mutation frequency, and immune checkpoint gene expression and a lower tumor TIDE score, suggesting the potential for better immunotherapy outcomes. Additionally, scRNA-seq analysis revealed higher ITGA3 expression in tumor cells compared with normal hepatocytes. Wound healing scratch and transwell cell migration assays revealed that overexpression of the MBRG ITGA3 enhanced migration of HCC HepG2 cells. CONCLUSION: This study established a direct molecular correlation between metastasis and BM, encompassing clinical features, tumor microenvironment, and immune response, thereby offering valuable insights for predicting clinical outcomes and immunotherapy responses in HCC.

Licochalcone A induces endoplasmic reticulum stress-mediated apoptosis of endometrial cancer cells via upregulation of GRP78 expression.

Licochalcone A (LicA), a natural compound extracted from licorice root, has been shown to exert a variety of anticancer activities. Whether LicA has such effects on endometrial cancer (EMC) is unclear. This study aims to investigate the antitumor effects of LicA on EMC. Our results show that LicA significantly reduced the viability and induced apoptosis of EMC cells and EMC-7 cells from EMC patients. LicA was also found to induce endoplasmic reticulum (ER) stress, leading to increased expression of ER-related proteins (GRP78/PERK/IRE1α/CHOP) in EMC cell lines. Suppression of GRP78 expression in human EMC cells treated with LicA significantly attenuated the effects of LicA, resulting in reduced ER-stress mediated cell apoptosis and decreased expression of ER- and apoptosis-related proteins. Our findings demonstrate that LicA induces apoptosis in EMC cells through the GRP78-mediated ER-stress pathway, emphasizing the potential of LicA as an anticancer therapy for EMC.

[Using Cloud Healthcare to Improve Home Learning Efficacy in Congenital Heart Disease Infants].

BACKGROUND: During the COVID-19 pandemic, visitation restrictions in line with infection control policies curtailed opportunities for family members to learn essential caregiving skills in the intensive care unit. This limitation decreased satisfaction among family members, possibly indicating their increased difficulties in care due to the lack of face-to-face guidance. Thus, increasing family member understanding of and ability to apply learning content without direct interaction presents a significant and urgent challenge. Moreover, because of lack of caregiving confidence, some family members may be reluctant to facilitate the discharge of critically ill patients, causing delays in discharge planning. These challenges underscore the obstacles faced by nursing health education during the pandemic. PURPOSE: This study was designed to utilize cloud technology to enhance the knowledge and skills of families caring for infants with congenital heart disease at home and to assess their satisfaction with the associated homecare learning platform. RESOLUTION: Based on our hospital's cloud-based health education platform, a series of personalized instructional video materials was developed for families of infants with congenital heart disease. These materials cover comprehensively the entire treatment process, from diagnosis to post-discharge home care skills, for these patients. To facilitate autonomous learning, the videos in this series were made accessible to the families anytime, anywhere via personal devices such as smartphones and tablets. Concurrently, a chatbot tool was integrated to provide guidance on inpatient care for infants with congenital heart disease, including fundamental aspects of newborn care, with the aim of equipping parents and caregivers with the knowledge and skills necessary to provide basic post-discharge care. To ensure the families acquired personalized care skills, after completing the learning modules, practical bedside training sessions incorporating knowledge and skills assessments were organized for family members. RESULTS: After project implementation, the average knowledge score for family members increased significantly from 79.1 to 100 (perfect score). The proportion of family members proficient in executing caregiving techniques autonomously also rose impressively from 30% to 95%. Furthermore, average overall satisfaction with cloud-based technology-assisted caregiving learning among the family members rose 31.4% from 3.5 to 4.6. CONCLUSIONS: This project represents a viable solution to providing clinical nursing guidance independent of the constraints of time and location, and effectively enhances homecare-skill-related learning outcomes in family members, especially with regard to caring for infants with congenital heart disease.

Hsa_circ_0000129 drives tumor growth via sequestering miR-485-3p and upregulating SPIN1 in breast cancer.

BACKGROUND: Breast cancer (BC) is second cancer frequently occurring worldwide. Circular RNA hsa_circ_0000129 (circ_0000129) exerts a tumor-promoting effect in BC. Nevertheless, the molecular mechanisms mediated by the upregulation of circ_0000129 during BC progression are not well understood. METHODS: Forty-five BC patients were recruited for the research. Changes in circ_0000129 levels were detected with quantitative reverse transcription-polymerase chain reaction. Cell proliferation, apoptosis, migration, invasion, and angiopoiesis were determined by cell counting, 5-ethynyl-2'-deoxyuridine (EdU), flow cytometry, transwell, and tube formation assays. Protein levels were detected by western blot analysis. The regulatory mechanism of circ_0000129 was predicted by bioinformatics analysis and validated by dual-luciferase reporter and RNA immunoprecipitation assays. In vivo experiments were carried out to verify the function of circ_0000129. RESULTS: Circ_0000129 was overexpressed in BC samples and cell lines. Functionally, circ_0000129 silencing reduced cell proliferation, migration, invasion, and promoted cell apoptosis, as well as induced HUVEC angiopoiesis in vitro. Furthermore, circ_0000129 knockdown decreased BC cell growth in mouse xenograft models. Mechanically, circ_0000129 interacted with miR-485-3p to mediate the inhibiting effect of miR-485-3p on SPIN1. Silenced miR-485-3p expression weakened the inhibiting effect of circ_0000129 knockdown on BC cell malignant behaviors. Also, forced SPIN1 expression weakened miR-485-3p upregulation mediated effects on BC cell malignant behaviors. CONCLUSION: Circ_0000129 acted as a miR-485-3p sponge molecular to mediate expression, thus promoting BC progression.

Rosavin: Research Advances in Extraction and Synthesis, Pharmacological Activities and Therapeutic Effects on Diseases of the Characteristic Active Ingredients of Rhodiola rosea L.

Rhodiola rosea L. (RRL) is a popular plant in traditional medicine, and Rosavin, a characteristic ingredient of RRL, is considered one of the most important active ingredients in it. In recent years, with deepening research on its pharmacological actions, the clinical application value and demand for Rosavin have been steadily increasing. Various routes for the extraction and all-chemical or biological synthesis of Rosavin have been gradually developed for the large-scale production and broad application of Rosavin. Pharmacological studies have demonstrated that Rosavin has a variety of biological activities, including antioxidant, lipid-lowering, analgesic, antiradiation, antitumor and immunomodulation effects. Rosavin showed significant therapeutic effects on a range of chronic diseases, including neurological, digestive, respiratory and bone-related disorders during in vitro and vivo experiments, demonstrating the great potential of Rosavin as a therapeutic drug for diseases. This paper gives a comprehensive and insightful overview of Rosavin, focusing on its extraction and synthesis, pharmacological activities, progress in disease-treatment research and formulation studies, providing a reference for the production and preparation, further clinical research and applications of Rosavin in the future.

PARP-1 modulates the expression of miR-223 through histone acetylation to involve in the hydroquinone-induced carcinogenesis of TK6 cells.

The upstream regulators of microRNAs were rarely reported. Hydroquinone (HQ) is the main metabolite of benzene, one of the important environmental factors contributing to leukemia and lymphoma. In HQ-induced malignant transformed TK6 (TK6-HT) cells, the expression of PARP-1 and miR-223 were upregulated. When in PARP-1 silencing TK6-HT cells, miR-223 was downregulated and the apoptotic cell number correspondingly increased. In TK6 cells treated with HQ for different terms, the expression of miR-223 and PARP-1 were dynamically observed and found to be decreased and increased, respectively. Trichostatin A could increase the expression of miR-223, then the expression of HDAC1-2 and nuclear factor kappa B were found to be increased, but that of mH2A was decreased. PARP-1 silencing inhibited the protein expression of H3Ac, mH2A, and H3K27ac. By co-immunoprecipitation experiment, PARP-1 and HDAC2 were found to form a regulatory complex. In conclusion, we demonstrated that the upregulation of PARP-1 mediated activation of acetylation to promote the transcription of miR-223 possibly via coregulating with HDAC2, an epigenetic regulation mechanism involved in cell malignant transformation resulting from long-term exposure to HQ, in which course, H3K27ac might be a specific marker for the activation of histone H3, which also gives hints for benzene exposure research.

Targeting of Mcl-1 Expression by MiRNA-3614-5p Promotes Cell Apoptosis of Human Prostate Cancer Cells.

MicroRNA (miRNA) acts as a critical regulator of growth in various human malignancies. However, the role of miRNA-3614 in the progression of human prostate cancer remains unknown. In this study, our results demonstrated that miRNA-3614-5p exerts a significant inhibitory effect on cell viability and colony formation and induces sub-G1 cell cycle arrest and apoptosis in human prostate cancer cells. Myeloid cell leukemia-1 (Mcl-1) acts as a master regulator of cell survival. Using the miRNA databases, miRNA-3614-5p was found to regulate Mcl-1 expression by targeting positions of the Mcl-1-3' UTR. The reduction of Mcl-1 expression by miRNA-3614-5p was further confirmed using an immunoblotting assay. Pro-apoptotic caspase-3 and poly (ADP-ribose) polymerase (PARP) were significantly activated by miRNA-3614-5p to generate cleaved caspase-3 (active caspase-3) and cleaved PARP (active PARP), accompanied by the inhibited Mcl-1 expression. These findings were the first to demonstrate the anti-growth effects of miRNA-3614-5p through downregulating Mcl-1 expression in human prostate cancer cells.

Norcantharidin combined with paclitaxel induces endoplasmic reticulum stress mediated apoptotic effect in prostate cancer cells by targeting SIRT7 expression.

Prostate cancer (PCa), an extremely common malignancy in males, is the most prevalent disease in several countries. Norcantharidin (NCTD) has antiproliferation, antimetastasis, apoptosis, and autophagy effects in various tumor cells. Nevertheless, the antitumor effect of NCTD combined with paclitaxel (PTX), a chemotherapeutic drug, in PCa remains unknown. The cell growth, proliferative rate, cell cycle distribution, and cell death were determined by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide, colony formation assay, PI staining, and Annexin V/PI staining by flow cytomertry, whereas the mitochondrial membrane potential (MMP) and endoplasmic reticulum (ER) stress was evaluated using the MitoPotential assay and ER-ID red assay. We also evaluated the protein and mRNA expression of SIRTs by Western blotting and qRTPCR assay. Overexpression effectivity was measured by DNA transfection assay. Our study showed that cell viability and proliferative PC3 and DU145 rates were effectively inhibited after NCTD-PTX combination. We also found that NCTD-PTX combination treatment significantly enhance G2/M phase arrest, induction of cell death and ER stress, loss of MMP, and ER- or apoptotic-related protein expression. Furthermore, NCTD-PTX combination treatment was significantly decreasing the protein and mRNA expression of SIRT7 in PCa cells. Combination therapy effectively reduced cell viability, ER stress-mediated apoptosis and p-eIF2α/ATF4/CHOP/cleaved-PARP expression inhibition in SIRT7 overexpression of PCa cells. These results indicate that NCTD combined with PTX induces ER stress-mediated apoptosis of PCa cells by regulating the SIRT7 expression axis. Moreover, combination therapy may become a potential therapeutic strategy against human PCa.

Silencing PROK2 Inhibits Invasion of Human Cervical Cancer Cells by Targeting MMP15 Expression.

Cervical cancer is the second most frequent type of gynecologic cancer worldwide. Prokineticin 2 (PROK2) is reported to be involved in tumor progression in some malignant tumors. However, the role of PROK2 in the development of cervical cancer remains unknown. Our results indicate that PROK2 is overexpressed in the human cervical cancer. Cervical cancer patients with high PROK2 expression have a shorter overall survival rate (OS) and disease-free survival rate (DFS). PROK2 acts as a potential biomarker for predicting OS and DFS of cervical cancer patients. We further show that PROK2 is important factor for oncogenic migration and invasion in human cervical cancer cells. Knockdown PROK2 significantly inhibited cell migration, invasion, and MMP15 protein expression in HeLa cells. High expression of MMP15 is confirmed in the human cervical cancer, is significantly associated with the shorter overall survival rate (OS) and is correlated with PROK2 expression. Overexpression of PROK2 using PROK2 plasmid significantly reverses the function of knockdown PROK2, and further upregulates MMP15 expression, migration and invasion of human cervical cancer cells. In conclusion, our findings are the first to demonstrate the role of PROK2 as a novel and potential biomarker for clinical use, and reveal the oncogenic functions of PROK2 as therapeutic target for cervical cancer.

The EMT-related transcription factor snail up-regulates FAPα in malignant melanoma cells.

FAPα is a cell surface serine protease, mainly expressed in tumor stromal fibroblasts in more than 90% of human epithelial carcinomas. Due to its almost no expression in normal tissues and its tumor-promoting effects, FAPα has been studied as a novel potential target for antitumor therapy. However, the regulation mechanism on FAPα expression is poorly understood. In this study, we found that overexpression of snail significantly increased the mRNA and protein expression levels of FAPα in malignant melanoma B16 and SK-MEL-28 cells. Overexpression of snail increased FAPα promoter activity remarkably. Snail could directly bind to FAPα promoter to regulate FAPα expression. Moreover, snail expression was positively correlated to FAPα expression in human cutaneous malignant melanoma. Furthermore, knockdown of FAPα markedly reduced snail-induced cell migration. Overall, our findings provide a novel regulation mechanism on FAPα expression and highlight the role of snail/FAPα axis as a novel target for melanoma treatment.

Cancer-associated fibroblasts promote epithelial-mesenchymal transition and EGFR-TKI resistance of non-small cell lung cancers via HGF/IGF-1/ANXA2 signaling.

The involvement of the tumor stromal cells in acquired resistance of non-small cell lung cancers (NSCLCs) to tyrosine kinase inhibitors (TKIs) has previously been reported, but the precise mechanism remains unclear. In the present study, we investigated the role and mechanism underlying Cancer-associated fibroblasts (CAFs) in TKI resistance of NSCLCs. In vitro and in vivo experiments showed that HCC827 and PC9 cells, non-small cell lung cancer cells with EGFR-activating mutations, became resistant to the EGFR-TKI gefitinib when cultured with CAFs isolated from NSCLC tissues. Moreover, we showed that CAFs could induce epithelial-mesenchymal transition (EMT) phenotype of HCC827 and PC9 cells, with an associated change in the expression of epithelial to mesenchymal transition markers. Using proteomics-based method, we identified that CAFs significantly increased the expression of the Annexin A2 (ANXA2). More importantly, knockdown of ANXA2 completely reversed EMT phenotype and gefitinib resistance induced by CAFs. Furthermore, we found that CAFs increased the expression and phosphorylation of ANXA2 by secretion of growth factors HGF and IGF-1 and by activation of the corresponding receptors c-met and IGF-1R. Dual inhibition of HGF/c-met and IGF-1/IGF-1R pathways could significantly suppress ANXA2, and markedly reduced CAFs-induced EMT and gefitinib resistance. Taken together, these findings indicate that CAFs promote EGFR-TKIs resistance through HGF/IGF-1/ANXA2/EMT signaling and may be an ideal therapeutic target in NSCLCs with EGFR-activating mutations.

The hypermethylation of p16 gene exon 1 and exon 2: potential biomarkers for colorectal cancer and are associated with cancer pathological staging.

BACKGROUND: Tumor suppressor gene p16 promoter hypermethylation has been widely studied in colorectal cancer (CRC), yet its clinicopathological significance remains controversial. The methylation alterations of other regions within p16 gene are still rarely researched. The present study aimed to explore the methylation changes of p16 gene body in CRC and to find whether they were associated with clinicopathological staging of CRC. METHODS: Paired colorectal cancer tissues and corresponding adjacent normal tissues from 30 CRC patients were collected. The methylation levels of two CpG islands within p16 gene body, exon 1 and exon 2, were accurately assessed simultaneously by a LC-MS/MS method. The p16 protein expressions were assessed by immunohistochemistry assay. Statistical analyses were carried out using SPSS 17.0 software. Heat-map analysis was carried out by HemI 1.0 software. RESULTS: In the present study, CRC tissues showed more highly methylated than adjacent normal tissues at both CpG islands of p16 gene. And exon 2 hypermethylation was higher and more frequent than exon 1. The ROC curve analysis showed that the simultaneous use of both indicators had excellent sensitivity and specificity for distinguishing CRC tissues and adjacent normal tissues. Following, the methylation level of p16 exon 1/2 was negatively related to p16 protein expression. Further correlation analysis revealed that p16 exon 1 hypermethylation was associated with N/Dukes staging (p = 0.033), and p16 exon 2 hypermethylaiton was associated with T staging (p = 0.035). CONCLUSIONS: The p16 gene body was remarkably hyper-methylated in CRC tissues and associated with p16 protein expression and cancer clinicopathological staging. The combination of p16 exon 1 and exon 2 could better reflect the overall methylation status of p16 gene body and provide potential biomarkers of CRC.

Synergistic antimetastatic effect of cotreatment with licochalcone A and sorafenib on human hepatocellular carcinoma cells through the inactivation of MKK4/JNK and uPA expression.

To improve the clinical outcome of tumor chemotherapy, more effective combination treatments against tumor metastasis and recurrence are required. Licochalcone A (LicA) is the root of Glycyrrhiza inflata and has been reported to possess anti-inflammatory, antimicrobial, and antitumor effects. Sorafenib (Sor), a multikinase inhibitor, is used to treat patients with solid tumors such as advanced hepatocellular carcinoma (HCC). However, the synergistic effects of LicA and Sor on the metastasis of human HCC cells have not been reported. We found that LicA and Sor did not have cytotoxic effects or arrest growth in human SK-Hep-1 and Huh-7 cells. In addition, treatment with LicA or Sor alone inhibited migration and invasion in human SK-Hep-1 and Huh-7 HCC cells. Furthermore, cotreatment with LicA and Sor synergistically inhibited the migration and invasion of HCC cells and significantly inhibited uPA protein expression. Notably, cotreatment of LicA and Sor synergistically and significantly downregulated MKK4-JNK expression. Through tail vein injection in nude mice, the aforementioned cotreatment synergistically suppressed SK-Hep-1 cell-mediated lung metastasis. These findings first revealed the synergistic effects of LicA and Sor cotreatment against human HCC cells, further suggesting that beneficial effects on tumor regression could be confirmed through prospective clinical trials.

Rb silencing mediated by the down-regulation of MeCP2 is involved in cell transformation induced by long-term exposure to hydroquinone.

Hydroquinone (HQ), a metabolite of benzene, is a well-known human carcinogen; however, its molecular mechanisms of action remain unclear. MeCP2 has been traditionally described as a transcriptional repressor, though growing evidence indicates that it also activates gene expression. Here, we investigated whether some epigenetic machinery genes are aberrantly expressed as target tumor suppressor genes in HQ-transformed TK6 lymphoblastoid cells. Our results showed that treatment with 5-Aza-2'-deoxycytidine or trichostatin A enhanced the expression of Rb, resulting in cell arrest in G1-phase, and subsequently, an increase in apoptosis and a decrease in cell growth. Moreover, we hypothesised that Rb was silenced by the down-regulation of MeCP2 in HQ-transformed cells, resulting in the dynamic expression of Rb and epigenetic machinery proteins in HQ-transformed cells at different time points. The expression of Rb and MeCP2 in patients with B-cell non-Hodgkin's lymphoma (B-NHL) showed that positive staining for MeCP2 or Rb was significantly lower in B-NHL tumor tissues, and these changes were significantly and negatively correlated with the grade of B-NHL. The restoration of MeCP2 in HQ-transformed cells enhanced the expression of Rb, promoted cell apoptosis, and inhibited cell growth. The changes in the expression patterns of MeCP2 and Rb were inversely correlated with the degree of DNA methylation. A ChiP assay revealed that MeCP2 proteins were recruited to the Rb promoter with lower 5'-methylcytosine levels. In conclusion, we demonstrated that the down-regulation of MeCP2 silences Rb, a process involved in cell transformation resulting from long-term exposure to HQ. © 2016 Wiley Periodicals, Inc.

Praeruptorin A Inhibits Human Cervical Cancer Cell Growth and Invasion by Suppressing MMP-2 Expression and ERK1/2 Signaling.

Praeruptorin A (PA) is a pyranocumarin present in the dried root of Peucedanumpraeruptorum Dunn that has anticancer effects against several types of cells. However, the effect of PA on human cervical cancer cells is unknown. Our results indicate that PA significantly inhibited cell proliferation, colony formation, migration, invasion, and wound closure of HeLa and SiHa cells, induced cell cycle arrest at G0/G1 phase, upregulated Rb, p16, p21 and p27 proteins and downregulated cyclin D1 and S-phase kinase-associated protein 2 (Skp2) proteins. PA also significantly reduced expression of matrix metalloproteinase-2 (MMP-2) and increased expression of tissue inhibitor of metalloproteinase-2 (TIMP-2). In addition, PA suppressed ERK1/2 activation and increased the effect of PD98059 (a specific MEK1/2 inhibitor) in downregulation of MMP-2 and upregulation of TIMP-2. PA treatment inhibited the effect of 12-O-tetradecanoylphorbol-13-acetate (TPA) on upregulation of ERK1/2 activation, MMP-2 expression, cellular migration, and invasion of HeLa cells. Our findings are the first to demonstrate the activity of PA against cervical cancer cells, and suggest this agent has promise as a therapeutic agent in treatment of human cervical cancer.

UBE2C is a diagnosis and therapeutic biomarker involved in immune infiltration of cancers including lung adenocarcinoma.

The mechanism of action of UBE2C in lung adenocarcinoma (LUAD) and its significance in cancer diagnosis, targeted therapy and immunotherapy, even in pan-cancer, are still unclear. Several large public databases and online analysis tools were used for big data mining analysis. RNA interference technology, CCK8 assay, flow cytometry and apoptosis detection, and western blot were used for in vitro experiments. UBE2C was found to be overexpressed in various of tumors, including LUAD, and its expression level was found to be significantly related to gender, weight, tumor stage, grade and prognosis in LUAD. Downregulation of UBE2C expression induced proliferation suppression and G2/M phase arrest and cell apoptosis in LUAD cells and suppressed LUAD cell growth through inhibiting the Akt-mTOR signaling pathway. Expression level of UBE2C was negatively correlated with B cells and CD4+ T cell, and also with immune checkpoint genes in LUAD. Pan-cancer assay shown that UBE2C was significantly overexpressed in 28 cancers and was correlated with Ki-67 index in many cancers. Overexpression of UBE2C in BRCA, LUAD and MESO indicated worse Overall Survival (OS). UBE2C expression levels were positively associated with immunocyte infiltration, immune regulatory genes, immune checkpoints, TMB, MSI and MMRs in some cancers. Additionally, Single-cell functional analysis showed that UBE2C was positively correlated with cell cycle, proliferation, DNA damage, EMT, DNA repair, invasion and differentiation in some cancers. These findings suggested that UBE2C could be regarded as a latent diagnosis and prognostic biomarker and a new target for immunological therapy of cancers including LUAD.

Identification and validation of immune-related methylated genes as diagnostic and prognostic biomarkers of nasopharyngeal carcinoma.

BACKGROUND: Nasopharyngeal carcinoma (NPC) is a common malignancy occurring in the head and neck. Identification of immune-related methylated biomarkers might be helpful for NPC detection and prognostic evaluation. METHODS: A co-methylation network based on WGCNA was constructed to identify modules associated with NPC and immune cells. In combination with differentially expressed genes (DEGs) and immune-related genes from ImmPort database, the candidate immune-related methylated genes (IRMGs) were obtained. RESULTS: Our combined analysis identified 12 IRMGs. Among them, both the methylation and mRNA expression of CCL28, CSK, and PRKCB were correlated with the infiltration of B cells. CD1D, CR2, and GDF10 were favorable markers. Demethylation experiments validated that downregulation of GDF10, PRKCB, SLC40A1, and TGFBR3 in NPC resulted from promoter hypermethylation. Additionally, a diagnostic model was developed and exhibited high discriminative accuracy. CONCLUSIONS: These results provided a group of immune-related methylated biomarkers that may help with the diagnosis and prognosis of NPC.

MiRNA-296-5p promotes the sensitivity of nasopharyngeal carcinoma cells to cisplatin via targeted inhibition of STAT3/KLF4 signaling axis.

Improving drug sensitivity is an important strategy in chemotherapy of cancer and accumulating evidence indicates that miRNAs are involved in the regulation of drug sensitivity, but the specific mechanism is still unclear. Our previous study has found that miR-296-5p was significantly downregulated in nasopharyngeal carcinoma (NPC). Here, we aim to explore whether miR-296-5p is involved in regulating cisplatin sensitivity in NPC by regulating STAT3/KLF4 signaling axis. The cell proliferation and clonogenic capacity of NPC cells were evaluated by CCK8 Assay and plate colony assay, respectively. The Annexin V-FITC staining kit was used to determine and quantify the apoptotic cells using flow cytometry. The drug efflux ability of NPC cells were determined by Rhodamine 123 efflux experiment. The expression of miR-296-5p, apoptosis-related genes and protein in NPC cell lines were detected by qPCR and Western blot, respectively. Animal study was used to evaluate the sensitivity of NPC cells to DDP treatment in vivo. Our results showed that elevated miR-296-5p expression obviously promoted the sensitivity of NPC cells to DDP by inhibiting cell proliferation and clonogenic capacity, and inducing apoptosis. In addition, we found that miR-296-5p inhibited the expression of STAT3 and KLF4 in NPC cells, while overexpression of exogenous STAT3 reversed miR-296-5p-mediated enhancement in cell death of DDP-treated NPC cells. In vivo studies further confirmed that miR-296-5p promotes the sensitivity of NPC cells to DDP treatment. miRNA-296-5p enhances the drug sensitivity of nasopharyngeal carcinoma cells to cisplatin via STAT3/KLF4 signaling pathway.

Signal flow in the NMDA receptor-dependent phosphoproteome regulates postsynaptic plasticity for aversive learning.

Structural plasticity of dendritic spines in the nucleus accumbens (NAc) is crucial for learning from aversive experiences. Activation of NMDA receptors (NMDARs) stimulates Ca2+-dependent signaling that leads to changes in the actin cytoskeleton, mediated by the Rho family of GTPases, resulting in postsynaptic remodeling essential for learning. We investigated how phosphorylation events downstream of NMDAR activation drive the changes in synaptic morphology that underlie aversive learning. Large-scale phosphoproteomic analyses of protein kinase targets in mouse striatal/accumbal slices revealed that NMDAR activation resulted in the phosphorylation of 194 proteins, including RhoA regulators such as ARHGEF2 and ARHGAP21. Phosphorylation of ARHGEF2 by the Ca2+-dependent protein kinase CaMKII enhanced its RhoGEF activity, thereby activating RhoA and its downstream effector Rho-associated kinase (ROCK/Rho-kinase). Further phosphoproteomic analysis identified 221 ROCK targets, including the postsynaptic scaffolding protein SHANK3, which is crucial for its interaction with NMDARs and other postsynaptic scaffolding proteins. ROCK-mediated phosphorylation of SHANK3 in the NAc was essential for spine growth and aversive learning. These findings demonstrate that NMDAR activation initiates a phosphorylation cascade crucial for learning and memory.

miR-124 radiosensitizes human glioma cells by targeting CDK4.

The aberrant expression of cyclin-dependent kinase-4 (CDK4) has previously been observed in human brain glioma. Furthermore, it is observed that up-regulation of CDK4 is associated with therapy resistance and relapse. However, the mechanisms behind these phenomena remain unclear. Here, we demonstrated that elevated CDK4 expression is correlated with poor prognosis in glioma after radiotherapy and that CDK4 knockdown conferred radiosensitivity in glioma cell lines. CDK4 was identified as potential downstream target of miR-124 through bioinformatics analysis and dual-firefly luciferase reporter assay. Furthermore, restoration of miR-124 could confer radiosensitivity. Cell differentiation agent-2 (CDA-2) mimicked the effect of miR-124 restoration and CDK4 knockdown, and sensitized xenografts to radiation in an animal model. Our findings demonstrated for the first time that CDK4 was a downstream target of miR-124 and that CDA-2 could radiosensitize Glioblastoma multiforme cells through the MiR-124-CDK4 axis.