Creatine kinase mitochondrial 2 promotes the growth and progression of colorectal cancer via enhancing Warburg effect through lactate dehydrogenase B.

Background: Mitochondrial creatine kinase (MtCK) plays a pivotal role in cellular energy metabolism, exhibiting enhanced expression in various tumors, including colorectal cancer (CRC). Creatine kinase mitochondrial 2 (CKMT2) is a subtype of MtCK; however, its clinical significance, biological functions, and underlying molecular mechanisms in CRC remain elusive. Methods: We employed immunohistochemical staining to discern the expression of CKMT2 in CRC and adjacent nontumor tissues of patients. The correlation between CKMT2 levels and clinical pathological factors was assessed. Additionally, we evaluated the association between CKMT2 and the prognosis of CRC patients using Kaplan-Meier survival curves and Cox regression analysis. Meanwhile, quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to detect the expression levels of CKMT2 in different CRC cell lines. Finally, we explored the biological functions and potential molecular mechanisms of CKMT2 in CRC cells through various techniques, including qRT-PCR, cell culture, cell transfection, western blot, Transwell chamber assays, flow cytometry, and co-immunoprecipitation. Results: We found that CKMT2 was significantly overexpressed in CRC tissues compared with adjacent nontumor tissues. The expression of CKMT2 is correlated with pathological types, tumor size, distant metastasis, and survival in CRC patients. Importantly, CKMT2 emerged as an independent prognostic factor through Cox regression analysis. Experimental downregulation of CKMT2 expression in CRC cell lines inhibited the migration and promoted apoptosis of these cells. Furthermore, we identified a novel role for CKMT2 in promoting aerobic glycolysis in CRC cells through interaction with lactate dehydrogenase B (LDHB). Conclusion: In this study, we found the elevated expression of CKMT2 in CRC, and it was a robust prognostic indicator in CRC patients. CKMT2 regulates glucose metabolism via amplifying the Warburg effect through interaction with LDHB, which promotes the growth and progression of CRC. These insights unveil a novel regulatory mechanism by which CKMT2 influences CRC and provide promising targets for future CRC therapeutic interventions.

Context-dependent roles for ubiquitous mitochondrial creatine kinase CKMT1 in breast cancer progression.

Metabolic reprogramming is a hallmark of cancer, enabling cancer cells to rapidly proliferate, invade, and metastasize. We show that creatine levels in metastatic breast cancer cell lines and secondary metastatic tumors are driven by the ubiquitous mitochondrial creatine kinase (CKMT1). We discover that, while CKMT1 is highly expressed in primary tumors and promotes cell viability, it is downregulated in metastasis. We further show that CKMT1 downregulation, as seen in breast cancer metastasis, drives up mitochondrial reactive oxygen species (ROS) levels. CKMT1 downregulation contributes to the migratory and invasive potential of cells by ROS-induced upregulation of adhesion and degradative factors, which can be reversed by antioxidant treatment. Our study thus reconciles conflicting evidence about the roles of metabolites in the creatine metabolic pathway in breast cancer progression and reveals that tight, context-dependent regulation of CKMT1 expression facilitates cell viability, cell migration, and cell invasion, which are hallmarks of metastatic spread.

Integrated analysis reveals FLI1 regulates the tumor immune microenvironment via its cell-type-specific expression and transcriptional regulation of distinct target genes of immune cells in breast cancer.

BACKGROUND: Immunotherapy is a practical therapeutic approach in breast cancer (BRCA), and the role of FLI1 in immune regulation has gradually been unveiled. However, the specific role of FLI1 in BRCA was conflicted; thus, additional convincing evidence is needed. METHODS: We explored the upstream regulation of FLI1 expression via summary data-based Mendelian randomization (SMR) analysis and ncRNA network construction centering on FLI1 using BRCA genome-wide association study (GWAS) summary data with expression quantitative trait loci (eQTLs) and DNA methylation quantitative trait loci (mQTLs) from the blood and a series of in silico analyses, respectively. We illuminated the downstream function of FLI1 in immune regulation by integrating a series of analyses of single-cell RNA sequence data (scRNA-seq). RESULTS: We verified a causal pathway from FLI1 methylation to FLI1 gene expression to BRCA onset and demonstrated that FLI1 was downregulated in BRCA. FLI1, a transcription factor, served as myeloid and T cells' communication regulator by targeting immune-related ligands and receptor transcription in BRCA tissues. We constructed a ceRNA network centering on FLI1 that consisted of three LncRNAs (CKMT2-AS1, PSMA3-AS1, and DIO3OS) and a miRNA (hsa-miR-324-5p), and the expression of FLI1 was positively related to a series of immune-related markers, including immune cell infiltration, biomarkers of immune cells, and immune checkpoints. CONCLUSION: Low-methylation-induced or ncRNA-mediated downregulation of FLI1 is associated with poor prognosis, and FLI1 might regulate the tumor immune microenvironment via a cell-type-specific target genes manner in BRCA.

Prognostic value of mitochondrial CKMT2 in Pan-cancer and its tumor immune correlation analysis.

Mitochondrial metabolism has been shown to play a key role in immune cell survival and function, but mitochondrial creatine kinase 2 (CKMT2) has been relatively little studied about tumor immunity. We aimed to explore the prognostic value of CKMT2 in 33 cancer types and investigate its potential immune function. We used a range of bioinformatics approaches to explore the potential carcinogenic role of CKMT2 in multiple cancers. CKMT2 was lowly expressed in 14 tumor tissues and highly expressed in 4 tumor tissues. Immunohistochemical assays showed overexpression of CKMT2 in colon cancer and rectal cancer. CKMT2 overexpression was positively correlated with the prognosis of lung adenocarcinoma and prostate cancer. CKMT2 overexpression is mainly enriched in the adaptive immune system and immune regulatory pathways of immunoglobulins. Seven cancers were positively correlated with low CKMT2 expression in tumor microenvironment analysis. Among the five cancers, low expression of CKMT2 resulted in better immunotherapy treatment outcomes. There was a strong correlation between CKMT2 and most immune-related genes in specific cancer types. CKMT2 plays an important role in tumorigenesis and cancer immunity and can be used as a prognostic biomarker and potential target for cancer immunotherapy.

Construction of a 5-gene prognostic signature based on oxidative stress related genes for predicting prognosis in osteosarcoma.

BACKGROUND: The understanding of the complex biological scenario of osteosarcoma will open the way to identifying new strategies for its treatment. Oxidative stress is a cancer-related biological scenario. At present, it is not clear the oxidative stress genes in affecting the prognosis and progression of osteosarcoma, the underlying mechanism as well as their impact on the classification of osteosarcoma subtypes. METHODS: We selected samples and sequencing data from TARGET data set and GSE21257 data set, and downloaded oxidative stress related-genes (OSRGs) from MsigDB. Univariate Cox analysis of OSRG was conducted using TARGET data, and the prognostic OSRG was screened to conduct unsupervised clustering analysis to identify the molecular subtypes of osteosarcoma. Through least absolute shrinkage and selection operator (LASSO) regression analysis and COX regression analysis of differentially expressed genes (DEGs) between subgroups, a risk assessment system for osteosarcoma was developed. RESULTS: 45 prognosis-related OSRGs genes were acquired, and two molecular subtypes of osteosarcoma were clustered. C2 cluster displayed prolonged overall survival (OS) accompanied with high degree of immune infiltration and enriched immune pathways. While cell cycle related pathways were enriched in C2 cluster. Based on DEGs between subgroups and Lasso analysis, 5 hub genes (ZYX, GJA5, GAL, GRAMD1B, and CKMT2) were screened to establish a robust prognostic risk model independent of clinicopathological features. High-risk group had more patients with cancer metastasis and death as well as C1 subtype with poor prognosis. Low-risk group exhibited favorable OS and high immune infiltration status. Additionally, the risk assessment system was optimized by building decision tree and nomogram. CONCLUSIONS: This study defined two molecular subtypes of osteosarcoma with different prognosis and tumor immune microenvironment status based on the expression of OSRGs, and provided a new risk assessment system for the prognosis of osteosarcoma.

Role of creatine shuttle in colorectal cancer cells.

The creatine shuttle translocates the energy generated by oxidative phosphorylation to the cytoplasm via mitochondrial creatine kinase (MTCK) and creatine kinase B (CKB) in the cytoplasm. It is not apparent how the creatine shuttle is related to cancer. Here, we analyzed the expression and function of CKB and MTCK in colorectal cancer (CRC) and investigated the role of the creatine shuttle in CRC. Compared with normal mucosa, 184 CRC tissues had higher levels of CKB and MTCK, and these levels were associated with histological grade, tumor invasion, and distant metastasis. CK inhibitor dinitrofluorobenzene (DNFB) on CRC cell lines HT29 and CT26 inhibited cell proliferation and stemness to less than 2/3 and 1/20 of their control levels, respectively. In this treatment, the production of reactive oxygen species increased, mitochondrial respiration decreased, and mitochondrial volume and membrane potential decreased. In a syngeneic BALB/c mouse model using CT26 cells pretreated with DNFB, peritoneal metastasis was suppressed to 70%. Phosphorylation of EGFR, AKT, and ERK1/2 was inhibited in DNFB-treated tumors. High ATP concentrations prevented EGFR phosphorylation in HT29 cells following DNFB treatment, CKB or MTCK knockdown, and cyclocreatine administration. Despite not being immunoprecipitated, CKB and EGFR were brought closer together by EGF stimulation. These findings imply that blocking the creatine shuttle decreases the energy supply, suppresses oxidative phosphorylation, and blocks ATP delivery to phosphorylation signals, preventing signal transduction. These findings highlight the critical role of the creatine shuttle in cancer cells and suggest a potential new cancer treatment target.

Mitochondrial creatine kinase 1 regulates the cell cycle in non-small cell lung cancer via activation of cyclin-dependent kinase 4.

BACKGROUND: Non-small cell lung cancer (NSCLC) is the main type of the most common malignant tumor in the world. Previous studies have shown that the expression level of mitochondrial creatine kinase 1 (CKMT1) is abnormal in NSCLC, but the mechanism of its effect remains unclear. Therefore, in this study, we intend to clarify the potential mechanism of CKMT1 in NSCLC and provide the theoretical basis for the clinical application of CKMT1. METHODS: The function of CKMT1 in NSCLC was identified by analyzing the GEO dataset and evaluating using in vitro and in vivo models. Protein mass spectrometry was used to find proteins interacting with CKMT1, and Co-immunoprecipitation (Co-IP) and GST-pull down experiments were used to verify the interaction between proteins. The immunofluorescence (IF) assay was used to explore the functional position of CKMT1 in cells. The effect of CKMT1 expression level on the efficacy of paclitaxel (TAX) in the treatment of NSCLC was analyzed by a combined TAX experiment in vivo and in vitro. RESULTS: CKMT1 expression was increased in NSCLC and CKMT1 promoted the proliferation of NSCLC cells in vitro and in vivo. CKMT1 knockdown resulted in a significantly increased G0/G1 fraction and decreased S phase cell fraction, indicating G1 phase arrest. Mechanically, the cyclin-dependent kinase 4 (CDK4) was identified to interact with CKMT1, and the crucial binding areas were focused on the DH domain of CKMT1 and the N- and C-terminal of CDK4. A fraction of the CDK4 proteins colocalize and interact with the CKMT1 at mitochondria, the level of phosphorylated CDK4 was regulated by CKMT1. Hence, the decrease in CKMT1 expression level could increase the antitumor effect of G2/M cell cycle antagonist-TAX in NSCLC in vitro and in vivo. CONCLUSIONS: CKMT1 could interact with CDK4 in mitochondria and regulate the phosphorylated level of CDK4, thus contributing to the proliferation and cell cycle transition of NSCLC cells. And CKMT1 could be a potential target to improve the sensitivity of chemotherapy based on TAX.

Cuproptosis-Related LncRNA-Based Prediction of the Prognosis and Immunotherapy Response in Papillary Renal Cell Carcinoma.

Cuproptosis, a new cell death pattern, is promising as an intervention target to treat tumors. Abnormal long non-coding RNA (lncRNA) expression is closely associated with the occurrence and development of papillary renal cell carcinoma (pRCC). However, cuproptosis-related lncRNAs (CRLs) remain largely unknown as prognostic markers for pRCC. We aimed to forecast the prognosis of pRCC patients by constructing models according to CRLs and to examine the correlation between the signatures and the inflammatory microenvironment. From the Cancer Genome Atlas (TCGA), RNA sequencing, genomic mutations and clinical data of TCGA-KIRP (Kidney renal papillary cell carcinoma) were analyzed. Randomly selected pRCC patients were allotted to the training and testing sets. To determine the independent prognostic impact of the training characteristic, the least absolute shrinkage and selection operator (LASSO) algorithm was utilized, together with univariate and multivariate Cox regression models. Further validation was performed in the testing and whole cohorts. External datasets were utilized to verify the prognostic value of CRLs as well. The CRLs prognostic features in pRCC were established based on the five CRLs (AC244033.2, LINC00886, AP000866.1, MRPS9-AS1 and CKMT2-AS1). The utility of CRLs was evaluated and validated in training, testing and all sets on the basis of the Kaplan-Meier (KM) survival analysis. The risk score could be a robust prognostic factor to forecast clinical outcomes for pRCC patients by the LASSO algorithm and univariate and multivariate Cox regression. Analysis of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) data demonstrated that differentially expressed genes (DEGs) are primarily important for immune responses and the PI3K-Akt pathway. Arachidonic acid metabolism was enriched in the high-risk set by Gene Set Enrichment Analysis (GSEA). In addition, Tumor Immune Dysfunction and Exclusion (TIDE) analysis suggested that there was a high risk of immune escape in the high-risk cohort. The immune functions of the low- and high-risk sets differed significantly based on immune microenvironment analysis. Finally, four drugs were screened with a higher sensitivity to the high-risk set. Taken together, a novel model according to five CRLs was set up to forecast the prognosis of pRCC patients, which provides a potential strategy to treat pRCC by a combination of cuproptosis and immunotherapy.

Identification of Biomarkers Associated with the Prognoses of Colorectal Cancer Patients.

INTRODUCTION: Colorectal cancer (CRC) is a common cancer. As metastasis and recurrence are main causes of CRC death, it is of great significance to find prognostic biomarkers. METHODS: Data related to CRC were collected from GEO database. The patients were grouped based on clinical information, and the differentially expressed genes (DEGs) were obtained by differential analysis. GO and KEGG pathway enrichment analyses were conducted based on DEGs. Cox combined with LASSO regression analysis was applied to screen out the key genes that used to build the prognostic model. Survival curve and receiver operating characteristic curve were employed to evaluate the validity and reliability of the model. Cox regression analysis was applied to determine the independence of risk score. GSEA and GSVA analyses were performed on patients with different risks according to the risk model scores, and the prognostic nomogram was plotted combined with clinical data. Also, qRT-PCR was applied to examine the expression status of the screened signatures in clinical cases. RESULTS: We obtained 302 DEGs by dividing CRC patients into early-stage and advanced-stage groups. The results of enrichment analyses demonstrated that the DEGs were mainly concentrated in tissues of extracellular matrix, epithelial cell proliferation, and cell adhesion-related pathways. Regression identified 9 hub genes notably correlated with prognosis, including CLK1, SLC2A3, LIPG, EPHB2, ATOH1, PLCB4, GZMB, CKMT2, and CXCL11. The validation of the risk model proved that the risk model was accurate and could independently determine the prognosis of patients. Finally, differences were found in pathway activity of extracellular matrix secretion, plaque secretion, Notch signaling pathway, and tight junctions in high-risk and low-risk patients. In addition to LIPG and CKMT2, other feature genes were notably overexpressed in CRC tumor tissues. CONCLUSION: The results proved that the expression levels of the 9 biomarkers could be used to predict the prognosis of CRC patients.

Ubiquitous mitochondrial creatine kinase promotes the progression of gastric cancer through a JNK-MAPK/JUN/HK2 axis regulated glycolysis.

BACKGROUND: Ubiquitous mitochondrial creatine kinase (uMtCK) transfers high-energy phosphates from mitochondrially generated ATP to creatine to generate phosphocreatine. uMtCK overexpression has been reported in several malignant tumors, however, the clinical significance and impact of uMtCK in gastric cancer (GC) has not been comprehensively studied. METHODS: We first examined uMtCK expression in GC by quantitative real-time PCR and western blot assays. Then the clinicopathological significance of aberrant uMtCK expression was determined by immunohistochemical staining in a GC tissue microarray. Kaplan-Meier analysis was used for survival analysis. The biological functions of uMtCK in GC cells were explored by wound-healing, transwell assays and glucose metabolism assays in vitro as well as a liver metastasis model by spleen injection in nude mice in vivo. RESULTS: We verified that the expression of uMtCK was substantially elevated in GC tissues, significantly associating with a poorer prognosis in GC patients, especially for those with advanced stage. In univariate and multivariate analyses, uMtCK expression emerged as an independent prognostic factor for both disease-free survival and overall survival. Functionally, we demonstrated that uMtCK promoted glycolysis in GC cells and facilitated their migration, invasion and liver metastasis in vitro and in vivo. Mechanistically, uMtCK enhanced GC progression in a HK2-dependent glycolysis via acting the JNK-MAPK/JUN signaling pathway. CONCLUSIONS: uMtCK could serve as a novel independent prognostic biomarker as well as potential therapeutic target for GC patients, particularly for GC patients with an advanced UICC stage and tumor recurrence.

Interaction between glycolysis‒cholesterol synthesis axis and tumor microenvironment reveal that gamma-glutamyl hydrolase suppresses glycolysis in colon cancer.

Background: Metabolic reprogramming is a feature of cancer. However, colon cancer subtypes based on the glycolysis‒cholesterol synthesis axis have not been identified, and little is known about connections between metabolic features and the tumor microenvironment. Methods: Data for 430 colon cancer cases were extracted from The Cancer Genome Atlas, including transcriptome data, clinical information, and survival outcomes. Glycolysis and cholesterol synthesis-related gene sets were obtained from the Molecular Signatures Database for a gene set variation analysis. The relationship between the genomic landscape and immune landscape were investigated among four metabolic subtypes. Hub genes were determined. The clinical significance of candidate hub gene was evaluated in 264 clinical samples and potential functions were validated in vitro and in vivo. Results: Colon cancer cases were clustered into four metabolic subtypes: quiescent, glycolytic, cholesterogenic, and mixed. The metabolic subtypes differed with respect to the immune score, stromal score, and estimate score using the ESTIMATE algorithm, cancer-immunity cycle, immunomodulator signatures, and signatures of immunotherapy responses. Patients in the cholesterogenic group had better survival outcomes than those for other subtypes, especially glycolytic. The glycolytic subtype was related to unfavorable clinical characteristics, including high mutation rates in TTN, APC, and TP53, high mutation burden, vascular invasion, right colon cancer, and low-frequency microsatellite instability. GGH, CACNG4, MME, SLC30A2, CKMT2, SYN3, and SLC22A31 were identified as differentially expressed both in glycolytic-cholesterogenic subgroups as well as between colon cancers and healthy samples, and were involved in glycolysis‒cholesterol synthesis. GGH was upregulated in colon cancer; its high expression was correlated with CD4+ T cell infiltration and longer overall survival and it was identified as a favorable independent prognostic factor. The overexpression of GGH in colon cancer-derived cell lines (SW48 and SW480) inhibited PKM, GLUT1, and LDHA expression and decreased the extracellular lactate content and intracellular ATP level. The opposite effects were obtained by GGH silencing. The phenotype associated with GGH was also validated in a xenograft nude mouse model. Conclusions: Our results provide insight into the connection between metabolism and the tumor microenvironment in colon cancer and provides preliminary evidence for the role of GGH, providing a basis for subsequent studies.

An integrative pan-cancer analysis of molecular characteristics and oncogenic role of mitochondrial creatine kinase 1A (CKMT1A) in human tumors.

In recent years, several studies have suggested that mitochondrial creatine kinase 1A (CKMT1A) plays a key role in various cancer types. However, there is still a lack of systematic understanding of the contribution of CKMT1A in different types of cancer. Therefore, this study aims to explore the potential role of CKMT1A in human tumors. Firstly, we evaluated the expression level of CKMT1A in 33 types of tumors. Secondly, we used the GEPIA2 and Kaplan-Meier plotter to explore the relationship between CKMT1A expression and survival prognosis. Furthermore, the genetic alterations of CKMT1A were analyzed by the cBioPortal web. In addition, we performed immune infiltration analysis and gene enrichment pathway analysis. CKMT1A was highly expressed in most types of cancers and there was a significant correlation between CKMT1A expression and the prognosis of patients for certain tumors. Non-Small Cell Lung Cancer cases with altered CKMT1A showed a poorer overall survival. CKMT1A expression was negatively correlated with the infiltration of cancer-associated fibroblasts in most tumors. We also found that its expression was negatively associated with CD8+ T-cell infiltration in several tumors. Furthermore, enrichment analysis revealed that "Glycolysis/ Gluconeogenesis" and "metabolic pathways" functions were involved in the functional mechanism of CKMT1A. Taken together, our studies will provide a relatively clear and integrative understanding of the role of CKMT1A across different tumors. All these findings will lay a solid foundation for further molecular assays of CKMT1A in tumorigenesis and provide the rationale for developing novel therapeutic strategies.

Post-COVID Complications after Pressure Ulcer Surgery in Patients with Spinal Cord Injury Associate with Creatine Kinase Upregulation in Adipose Tissue.

The risk of complications following surgical procedures is significantly increased in patients with SARS-CoV-2 infection. However, the mechanisms underlying these correlations are not fully known. Spinal cord injury (SCI) patients who underwent reconstructive surgery for pressure ulcers (PUs) before and during the COVID-19 pandemic were included in this study. The patient's postoperative progression was registered, and the subcutaneous white adipose tissue (s-WAT) surrounding the ulcers was analyzed by proteomic and immunohistochemical assays to identify the molecular/cellular signatures of impaired recovery. Patients with SCI and a COVID-19-positive diagnosis showed worse recovery and severe postoperative complications, requiring reintervention. Several proteins were upregulated in the adipose tissue of these patients. Among them, CKMT2 and CKM stood out, and CKM increased for up to 60 days after the COVID-19 diagnosis. Moreover, CKMT2 and CKM were largely found in MGCs within the s-WAT of COVID patients. Some of these proteins presented post-translational modifications and were targeted by autoantibodies in the serum of COVID patients. Overall, our results indicate that CKMT2, CKM, and the presence of MGCs in the adipose tissue surrounding PUs in post-COVID patients could be predictive biomarkers of postsurgical complications. These results suggest that the inflammatory response in adipose tissue may underlie the defective repair seen after surgery.

Mitochondrial Creatine Kinase Attenuates Pathologic Remodeling in Heart Failure.

BACKGROUND: Abnormalities in cardiac energy metabolism occur in heart failure (HF) and contribute to contractile dysfunction, but their role, if any, in HF-related pathologic remodeling is much less established. CK (creatine kinase), the primary muscle energy reserve reaction which rapidly provides ATP at the myofibrils and regenerates mitochondrial ADP, is down-regulated in experimental and human HF. We tested the hypotheses that pathologic remodeling in human HF is related to impaired cardiac CK energy metabolism and that rescuing CK attenuates maladaptive hypertrophy in experimental HF. METHODS: First, in 27 HF patients and 14 healthy subjects, we measured cardiac energetics and left ventricular remodeling using noninvasive magnetic resonance 31P spectroscopy and magnetic resonance imaging, respectively. Second, we tested the impact of metabolic rescue with cardiac-specific overexpression of either Ckmyofib (myofibrillar CK) or Ckmito (mitochondrial CK) on HF-related maladaptive hypertrophy in mice. RESULTS: In people, pathologic left ventricular hypertrophy and dilatation correlate closely with reduced myocardial ATP levels and rates of ATP synthesis through CK. In mice, transverse aortic constriction-induced left ventricular hypertrophy and dilatation are attenuated by overexpression of CKmito, but not by overexpression of CKmyofib. CKmito overexpression also attenuates hypertrophy after chronic isoproterenol stimulation. CKmito lowers mitochondrial reactive oxygen species, tissue reactive oxygen species levels, and upregulates antioxidants and their promoters. When the CK capacity of CKmito-overexpressing mice is limited by creatine substrate depletion, the protection against pathologic remodeling is lost, suggesting the ADP regenerating capacity of the CKmito reaction rather than CK protein per se is critical in limiting adverse HF remodeling. CONCLUSIONS: In the failing human heart, pathologic hypertrophy and adverse remodeling are closely related to deficits in ATP levels and in the CK energy reserve reaction. CKmito, sitting at the intersection of cardiac energetics and redox balance, plays a crucial role in attenuating pathologic remodeling in HF. Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT00181259.

System analysis of VEGFA in renal cell carcinoma: The expression, prognosis, gene regulation network and regulation targets.

BACKGROUND: VEGFA is one of the most important regulators of angiogenesis and plays a crucial role in cancer angiogenesis and progression. Recent studies have highlighted a relationship between VEGFA expression and renal cell carcinoma occurrence. However, the expression level, gene regulation network, prognostic value, and target prediction of VEGFA in renal cell carcinoma remain unclear. Therefore, system analysis of the expression, gene regulation network, prognostic value, and target prediction of VEGFA in patients with renal cell carcinoma is of great theoretical significance as there is a clinical demand for the discovery of new renal cell carcinoma treatment targets and strategies to further improve renal cell carcinoma treatment efficacy. METHODS: This study used multiple free online databases, including cBioPortal, TRRUST, GeneMANIA, GEPIA, Metascape, UALCAN, LinkedOmics, Metascape, and TIMER for the abovementioned analysis. RESULTS: VEGFA was upregulated in patients with kidney renal clear cell carcinoma (KIRC) and kidney chromophobe (KICH), and downregulated in patients with kidney renal papillary cell carcinoma (KIRP). Moreover, genetic alterations of VEGFA were found in patients with renal cell carcinoma as follows: 4% (KIRC), 8% (KICH), and 4% (KIRP). The promoter methylation of VEGFA was lower and higher in patients with clinical stages of KIRC and stage 1 KIRP, respectively. VEGFA expression significantly correlated with KIRC and KIRP pathological stages. Furthermore, patients with KICH and KIRP having low VEGFA expression levels had a longer survival than those having high VEGFA expression levels. VEGFA and its neighboring genes functioned in the regulation of protein methylation and glycosylation, as well as muscle fiber growth and differentiation in patients with renal cell carcinoma. Gene Ontology enrichment analysis revealed that the functions of VEGFA and its neighboring genes in patients with renal cell carcinoma are mainly related to cell adhesion molecule binding, catalytic activity, acting on RNA, ATPase activity, actin filament binding, protease binding, transcription coactivator activity, cysteine-type peptidase activity, and calmodulin binding. Transcription factor targets of VEGFA and its neighboring genes in patients with renal cell carcinoma were found: HIF1A, TFAP2A, and ESR1 in KIRC; STAT3, NFKB1, and HIPK2 in KICH; and FOXO3, TFAP2A, and ETS1 in KIRP. We further explored the VEGFA-associated kinase (ATM in KICH as well as CDK1 and AURKB in KIRP) and VEGFA-associated microRNA (miRNA) targets (MIR-21 in KICH as well as MIR-213, MIR-383, and MIR-492 in KIRP). Furthermore, the following genes had the strongest correlation with VEGFA expression in patients with renal cell carcinoma: NOTCH4, GPR4, and TRIB2 in KIRC; CKMT2, RRAGD, and PPARGC1A in KICH; and FLT1, C6orf223, and ESM1 in KIRP. VEGFA expression in patients with renal cell carcinoma was positively associated with immune cell infiltration, including CD8+T cells, CD4+T cells, macrophages, neutrophils, and dendritic cells. CONCLUSIONS: This study revealed VEGFA expression and potential gene regulatory network in patients with renal cell carcinoma, thereby laying a foundation for further research on the role of VEGFA in renal cell carcinoma occurrence. Moreover, the study provides new renal cell carcinoma therapeutic targets and prognostic biomarkers as a reference for fundamental and clinical research.

CKMT1B is a potential prognostic biomarker and associated with immune infiltration in Lower-grade glioma.

BACKGROUND: Lower-grade glioma (LGG) is the most common histology identified in glioma. CKMT1B has not been investigated in glioma. The purpose of this research was to investigate the prognostic value of CKMT1B and its correlation with immune infiltration in LGG. METHODS: We used Gene Expression Profiling Interactive Analysis (GEPIA) to analyze the expression of CKMT1B in LGG. Univariate and multivariate Cox regression analyses were used to assess the effect of CKMT1B expression and screened variables on survival. The correlation between CKMT1B and immune infiltration was evaluated by TIMER and CIBERSORT. Moreover, the possible biological functions of CKMT1B were studied by GSEA. The statistical analysis was conducted by R software. RESULTS: The expression of CKMT1B was significantly lower than the normal samples in LGG. Low expression of CKMT1B predicts a worse prognosis. Multivariate Cox analyses revealed that CKMT1B might be an independent favorable prognostic indicator. TIMER analysis revealed that CKMT1B expression level was related to immune infiltration. CIBERSORT analysis showed that CKMT1B expression was positively related to the infiltration level of activated mast cells and negatively related to macrophage M2 in LGG. Moreover, GESA showed that multiple cancer-related and immune-related gene sets were enriched in the low-CKMT1B group in the top 5 of the most significant differences. CONCLUSION: CKMT1B is a prognostic biomarker with potential applications and associated with immune infiltration in Lower-grade glioma.

Zbtb20 deficiency causes cardiac contractile dysfunction in mice.

The zinc-finger protein ZBTB20 regulates development and metabolism in multiple systems, and is essential for postnatal survival in mice. However, its potential role in the cardiovascular system remains undefined. Here, we demonstrate that ZBTB20 is critically involved in the regulation of cardiac contractility and blood pressure in mice. At the age of 16 days, the relatively healthy Zbtb20-null mice exhibited hypotension without obvious change of heart rate or other evidence for heart failure. Moreover, Zbtb20 deletion led to a marked reduction in heart size, left ventricular wall thickness, and cell size of cardiomyocytes, which was largely proportional to the decreased body growth. Notably, echocardiographic and hemodynamic analyses showed that cardiac contractility was greatly impaired in the absence of ZBTB20. Mechanistically, ZBTB20 deficiency decreased cardiac ATP contents, and compromised the enzyme activity of mitochondrial complex I in heart as well as L-type calcium current density in cardiomyocytes. Furthermore, the developmental activation of some mitochondrial function-related genes was significantly attenuated in Zbtb20-null myocardium, which included Hspb8, Ckmt2, Cox7a1, Tfrc, and Ogdhl. Put together, these results suggest that ZBTB20 plays a crucial role in the regulation of heart development, energy metabolism, and contractility.

Structure and function of resistance arteries from BB-creatine kinase and ubiquitous Mt-creatine kinase double knockout mice.

Increasing evidence indicates that the enzyme creatine kinase (CK) is intimately involved in microvascular contractility. The mitochondrial isoenzyme catalyses phosphocreatine synthesis from ATP, while cytoplasmic CK, predominantly the BB isoenzyme in vascular tissue, is tightly bound near myosin ATPase, where it favours ATP production from phosphocreatine to metabolically support vascular contractility. However, the effect of CK gene inactivation on microvascular function is hitherto unknown. We studied functional and structural parameters of mesenteric resistance arteries isolated from 5 adult male mice lacking cytoplasmic BB-CK and ubiquitous mitochondrial CK (CK-/-) vs 6 sex/age-matched controls. Using a Mulvany Halpern myograph, we assessed the acute maximum contractile force with 125 mM K+ and 10-5 M norepinephrine, and the effect of two inhibitors, dinitrofluorobenzene, which inhibits phosphotransfer enzymes (0.1 µM), and the specific adenylate kinase inhibitor P1, P5-di(adenosine 5') pentaphosphate (10-6 to 10-5 M). WT and CK-/- did not significantly differ in media thickness, vascular elasticity parameters, or acute maximum contractile force. CK-/- arteries displayed greater reduction in contractility after dinitrofluorobenzene 38%; vs 14% in WT; and after AK inhibition, 14% vs 5.5% in WT, and displayed abnormal mitochondria, with a partial loss of the inner membrane. Thus, CK-/- mice display a surprisingly mild phenotype in vascular dysfunction. However, the mitochondrial abnormalities and greater effect of inhibitors on contractility may reflect a compromised energy metabolism. In CK-/- mice, compensatory mechanisms salvage energy metabolism, as described for other CK knock-out models.

The relationship between skin ultrasound images and muscle damage using skin blotting in wheelchair basketball athletes.

STUDY DESIGN: Secondary analysis of a cross-sectional observation study. OBJECTIVES: To determine the relationship between skin ultrasound images and muscle damage in wheelchair basketball athletes, using skin blotting examinations of the ischial regions. SETTING: Community, Japan. METHODS: Fourteen elite wheelchair basketball athletes were recruited. We obtained data regarding participants' characteristics. We undertook ultrasonographic images and quantitative skin blotting of the ischial region before and after training, and after rest. RESULTS: We identified Category II and III pressure injuries in 2 of the 12 participants. Structural features were classified into four categories based on ultrasonographic features, namely, normal skin structure, unclear superficial and deep fascia, cloudy fat layer, and fat infiltration and low-echoic lesion/anechoic lesions. The muscle-type creatinine kinase (CK-M) level (median [interquartile range: IQR], 2.98 [2.80-3.47]) in the fat infiltration and low-echoic lesion/anechoic lesion group was significantly higher (1.43 [1.41-1.49]) than in a nonfat infiltration and low-echoic lesion/anechoic lesion group after training (p = 0.03). The interleukin-6 (IL-6) level (median [IQR], 23.5 [16.15-58.97]) in the fat infiltration and low-echoic lesion/anechoic lesion group was significantly higher (1.94 [1.74-4.44]) than in the nonfat infiltration and low-echoic lesion/anechoic lesion group after rest (mean difference = -25.4, 95% CI -61.1 to 10.7, p = 0.03). CONCLUSIONS: The combination of ultrasonographic images and skin blotting using CK-M and IL-6, could detect early deep tissue damage in wheelchair athletes. These techniques could be potentially useful in the treatment and prevention of pressure injuries. SPONSORSHIP: This study was supported in part by YAMAHA Motor Foundation for Sports.

Overexpression of mitochondrial creatine kinase preserves cardiac energetics without ameliorating murine chronic heart failure.

Mitochondrial creatine kinase (Mt-CK) is a major determinant of cardiac energetic status and is down-regulated in chronic heart failure, which may contribute to disease progression. We hypothesised that cardiomyocyte-specific overexpression of Mt-CK would mitigate against these changes and thereby preserve cardiac function. Male Mt-CK overexpressing mice (OE) and WT littermates were subjected to transverse aortic constriction (TAC) or sham surgery and assessed by echocardiography at 0, 3 and 6 weeks alongside a final LV haemodynamic assessment. Regardless of genotype, TAC mice developed progressive LV hypertrophy, dilatation and contractile dysfunction commensurate with pressure overload-induced chronic heart failure. There was a trend for improved survival in OE-TAC mice (90% vs 73%, P = 0.08), however, OE-TAC mice exhibited greater LV dilatation compared to WT and no functional parameters were significantly different under baseline conditions or during dobutamine stress test. CK activity was 37% higher in OE-sham versus WT-sham hearts and reduced in both TAC groups, but was maintained above normal values in the OE-TAC hearts. A separate cohort of mice received in vivo cardiac 31P-MRS to measure high-energy phosphates. There was no difference in the ratio of phosphocreatine-to-ATP in the sham mice, however, PCr/ATP was reduced in WT-TAC but preserved in OE-TAC (1.04 ± 0.10 vs 2.04 ± 0.22; P = 0.007). In conclusion, overexpression of Mt-CK activity prevented the changes in cardiac energetics that are considered hallmarks of a failing heart. This had a positive effect on early survival but was not associated with improved LV remodelling or function during the development of chronic heart failure.