Inhibition of Pyruvate Dehydrogenase Kinase 4 Attenuates Myocardial and Mitochondrial Injury in Sepsis-Induced Cardiomyopathy.

BACKGROUND: Sepsis-induced cardiomyopathy (SIC) is a cardiac dysfunction caused by sepsis, with mitochondrial dysfunction being a critical contributor. Pyruvate dehydrogenase kinase 4 (PDK4) is a kinase of pyruvate dehydrogenase with multifaceted actions in mitochondrial metabolism. However, its role in SIC remains unknown. METHODS: Serum PDK4 levels were measured and analyzed in 27 children with SIC, 30 children with sepsis, and 29 healthy children. In addition, for mice exhibiting SIC, the effects of PDK4 knockdown or inhibition on the function and structure of the myocardium and mitochondria were assessed. RESULTS: The findings from the analysis of children with SIC revealed that PDK4 was significantly elevated and correlated with disease severity and organ injury. Nonsurvivors displayed higher serum PDK4 levels than survivors. Furthermore, mice with SIC benefited from PDK4 knockdown or inhibition, showing improved myocardial contractile function, reduced myocardial injury, and decreased mitochondrial structural injury and dysfunction. In addition, inhibition of PDK4 decreased the inhibitory phosphorylation of PDHE1α (pyruvate dehydrogenase complex E1 subunit α) and improved abnormal pyruvate metabolism and mitochondrial dysfunction. CONCLUSIONS: PDK4 is a potential biomarker for the diagnosis and prognosis of SIC. In experimental SIC, PDK4 promoted mitochondrial dysfunction with increased phosphorylation of PDHE1α and abnormal pyruvate metabolism.

Unveiling the PDK4-centered rituximab-resistant mechanism in DLBCL: the potential of the "Smart" exosome nanoparticle therapy.

BACKGROUND: Diffuse large B-cell lymphoma (DLBCL) represents a prevalent malignant tumor, with approximately 40% of patients encountering treatment challenges or relapse attributed to rituximab resistance, primarily due to diminished or absent CD20 expression. Our prior research identified PDK4 as a key driver of rituximab resistance through its negative regulation of CD20 expression. Further investigation into PDK4's resistance mechanism and the development of advanced exosome nanoparticle complexes may unveil novel resistance targets and pave the way for innovative, effective treatment modalities for DLBCL. METHODS: We utilized a DLBCL-resistant cell line with high PDK4 expression (SU-DHL-2/R). We infected it with short hairpin RNA (shRNA) lentivirus for RNA sequencing, aiming to identify significantly downregulated mRNA in resistant cells. Techniques including immunofluorescence, immunohistochemistry, and Western blotting were employed to determine PDK4's localization and expression in resistant cells and its regulatory role in phosphorylation of Histone deacetylase 8 (HDAC8). Furthermore, we engineered advanced exosome nanoparticle complexes, aCD20@ExoCTX/siPDK4, through cellular, genetic, and chemical engineering methods. These nanoparticles underwent characterization via Dynamic Light Scattering (DLS) and Transmission Electron Microscopy (TEM), and their cellular uptake was assessed through flow cytometry. We evaluated the nanoparticles' effects on apoptosis in DLBCL-resistant cells and immune cells using CCK-8 assays and flow cytometry. Additionally, their capacity to counteract resistance and exert anti-tumor effects was tested in a resistant DLBCL mouse model. RESULTS: We found that PDK4 initiates HDAC8 activation by phosphorylating the Ser-39 site, suppressing CD20 protein expression through deacetylation. The aCD20@ExoCTX/siPDK4 nanoparticles served as effective intracellular delivery mechanisms for gene therapy and monoclonal antibodies, simultaneously inducing apoptosis in resistant DLBCL cells and triggering immunogenic cell death in tumor cells. This dual action effectively reversed the immunosuppressive tumor microenvironment, showcasing a synergistic therapeutic effect in a subcutaneous mouse tumor resistance model. CONCLUSIONS: This study demonstrates that PDK4 contributes to rituximab resistance in DLBCL by modulating CD20 expression via HDAC8 phosphorylation. The designed exosome nanoparticles effectively overcome this resistance by targeting the PDK4/HDAC8/CD20 pathway, representing a promising approach for drug delivery and treating patients with Rituximab-resistant DLBCL.

Ginsenoside Rh2 shifts tumor metabolism from aerobic glycolysis to oxidative phosphorylation through regulating the HIF1-α/PDK4 axis in non-small cell lung cancer.

BACKGROUND: Ginsenoside Rh2 (G-Rh2), a steroidal compound extracted from roots of ginseng, has been extensively studied in tumor therapy. However, its specific regulatory mechanism in non-small cell lung cancer (NSCLC) is not well understood. Pyruvate dehydrogenase kinase 4 (PDK4), a central regulator of cellular energy metabolism, is highly expressed in various malignant tumors. We investigated the impact of G-Rh2 on the malignant progression of NSCLC and how it regulated PDK4 to influence tumor aerobic glycolysis and mitochondrial function. METHOD: We examined the inhibitory effect of G-Rh2 on NSCLC through I proliferation assay, migration assay and flow cytometry in vitro. Subsequently, we verified the ability of G-Rh2 to inhibit tumor growth and metastasis by constructing subcutaneous tumor and metastasis models in nude mice. Proteomics analysis was conducted to analyze the action pathways of G-Rh2. Additionally, we assessed glycolysis and mitochondrial function using seahorse, PET-CT, Western blot, and RT-qPCR. RESULT: Treatment with G-Rh2 significantly inhibited tumor proliferation and migration ability both in vitro and in vivo. Furthermore, G-Rh2 inhibited the tumor's aerobic glycolytic capacity, including glucose uptake and lactate production, through the HIF1-α/PDK4 pathway. Overexpression of PDK4 demonstrated that G-Rh2 targeted the inhibition of PDK4 expression, thereby restoring mitochondrial function, promoting reactive oxygen species (ROS) accumulation, and inducing apoptosis. When combined with sodium dichloroacetate, a PDK inhibitor, it complemented the inhibitory capacity of PDKs, acting synergistically as a detoxifier. CONCLUSION: G-Rh2 could target and down-regulate the expression of HIF-1α, resulting in decreased expression of glycolytic enzymes and inhibition of aerobic glycolysis in tumors. Additionally, by directly targeting mitochondrial PDK, it elevated mitochondrial oxidative phosphorylation and enhanced ROS accumulation, thereby promoting tumor cells to undergo normal apoptotic processes.

circFTO from M2 macrophage-derived small extracellular vesicles (sEV) enhances NSCLC malignancy by regulation miR-148a-3pPDK4 axis.

BACKGROUND: Accumulation studies found that tumor-associated macrophages (TAMs) are a predominant cell in tumor microenvironment (TME), which function essentially during tumor progression. By releasing bioactive molecules, including circRNA, small extracellular vesicles (sEV) modulate immune cell functions in the TME, thereby affecting non-small cell lung cancer (NSCLC) progression. Nevertheless, biology functions and molecular mechanisms of M2 macrophage-derived sEV circRNAs in NSCLC are unclear. METHODS: Cellular experiments were conducted to verify the M2 macrophage-derived sEV (M2-EV) roles in NSCLC. Differential circRNA expression in M0 and M2-EV was validated by RNA sequencing. circFTO expression in NSCLC patients and cells was investigated via real-time PCR and FISH. The biological mechanism of circFTO in NSCLC was validated by experiments. Our team isolated sEV from M2 macrophages (M2Ms) and found that M2-EV treatment promoted NSCLC CP, migration, and glycolysis. RESULTS: High-throughput sequencing found that circFTO was highly enriched in M2-EV. FISH and RT-qPCR confirmed that circFTO expression incremented in NSCLC tissues and cell lines. Clinical studies confirmed that high circFTO expression correlated negatively with NSCLC patient survival. Luciferase reporter analysis confirmed that miR-148a-3p and PDK4 were downstream targets of circFTO. circFTO knockdown inhibited NSCLC cell growth and metastasis in in vivo experiments. Downregulating miR-148a-3p or overexpressing PDK4 restored the malignancy of NSCLC, including proliferation, migration, and aerobic glycolysis after circFTO silencing. CONCLUSION: The study found that circFTO from M2-EV promoted NSCLC cell progression and glycolysis through miR-148a-3p/PDK4 axis. circFTO is a promising prognostic and diagnostic NSCLC biomarker and has the potential to be a candidate NSCLC therapy target.

PDK4 facilitates fibroblast functions and diabetic wound healing through regulation of HIF-1α protein stability and gene expression.

Fibroblast activation disorder is one of the main pathogenic characteristics of diabetic wounds. Orchestrated fibroblast functions and myofibroblast differentiation are crucial for wound contracture and extracellular matrix (ECM) formation. Pyruvate dehydrogenase kinase 4 (PDK4), a key enzyme regulating energy metabolism, has been implicated in modulating fibroblast function, but its specific role in diabetic wounds remains poorly understood. In this study, we investigated the impact of PDK4 on diabetic wounds and its underlying mechanisms. To assess the effect of PDK4 on human dermal fibroblasts (HDFs), we conducted CCK-8, EdU proliferation assay, wound healing assay, transwell assay, flow cytometry, and western blot analyses. Metabolic shifts were analyzed using the Seahorse XF analyzer, while changes in metabolite expression were measured through LC-MS. Local recombinant PDK4 administration was implemented to evaluate its influence on wound healing in diabetic mice. Finally, we found that sufficient PDK4 expression is essential for a normal wound-healing process, while PDK4 is low expressed in diabetic wound tissues and fibroblasts. PDK4 promotes proliferation, migration, and myofibroblast differentiation of HDFs and accelerates wound healing in diabetic mice. Mechanistically, PDK4-induced metabolic reprogramming increases the level of succinate that inhibits PHD2 enzyme activity, thus leading to the stability of the HIF-1α protein, during which process the elevated HIF-1α mRNA by PDK4 is also indispensable. In conclusion, PDK4 promotes fibroblast functions through regulation of HIF-1α protein stability and gene expression. Local recombinant PDK4 administration accelerates wound healing in diabetic mice.

PRKCA Promotes Mitophagy through the miR-15a-5p/PDK4 Axis to Relieve Sepsis-Induced Acute Lung Injury.

Acute lung injury (ALI) caused by sepsis is a common respiratory critical illness with high morbidity and mortality. Protein kinase C-alpha (PRKCA) plays a protective role in sepsis-induced ALI. However, the detailed molecular mechanism of PRKCA in ALI caused by sepsis is unclear. Animal and cell models of sepsis were established by cecal ligation and puncture (CLP)-surgery and lipopolysaccharide (LPS)/interferon-gamma (IFN-γ) treatment, respectively. Lentivirus transfection was used to overexpress PRKCA. H&E staining and lung injury in CLP-surgery mice were evaluated. Gene expression was evaluated using qPCR and Western blotting. The expression of TNF-α, IL-1ß, and IL-6 was examined using qPCR and ELISA. The expression of LC3 and TOM20 was evaluated using immunofluorescence assays. Cell apoptosis was assessed using a flow cytometry assay. The bond between miR-15a-5p and PDK4 was confirmed by dual-luciferase reporter gene and RNA immunoprecipitation assays. In vivo and in vitro, PRKCA overexpression reduced lung injury to prompt mitophagy and inhibit the inflammatory response, ROS production, and cell apoptosis. miR-15a-5p was highly expressed in macrophages treated with LPS/IFN-γ and was negatively mediated by PRKCA. The overexpression of miR-15a-5p reduced the effects of PRKCA upregulation in macrophages. miR-15a-5p could restrain mitophagy in LPS/IFN-γ-treated macrophages by directly targeting PDK4. Furthermore, PDK4 knockdown reversed the inhibition of cell apoptosis and inflammatory factor release caused by miR-15a-5p silencing. The PRKCA/miR-15a-5p/PDK4 axis alleviated ALI caused by sepsis by promoting mitophagy and repressing anti-inflammatory response.

PPAR/PDK4 pathway is involved in the anticancer effects of cGMP in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is a type of cancer with a high mortality rate. Current treatments for PDACs often have side effects, and drug resistance in cancer stem cells (CSCs) would be also a problem. Cyclic guanosine monophosphate (cGMP) suppresses the mitochondrial function of PDACs and inhibits their CSC properties. Metabolic regulation plays a crucial role in the maintenance of CSC phenotype, and we hypothesized that cGMP induction suppresses cancer stem cell properties in the cancer cell through energy-related signaling pathways. We demonstrated that induction of cGMP upregulated the PPARα/PDK4 pathway and suppressed CSC properties in PDAC, and patients with pancreatic cancer with high PDK4 gene expression had a better prognosis than those with low gene expression. Therefore, these mechanisms may provide new therapeutic targets for the eradication of pancreatic CSCs.

Identification of PDK4 as Hub Gene for Diabetic Nephropathy Using Co-Expression Network Analysis.

INTRODUCTION: Diabetic nephropathy (DN) is related to type 1 and type 2 diabetes. They are the leading cause of end-stage renal disease, but the underling specific pathogenesis of DN is not yet clear. Our study was conducted to explore how DN changed the transcriptome profiles in the kidney. METHODS: The gene expression profile of microdissected glomeruli of 41 type 2 DN patients and 20 healthy controls were included. The sample dataset GSE96804 was obtained from the GEO database. Differentially expressed genes (DEGs) were analyzed in R with the limma package and the important modules were found by weighted gene co-expression network analysis (WGCNA) clustering. The modules were then analyzed based on Gene Ontology (GO) gene set enrichment analysis, and the hub genes were found out. We next validated the hub gene, PDK4, in a cell model of DN. We also constructed the PDK4-related PPI network to investigate the correlation between PDK4 expression and other genes. RESULTS: Heatmap and volcano map were drawn to illustrate the mRNA expression profile of 1,204 DEGs in both samples of DN patients and the control group. Using WGCNA, we selected the blue module in which genes showed the strongest correlation with the phenotype and the smallest p value. We also identified PDK4 as a hub gene. PDK4 expression was upregulated in human diabetic kidney tissue. Moreover, PDK4 was speculated to play a role in glomerular basement membrane development and kidney development according to the enrichment of functions and signaling pathways. Furthermore, PDK4 and two key genes GSTA2 and G6PC protein expression were verified highly expressed in the cell model of DN. CONCLUSION: During the pathogenesis of DN, many genes may change expression in a coordinated manner. The discovery of PDK4 as key gene using WGCNA is of great significance for the development of new treatment strategies to block the development of DN.

MicroRNA-148a-3p in pericyte-derived extracellular vesicles improves erectile function in diabetic mice by promoting cavernous neurovascular regeneration.

BACKGROUND: To investigate the regulatory role of microRNA (miR)-148a-3p in mouse corpus cavernous pericyte (MCPs)-derived extracellular vesicles (EVs) in the treatment of diabetes-induced erectile dysfunction (ED). METHODS: Mouse corpus cavernous tissue was used for MCP primary culture and EV isolation. Small-RNA sequencing analysis was performed to assess the type and content of miRs in MCPs-EVs. Four groups of mice were used: control nondiabetic mice and streptozotocin-induced diabetic mice receiving two intracavernous injections (days - 3 and 0) of phosphate buffered saline, MCPs-EVs transfected with reagent control, or MCPs-EVs transfected with a miR-148a-3p inhibitor. miR-148a-3p function in MCPs-EVs was evaluated by tube-formation assay, migration assay, TUNEL assay, intracavernous pressure, immunofluorescence staining, and Western blotting. RESULTS: We extracted EVs from MCPs, and small-RNA sequencing analysis showed miR-148a-3p enrichment in MCPs-EVs. Exogenous MCPs-EV administration effectively promoted mouse cavernous endothelial cell (MCECs) tube formation, migration, and proliferation, and reduced MCECs apoptosis under high-glucose conditions. These effects were significantly attenuated in miR-148a-3p-depleted MCPs-EVs, which were extracted after inhibiting miR-148a-3p expression in MCPs. Repetitive intracavernous injections of MCPs-EVs improved erectile function by inducing cavernous neurovascular regeneration in diabetic mice. Using online bioinformatics databases and luciferase report assays, we predicted that pyruvate dehydrogenase kinase-4 (PDK4) is a potential target gene of miR-148a-3p. CONCLUSIONS: Our findings provide new and reliable evidence that miR-148a-3p in MCPs-EVs significantly enhances cavernous neurovascular regeneration by inhibiting PDK4 expression in diabetic mice.

microRNA-15b-5p shuttled by mesenchymal stem cell-derived extracellular vesicles protects podocytes from diabetic nephropathy via downregulation of VEGF/PDK4 axis.

Diabetic nephropathy (DN) is a severe complication of diabetes lethal for end-stage renal disease, with less treatment methodologies and uncertain pathogenesis. In the current study, we determined the role of mesenchymal stem cells (MSCs)-derived extracellular vesicles (EVs) containing microRNA (miR)-15b-5p in DN. After extraction and identification of MSC-derived EVs, mouse podocyte line MPC5 was selected to establish an in vitro high-glucose (HG) cell model, where expression of miR-15b-5p, pyruvate dehydrogenase kinase 4 (PDK4) and VEGFA expression in tissues and cells were determined. The loss- and gain- function assays were conducted to determine the roles of miR-15b-5p, PDK4 and VEGFA. MPC5 cells were then co-cultured with MSC-derived EVs and their biological behaviors were detected by Western blot, CCK-8 assay, and flow cytometry. The binding relationship between miR-15b-5p and PDK43 by dual luciferase reporter gene assay. The expression of miR-15b-5p was downregulated in podocytes under HG environment, but highly expressed in mouse MSCs-derived EVs. EVs-derived miR-15b-5p could protect MPC5 cell apoptosis and inflammation. miR-15b-5p inhibited the expression of PDK4 by directly bound to the 3'UTR region of PDK4 gene. miR-15b-5p inhibits VEGF expression by binding to PDK4. Inhibition of PDK4 decreased VEGFA expression and reduced apoptosis and inflammation. Collectively, miR-15b-5p shuttled by MSC-derived EV can play protective roles in HG-induced mouse podocyte injury, possibly by targeting PDK4 and decreasing the VEGFA expression.

LncRNA CASC2 Alleviates Sepsis-induced Acute Lung Injury by Regulating the miR-152-3p/PDK4 Axis.

BACKGROUND: Acute lung injury (ALI) is an early complication of sepsis and it is also considered as an important cause of high mortality in sepsis patients. This research aimed to explore the potential role and mechanism of long non-coding RNA (lncRNA) cancer susceptibility candidate 2 (CASC2) in sepsis-induced ALI. METHODS: The levels of CASC2, microRNA-152-3p (miR-152-3p) and pyruvate dehydrogenase kinase 4 (PDK4) in sepsis patients and LPS-treated HPAEpiC were detected by quantitative real-time PCR and western blot. Cell viability and apoptosis were assessed by Counting Kit-8 (CCK-8) assay and flow cytometry. The concentrations of inflammatory factors were tested by Enzyme-linked immunosorbent assay. Oxidative stress was evaluated by the levels of reactive oxygen species and superoxide dismutase using corresponding commercial kits. The targeting relationship between miR-152-3p and CASC2 or PDK4 was verified by dual-luciferase reporter, RNA immunoprecipitation (RIP) and RNA pull-down assays. RESULTS: CASC2 and PDK4 were down-regulated, while miR-152-3p was up-regulated in sepsis patients and LPS-stimulated HPAEpiC. Overexpression of CASC2 relieved the LPS-resulted cell viability inhibition, apoptosis promotion, inflammatory and oxidative damages in HPAEpiC. In addition, miR-152-3p was a miRNA target of CASC2 and CASC2 alleviated cell injury in LPS-disposed HPAEpiC by sponging miR-152-3p. Moreover, miR-152-3p directly targeted PDK4 and CASC2 increased the PDK4 expression by depending on the sponge effect on miR-152-3p. Meanwhile, inhibition of miR-152-3p attenuated LPS-triggered HPAEpiC injury by upregulating the level of PDK4. CONCLUSION: These results suggested that CASC2 ameliorated the LPS-induced injury in HPAEpiC via regulating miR-152-3p/PDK4 pathway.

Anti-Metastatic Effect of Pyruvate Dehydrogenase Kinase 4 Inhibition in Bladder Cancer via the ERK, SRC, and JNK Pathways.

Bladder cancer is a common global cancer with a high percentage of metastases and high mortality rate. Thus, it is necessary to identify new biomarkers that can be helpful in diagnosis. Pyruvate dehydrogenase kinase 4 (PDK4) belongs to the PDK family and plays an important role in glucose utilization in living organisms. In the present study, we evaluated the role of PDK4 in bladder cancer and its related protein changes. First, we observed elevated PDK4 expression in high-grade bladder cancers. To screen for changes in PDK4-related proteins in bladder cancer, we performed a comparative proteomic analysis using PDK4 knockdown cells. In bladder cancer cell lines, PDK4 silencing resulted in a lower rate of cell migration and invasion. In addition, a PDK4 knockdown xenograft model showed reduced bladder cancer growth in nude mice. Based on our results, PDK4 plays a critical role in the metastasis and growth of bladder cancer cells through changes in ERK, SRC, and JNK.

[Mining of Differentially Expressed Genes in Diabetic Cardiomyopathy Based on GEO Database].

Objective To screen out the key genes leading to diabetic cardiomyopathy by analyzing the mRNA array associated with diabetic cardiomyopathy in the GEO database. Methods The online tool GEO2R of GEO was used to mine the differentially expressed genes (DEG) in the datasets GSE4745 and GSE5606.R was used to draw the volcano map of the DEG,and the Venn diagram was established online to identify the common DEG shared by the two datasets.The clusterProfile package in R was used for gene ontology annotation and Kyoto encyclopedia of genes and genomes pathway enrichment of the DEG.GSEA was used for gene set enrichment analysis,and STRING for the construction of a protein-protein interaction network.The maximal clique centrality algorithm in the plug-in Cytohubba of Cytoscape was used to determine the top 10 key genes. The expression of key genes was studied in the primary cardiomyocytes of rats and compared between the normal control group and high glucose group. Results The expression of Pdk4,Ucp3,Hmgcs2,Asl6,and Slc2a4 was consistent with the array analysis results.The expression of Pdk4,Ucp3,and Hmgcs2 was up-regulated while that of Acsl6 and Slc2a4 was down-regulated in the cardiomyocytes stimulated by high glucose (25 mmol/L) for 72 h. Conclusion Pdk4,Ucp3,Hmgcs2,Asl6,and Slc2a4 may be associated with the occurrence and development of diabetic cardiomyopathy,and may serve as the potential biomarkers of diabetic cardiomyopathy.

Pyruvate dehydrogenase kinase 4-mediated metabolic reprogramming is involved in rituximab resistance in diffuse large B-cell lymphoma by affecting the expression of MS4A1/CD20.

Diffuse large B cell lymphoma (DLBCL) heterogeneity promotes recurrence and anti-CD20-based therapeutic resistance. Previous studies have shown that downregulation of MS4A1/CD20 expression after chemoimmunotherapy with rituximab leads to rituximab resistance. However, the mechanisms of CD20 loss remain unknown. We identified that pyruvate dehydrogenase kinase 4 (PDK4) is markedly elevated in DLBCL cells derived from both patients and cell lines with R-CHOP (rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone) resistance. We found that overexpression of PDK4 in DLBCL cells resulted in cell proliferation and resistance to rituximab in vitro and in vivo. Furthermore, loss of PDK4 expression or treatment with the PDK4 inhibitor dichloroacetate was able to significantly increase rituximab-induced cell apoptosis in DLBCL cells. Further studies suggested PDK4 mediates a metabolic shift, in that the main energy source was changed from oxidative phosphorylation to glycolysis, and the metabolic changes could play an important role in rituximab resistance. Importantly, by knocking down or overexpressing PDK4 in DLBCL cells, we showed that PDK4 has a negative regulation effect on MS4A1/CD20 expression. Collectively, this is the first study showing that targeting PDK4 has the potential to overcome rituximab resistance in DLBCL.

lncRNA GAS5 suppresses rheumatoid arthritis by inhibiting miR-361-5p and increasing PDK4.

Rheumatoid arthritis (RA) is an inflammatory disease that causes hyperplasia of synovial tissue and cartilage destruction. This research was to investigate the effects of lncRNA GAS5/miR-361-5p/PDK4 on rheumatoid arthritis. By qRT-PCR, GAS5 and PDK4 were found to be overexpressed in synovial tissue, fibroblast-like synoviocytes of RA patients and LPS-induced chondrocytes, while the miR-361-5p expression was significantly reduced. GAS5 overexpression resulted in a decrease in the proliferation and Bcl-2 protein expression, and an increase in the Bax protein level. On the contrary, miR-361-5p sponged by GAS5 could accelerate chondrocyte proliferation, inhibit apoptosis. PDK4 targeted by miR-361-5p could inhibit RA, and partially eliminated the effect of miR-361-5p on RA. Our study suggested that GAS5 suppressed RA by competitively adsorbing miR-361-5p to modulate PDK4 expression.

LINC00662 modulates cervical cancer cell proliferation, invasion, and apoptosis via sponging miR-103a-3p and upregulating PDK4.

Cervical cancer (CC) is one of the most common cancers among women with high recurrence rates all over the world. Recently, the molecular mechanism of CC has been gradually uncovered in accumulating reports. This study aimed to investigate the function and upstream regulation mechanism of pyruvate dehydrogenase kinase 4 (PDK4) in CC cells, which was verified as an oncogene in several cancers. Through RT-qPCR assay, we discovered that PDK4 was highly expressed in CC cells. Then, it was demonstrated in function assays that PDK4 facilitated CC cell proliferation and invasion, but inhibited CC cell apoptosis. Next, we sought to determine the upstream genes of PDK4, and miR-103a-3p was identified to target PDK4. Then, through bioinformatics tools and a range of mechanism assays, long intergenic non-protein coding RNA 662 (LINC00662) was verified as the sponge of miR-103a-3p. Moreover, LINC00662 positively modulated PDK4 expression via competitively binding to miR-103a-3p in CC cells. Subsequently, rescue assays demonstrated that LINC00662 accelerated CC cell proliferation and inhibited cell apoptosis through upregulating PDK4. Furthermore, forkhead box A1 (FOXA1) was verified to activate transcription of both LINC00662 and PDK4. Taken together, our study revealed a novel ceRNA pattern of LINC00662/miR-103a-3p/PDK4 with FOXA1 as a transcription factor of LINC00662 and PDK4 in CC cells.

STAT3-dependent analysis reveals PDK4 as independent predictor of recurrence in prostate cancer.

Prostate cancer (PCa) has a broad spectrum of clinical behavior; hence, biomarkers are urgently needed for risk stratification. Here, we aim to find potential biomarkers for risk stratification, by utilizing a gene co-expression network of transcriptomics data in addition to laser-microdissected proteomics from human and murine prostate FFPE samples. We show up-regulation of oxidative phosphorylation (OXPHOS) in PCa on the transcriptomic level and up-regulation of the TCA cycle/OXPHOS on the proteomic level, which is inversely correlated to STAT3 expression. We hereby identify gene expression of pyruvate dehydrogenase kinase 4 (PDK4), a key regulator of the TCA cycle, as a promising independent prognostic marker in PCa. PDK4 predicts disease recurrence independent of diagnostic risk factors such as grading, staging, and PSA level. Therefore, low PDK4 is a promising marker for PCa with dismal prognosis.

MiR-122-5p suppresses neuropathic pain development by targeting PDK4.

The complex pathogenesis and limited efficacy of available treatment make neuropathic pain difficult for long periods of time. Several findings suggested the regulatory role of microRNA in the development of neuropathic pain. This study aims to investigate the functional role of miR-122-5p in the development of neuropathic pain. Down-regulation of miR-122-5p was observed in spinal cords of rats with neuropathic pain. We also found that overexpressing miR-122-5p by intrathecal injection of miR-122-5p lentivirus in a mouse model of chronic sciatic nerve injury (CCI) prevented neuropathic pain behavior. In HEK-293 T cells, luciferase activity was significantly decreased in the transfection group with mimic-miR-122-5p in wild-type PDK4 reporter, compared with mutant PDK4 reporter. Increased PDK4 expression was also observed during the progression of neuropathic pain. Intrathecal injection of both mimic-miR-122-5p and shPDK4 in CCI mice downregulated PDK4 expression to a lower level when compared with injected with shPDK4. In CCI mice, transfection of shPDK4 suppressed mechanical allodynia and thermal hyperalgesia, while co-transfection of shPDK4 and LV-miR-122-5p resulted in stronger levels of mechanical allodynia and thermal hyperalgesia inhibition. Taken together, the data suggest that miR-122-5p inhibits PDK4 expression, attenuating neuropathic pain. This result suggests the potential role of miR-122-5p acting as a target for the treatment of neuropathic pain.

Long non-coding RNA LINC00243 promotes proliferation and glycolysis in non-small cell lung cancer cells by positively regulating PDK4 through sponging miR-507.

Non-small cell lung cancer (NSCLC) is the main subtype of lung cancer. The overall survival of NSCLC patients is relatively low even after various treatments. Accumulating evidence demonstrated that long non-coding RNA (LncRNA) plays crucial roles in different biological process. However, the role of a novel LncRNA, LINC00243, in NSCLC remains unclear. We aimed to explore the biological role of LINC00243 in NSCLC. The mRNA and protein levels were determined by real-time PCR and western blot, respectively. Cell viability in vitro was detected by Cell Counting Kit-8 (CCK-8) assay and colony-formation assay. Reporter assay was used to detect the interactions between molecules, and the interaction was assessed by RNA pull-down assay. LINC00243 expression increased in human NSCLC tissues and associated with poor prognosis of NSCLC patients. LINC00243 knockdown inhibited proliferation and glycolysis of NSCLC cells. Mechanically, LINC00243 acted as a molecular sponge for miR-507, and miR-507 reversed the effect of LINC00243 on promoting proliferation and glycolysis of NSCLC cells. Moreover, LINC00243 modulated expression of endogenous miR-507-targeted PDK4. LINC00243 promotes proliferation and glycolysis in NSCLC cells by positively regulating PDK4 through sponging miR-507. LINC00243 could be the potential target for NSCLC treatment clinically.

Down-regulation of lncRNA CASC9 aggravates sepsis-induced acute lung injury by regulating miR-195-5p/PDK4 axis.

BACKGROUND: Long non-coding RNA (lncRNA) cancer susceptibility candidate 9 (CASC9) is reported to be linked to cancers. This research aims to explore the role and possible mechanism of CASC9 in lung injury induced by sepsis. METHODS: Lipopolysaccharide (LPS) induced human small airway epithelial cells (HSAECs) were established in vitro to mimic sepsis-induced lung injury. The effects of CASC9 and miR-195-5p on HSAECs viability were studied by CCK-8 assay. Interactions between CASC9 and miR-195-5p were determined by bioinformatics analysis, RT-PCR, dual luciferase reporter assay, and RNA immunoprecipitation assay. Pyruvate dehydrogenase kinase 4 (PDK4) and apoptosis-related molecules including Bcl2 and Bad were detected by western blot. Additionally, sepsis-induced lung injury model in rats was established by intraperitoneal injection of LPS in vivo to validate the demonstrations of in vitro studies. RESULTS: CASC9 was markedly down-regulated while miR-195-5p was significantly up-regulated in HSAECs treated by LPS and lung tissues of rats with sepsis. CASC9 interacted with miR-195-5p, and negatively regulated its expression level. Overexpression of CASC9 or transfection of miR-195-5p inhibitors significantly promoted the viability of HSAECs. The transfection of miR-195-5p mimics effected oppositely. For mechanism, miR-195-5p targeted the 3'UTR of pyruvate dehydrogenase kinase 4 (PDK4) gene and depressed the protein level, and PDK4 was regulated indirectly by CASC9. Restoration of CASC9 in the lung tissues of rats with sepsis ameliorated lung injury. CONCLUSION: CASC9 protects lung epithelial cells from sepsis-induced injury via regulating miR-195-5p/PDK4 axis.