Expression Of Cdc42 Research Articles

Spatial dysregulation of Rho GTPases in Alzheimer’s Disease brain in human and in transgenic mouse model

AbstractBackgroundAlzheimer disease (AD) is histopathologically characterized by brain accumulation of amyloid plaques, neurofibrillary tangle formation, and loss of synapses. The small GTPases of Rho family such as RhoA, Rac1 and Cdc42 play an important role in neural plasticity by regulating synaptic actin and spine dynamics. However, the functions of these proteins in AD are paradoxical. To understand their contributions in AD pathogenesis, it is important to investigate the relationship between the expression of Rho GTPases and their functional states in signaling.MethodWe characterized the spatial dysregulation of Rho GTPases in human AD brains and that of aged Triple Transgenic (3xTg‐AD) mouse model carrying mutations in amyloid precursor protein, tau protein, and presenilin. For human brains, the paraffin‐embedded tissues were collected from age and gender matched AD and Non‐Dementia (ND) patients. Phosphorylated Rho GTPase antibodies were used as signaling indicators for immunofluorescence microscopyResultWe observed altered phospho‐RhoA (S188) and phospho‐Rac/Cdc42(S71) expression and distribution in human, wild type (WT), and 3xTg‐AD mouse brains. In the CA3 region and dentate gyrus (DG), pRac/Cdc42 immunostaining showed decreased punctate staining in WT compared to 3xTg‐AD mice. For CA3 pRhoA immunostaining scored a slightly higher level of expression in WT compared to 3xTg‐AD. In the CA2 region, pRac/Cdc42 immunostaining was perinuclear in WT and diffused in 3xTg‐AD, whereas the pRhoA immunostaining pattern was similar between WT and 3xTg‐AD. In the CA1 region, pRac/Cdc42 immunostaining showed perinuclear as well as dendritic localization in WT and 3xTg‐AD, while pRhoA immunostaining between WT and 3xTg‐AD was similar. In human brains,there was no significant difference in pRhoA in cortex between ND and AD whereas pRac/Cdc42 was slightly reduced in AD cortex. However, while Rac signaling was reduced, Cdc42 signaling was increased in ND compared to AD cortex. On the other hand, Rac signaling is decreased in ND DG compared to AD whereas the differences in Cdc42 signaling in DG were not significant.ConclusionSmall GTPases of Rho family are differentially dysregulated in expression, distribution, and functional states in different areas of the brain in AD patients as well as in AD mouse brains

TMIC-34. INVESTIGATING THE EFFECT OF IRON IN GLIOBLASTOMA CELL POLARIZATION AND MIGRATION

Abstract Glioblastoma represents one of the most difficult-to-treat malignancies as evidenced by the poor prognosis associated with a diagnosis. The ability of glioblastoma cells to diffusely infiltrate into healthy brain tissue renders complete surgical resection challenging. Consequently, a large majority of glioblastoma patients end up with recurrent disease despite receiving maximally feasible surgical resection and rigorous chemoradiation. This work examined how modulation of cellular iron levels in T98G and LN229 glioblastoma cells impacted migratory capacity. Treatment of T98G or LN229 glioblastoma cells with iron in the form of ferric ammonium citrate (FAC) resulted in significantly reduced migration as assessed by time-lapse phase contrast imaging and wound healing assays. The iron-induced reduction in migration was able to be rescued by the addition of equimolar concentrations of deferoxamine, an iron chelator. Cellular proliferation in response to the iron treatments was quantified using both optical confluence and nucleic-acid-based proliferation assays and it was found that iron treatment at the concentrations used for the migration assays (0 – 300 µM FAC) did not result in reduced proliferation. Mechanistically probing iron’s impact on cell migration revealed that addition of iron resulted in decreased expression of Cdc42, a Rho GTPase that is essential to determining cellular polarity during migration. Functional cellular polarization assays further confirmed that reduced expression of Cdc42 corresponded to reduced cellular polarization. Bioinformatic analysis of CDC42 transcripts revealed the presence of potential iron-responsive-elements that may drive the iron-induced reduction in Cdc42 expression. This work highlights the importance of iron biology in impacting glioblastoma cell phenotype and potentially glioblastoma patient outcomes.

EPCO-36. INVESTIGATING THE BAIAP2-CDC42 INTERACTION IN MEDULLOBLASTOMA

Abstract Medulloblastoma is the most common malignant brain tumor found in children. The standard of care is surgical removal, radiation, and chemotherapy. Despite advancements in treatment, there is still no significant improvement of survival rates using chemotherapy compared to no treatment. This indicates novel, more effective treatment options are necessary. In addition, there is a worst mortality rate when medulloblastoma is metastasized outside the central nervous system. Recent studies have identified that medulloblastoma tumors can disseminate via the bloodstream and cerebral spinal fluid. However, not much is known about the properties of metastasis and invasion in medulloblastoma. We set out to identify potential key regulators for the pathophysiology of medulloblastoma. I mined and analyzed Medulloblastoma patient tumor data along with data obtained from a 10X Genomics Chromium single-cell RNA sequencing experiment performed in the laboratory from a Tg (Neurod-Smoothened*A1) mouse to investigate cellular heterogeneity and identify potential regulators in the metastasis and invasion of medulloblastoma. We identified BAIAP2 and CDC42 to be differentially expressed in our data. Previous studies show that these genes are implicated in oncogenic properties, such as motility, filopodia, and invasion. We hypothesize that distinct cell populations within medulloblastoma should show different expressions of BAIAP2 and CDC42 and that there will be different expressions when compared to non-tumor brain cells. We determined that BAIAP2 and CDC42 have higher expression in medulloblastoma tumors compared to the control. Interestingly, BAIAP2 is enriched in the cell population for microtubule depolymerization. Moreover, we show the role BAIAP2 and CDC42 have in the invasion and metastasis of medulloblastoma. This study is important because BAIAP2 and CDC42 have yet to be characterized in medulloblastoma. We suggest that BAIAP2 and CDC42 have a role in the metastasis and invasion of medulloblastoma indicating it could be a possible target for future studies.

Electroacupuncture Enhances Cognitive Deficits in a Rat Model of Rapid Eye Movement Sleep Deprivation via Targeting MiR-132.

Deprivation of rapid eye movement sleep (REMSD) reduces the potential for learning and memory. The neuronal foundation of cognitive performance is synapse plasticity. MicroRNA-132 (MiR-132) is an important microRNA related to cognitive and synapse plasticity. Acupuncture is effective at improving cognitive impairment caused by sleep deprivation. Furthermore, its underlying principle is still unclear. Herein, whether electroacupuncture (EA) helps alleviate cognitive impairment in REMSD by targeting miR-132 was assessed. A rat model of REMSD was constructed using the developing multiplatform water environment technique, as well as EA therapy in Baihui (GV20) and Dazhui (GV14) was performed for 15 minutes, once daily for 7 days. Agomir or antagomir of MiR-132 was injected into the hippocampal CA1 areas to assess the EA mechanism in rats with REMSD. Then, the learning and memory abilities were detected by behavioral tests; synapse structure was assessed by transmission electron microscope (TCM); and dendrites branches and length were examined by Golgi staining. MiR-132-3p was assessed by real-time quantitative polymerase chain reaction (qRT-PCR). P250GAP, ras-related C3 botulinum toxin substrate 1 (Rac1), and cell division cycle 42 (Cdc42) expression levels in hippocampal tissues were evaluated by immunohistochemistry and Western blot. According to the research, EA therapy enhanced cognitive in REMSD rats, as evidenced by reduced escape latency; upregulated the performance of platform crossings and prolonged duration in the goal region; and improved spontaneous alternation. EA administration restored synaptic and dendritic structural damage in hippocampal neurons, enhanced miR-132 expression, and reduced p250GAP mRNA and protein levels. Additionally, EA boosted the protein level of Rac1 and Cdc42 associated with synaptic plasticity. MiR-132 agomir enhanced this effect, whereas miR-13 antagomir reversed this action. The current data demonstrate that EA at GV20 and GV14 attenuates cognitive impairment and modulates synaptic plasticity in hippocampal neurons via miR-132 in a sleep-deprived rat model.

Mucin 1 aggravates synovitis and joint damage of rheumatoid arthritis by regulating inflammation and aggression of fibroblast-like synoviocytes.

AimsTo explore the synovial expression of mucin 1 (MUC1) and its role in rheumatoid arthritis (RA), as well as the possible downstream mechanisms.MethodsPatients with qualified synovium samples were recruited from a RA cohort. Synovium from patients diagnosed as non-inflammatory orthopaedic arthropathies was obtained as control. The expression and localization of MUC1 in synovium and fibroblast-like synoviocytes were assessed by immunohistochemistry and immunofluorescence. Small interfering RNA and MUC1 inhibitor GO-203 were adopted for inhibition of MUC1. Lysophosphatidic acid (LPA) was used as an activator of Rho-associated pathway. Expression of inflammatory cytokines, cell migration, and invasion were evaluated using quantitative real-time polymerase chain reaction (PCR) and Transwell chamber assay.ResultsA total of 63 RA patients and ten controls were included. Expression of MUC1 was observed in both the synovial lining and sublining layer. The percentage of MUC1+ cells in the lining layer of synovium was significantly higher in RA than that in control, and positively correlated to joint destruction scores of RA. Meanwhile, MUC1+ cells in the sublining layer were positively correlated to the Krenn subscore of inflammatory infiltration. Knockdown of MUC1, rather than GO-203 treatment, ameliorated the expression of proinflammatory cytokines, cell migration, and invasion of rheumatoid synoviocytes. Knockdown of MUC1 decreased expression of RhoA, Cdc42, and Rac1. Treatment with LPA compromised the inhibition of migration and invasion, but not inflammation, of synoviocytes by MUC1 knockdown.ConclusionUpregulated MUC1 promotes the aggression of rheumatoid synoviocytes via Rho guanosine triphosphatases (GTPases), thereby facilitating synovitis and joint destruction during the pathological process of RA.Cite this article: Bone Joint Res 2022;11(9):639–651.

Hypoxia-Induced CD36 Expression in Gastric Cancer Cells Promotes Peritoneal Metastasis via Fatty Acid Uptake

BackgroundThe lipid scavenger receptor cluster of differentiation 36 (CD36) has been shown to have a pro-metastatic function in several cancers. Adipose tissue, a favorable site for peritoneal metastasis (PM) from gastric cancer (GC), promotes this process by providing free fatty acids (FFAs); however, the role of CD36 in PM progression from GC remains to be elucidated.Materials and MethodsWe evaluated CD36 expression in the GC cells under various conditions. CD36 overexpressing (CD36OE) MKN45 cells were prepared and their migration and invasive properties were assessed. A PM mouse model was used to investigate the biological effects of palmitic acid (PA) and CD36. Furthermore, we examined the clinical role of CD36 expression in 82 human PM samples by immunohistochemical staining.ResultsHypoxia markedly increased CD36 expression in GC cells. In normoxia, only CD36OE MKN45 cells treated with PA showed an increase in migration and invasion abilities. An increased expression of active Rac1 and Cdc42 was observed, which decreased following etomoxir treatment. Conversely, hypoxia increased those capacities of both vector and CD36OE MKN45 cells. In a mouse model transplanted with CD36OE MKN45 cells, more peritoneal tumors were observed in the high-fat diet group than those in the normal diet group. In clinical samples, 80% of PM lesions expressed CD36, consistent with hypoxic regions, indicating a significant association with prognosis.ConclusionOur findings indicate that a hypoxia in the peritoneal cavity induces CD36 expression in GC cells, which contributes to PM through the uptake of FFAs.

Hypermethylation of PRKCZ Regulated by E6 Inhibits Invasion and EMT via Cdc42 in HPV-Related Head and Neck Squamous Cell Carcinoma

Simple SummaryHPV+ and HPV- HNSCC share distinct epigenetic characteristics and clinicopathological features. The aim of our study was to assess whether DNA methylation plays a role in the progression of HPV+ HNSCC. We used a HumanMethylation450 BeadChip array (Illumina) to identify PRKCZ genes exhibiting different levels of DNA methylation between HPV+ and HPV- HNSCC. PRKCZ acts as a potent tumor promoter in HPV+ HNSCC. These findings may provide a possible explanation for the differences in the clinicopathological characteristics between HPV+ and HPV- HNSCC and promising ideas for the treatment of HPV+ HNSCC.Purpose: To study the role of target genes with aberrant DNA methylation in HPV+ HNSCC. Methods: A HumanMethylation450 BeadChip array (Illumina) was used to identify differentially methylated genes. CCK-8, flow cytometry, wound healing, and cell invasion assays were conducted to analyze the biological roles of PRKCZ. Western blot, qRT-PCR, immunohistochemistry, and animal studies were performed to explore the mechanisms underlying the functions of PRKCZ. Results: We selected PRKCZ, which is associated with HPV infection, as our target gene. PRKCZ was hypermethylated in HPV+ HNSCC patients, and PRKCZ methylation status was negatively related to the pathological grading of HNSCC patients. Silencing PRKCZ inhibited the malignant capacity of HPV+ HNSCC cells. Mechanistically, HPV might promote DNMT1 expression via E6 to increase PRKCZ methylation. Cdc42 was required for the PRKCZ-mediated mechanism of action, contributing to the occurrence of epithelial-mesenchymal transition (EMT) in HPV+ HNSCC cells. In addition, blocking PRKCZ delayed tumor growth in HPV16-E6/E7 transgenic mice. Cdc42 expression was decreased, whereas E-cadherin levels increased. Conclusion: We suggest that PRKCZ hypermethylation induces EMT via Cdc42 to act as a potent tumor promoter in HPV+ HNSCC.

Cdc42 upregulation under high glucose induces podocyte apoptosis and impairs β-cell insulin secretion.

ObjectivesThe progressive impairment of β-cell function results in prolonged deterioration in patients with type 2 diabetes mellitus (T2DM). Interestingly, the finding on pancreatitis secondary to renal injury suggests that potential communication exists between kidney and pancreas. Therefore, we aimed to investigate cell division cycle 42 (Cdc42)-mediated podocyte apoptosis and its effect on insulin secretion in islet β-cells.MethodsType 2 diabetic nephropathy mouse models were established to identify the expression of Cdc42 in podocytes by immunohistochemistry. An in vitro co-culture of mouse podocyte MPC5 and β-TC6 cells was preliminarily established. Subsequently, podocyte apoptosis induced by high glucose and Cdc42 was detected by TUNEL staining and western blotting. In addition, the JNK pathway was examined to determine the mechanism of apoptosis in MPC5 cells. Finally, insulin secretion and expression in β-TC6 cells as well as malondialdehyde (MDA) and superoxide dismutase (SOD) levels in both cell types were examined after the regulation of Cdc42 in MPC5 cells.ResultsCdc42 was highly expressed in the podocytes of diabetic nephropathy mice. Exposure to 25 mM glucose for 48 h induced a significant upregulation of Cdc42, Bax, and cleaved caspase-3 as well as a decreased Bcl-2 expression. In addition, marked apoptosis of MPC5 cells was observed compared to normal glucose treatment. After transfection with Cdc42 plasmid, apoptosis of MPC5 cells was enhanced with an increased expression of p-JNK, whereas inhibition of Cdc42 significantly alleviated podocyte apoptosis accompanied by a downregulation of p-JNK. The glucose-stimulated insulin secretion level of β-TC6 cells decreased after the upregulation of Cdc42 in MPC5 cells. Immunofluorescence staining for insulin showed that co-culture with MPC5 cells carrying the Cdc42 plasmid significantly reduced insulin expression, whereas inhibition of Cdc42 in MPC5 cells alleviated the above-mentioned abnormality of β-TC6 cells. The expression of Cdc42 and p-p38 in β-TC6 cells increased following the upregulation of Cdc42 in MPC5 cells; this was concurrent with augmented MDA levels and decreased SOD activity. The opposite result was observed for Cdc42 knockdown in MPC5 cells.ConclusionsCdc42 in podocytes plays a crucial role in insulin secretion by β-cells, which may provide a new therapeutic target to prevent the vicious cycle of β-cell dysfunction in T2DM.

The Effects of Poria cocos on Rho Signaling-Induced Regulation of Mobility and F-Actin Aggregation in MK-801-Treated B35 and C6 Cells

Methods B35 neuronal cells and C6 glial cells were incubated with MK-801 for 7 days followed by MK-801, MK801 in combination with water extracts of P. cocos (PRP for P. cocos cum Radix Pini or WP for White Poria) treatment for an additional 7 days. Analysis of cell mobility, F-actin aggregation, and Rho signaling modulation was performed to clarify the roles of PRP or WP in MK-801-treated B35 and C6 cells. Results MK-801 decreases B35 cell mobility, whereas the inhibited cell migration ability and F-actin aggregation in MK-801-treated B35 or C6 cells could be reversed by PRP or WP. The CDC42 expression in B35 or C6 cells would be reduced by MK-801 and restored by treating with PRP or WP. The RhoA expression was increased by MK-801 in both B35 and C6 cells but was differentially regulated by PRP or WP. In B35 cells, downregulation of PFN1, N-WASP, PAK1, and ARP2/3 induced by MK-801 can be reversely modulated by PRP or WP. PRP or WP reduced the increase in the p-MLC2 expression in B35 cells treated with MK-801. The reduction in ROCK1, PFN1, p-MLC2, and ARP2/3 expression in C6 cells induced by MK-801 was restored by PRP or WP. Reduced N-WASP and PAK1 expression was differentially regulated by PRP or WP in MK-801-treated C6 cells.

Downregulation of CDC42 inhibits the proliferation and stemness of human trophoblast stem cell via EZRIN/YAP inactivation.

Placental dysplasia increases the risk of recurrent spontaneous abortion (RSA). However, the underlying mechanism regulating placental development remains unclear. In this study, we showed that the expression of CDC42 was decreased in the villous tissue of RSA samples compared to healthy controls. Further examination demonstrated that CDC42 deficiency led to the differentiation of human trophoblast stem cells (hTSCs) and inhibited their proliferation. Genetic manipulation of YAP and EZRIN in hTSCs revealed that CDC42 regulates the stemness and proliferation of hTSCs; this is dependent on EZRIN, which translocates YAP into the nucleus. Moreover, the expression pattern of EZRIN, YAP, and Ki67 was also abnormal in the villous tissue of RSA samples, consistent with in vitro experiments. In summary, these findings suggest that the CDC42/EZRIN/YAP pathway plays an important role in placental development.

β2-adrenergic receptor drives the metastasis and invasion of pancreatic ductal adenocarcinoma through activating Cdc42 signaling pathway

Recent investigations indicate that β2-adrenergic receptor (β2-AR) signaling may facilitate the progression of various tumors, whose underlying mechanisms remain largely elusive. In the present study, we showed that β2-AR recruited Cdc42 in response to isoproterenol (ISO, a β-AR selective agonist) exposure in pancreatic ductal adenocarcinoma (PDAC) cells. The association of β2-AR and Cdc42 promoted the activation of Cdc42, as revealed by increased levels of Cdc42-GTP, and co-incubation with β2-AR antagonist abrogated ISO-induced activation of Cdc42. β2-AR-mediated Cdc42 activation further led to the phosphorylation of downstream PAK1, LIMK1 and Merlin. Furthermore, we showed that the activation of β2-AR/Cdc42 signaling facilitated the migration and invasion of PDAC cells. In addition, β2-AR and Cdc42 were overexpressed in PDAC specimens, compared with adjacent non-tumor tissues. High expression of β2-AR and Cdc42 were correlated with lymph node metastasis and TNM stage in PDAC patients. Finally, we showed that overexpression of β2-AR and Cdc42 were indicative of unfavorable prognosis in PDAC patients. Taken together, our findings suggested that β2-AR might facilitate Cdc42 signaling to drive the migration and invasion of PDAC cells, consequently resulting in the metastasis and dismal prognosis of PDAC. These studies highlight targeting β2-AR/Cdc42 signaling as a therapeutic strategy against PDAC.

Abstract 2430: The role of cellular iron and the homeostatic iron regulator (HFE) in high-grade brain tumor cell migration

Abstract High-grade brain tumors such as grade III astrocytoma and glioblastoma represent among the most difficult to manage malignancies facing oncological practice. Diffuse migration and invasion into adjacent healthy brain tissue yields complete surgical resection of these tumors unfeasible. As a result, despite receiving maximal surgical resection and an aggressive course of chemoradiation, the majority of high-grade brain tumor patients suffer from recurrent disease. Increased expression levels of the homeostatic iron regulator gene (HFE) in high-grade brain tumors have been correlated with poorer outcomes. HFE is known to influence cellular iron metabolism by inhibiting transferrin-mediated iron uptake yet little is known regarding how HFE or iron impact the migratory capabilities of high-grade brain tumor cells. In order to better understand how HFE expression and cellular iron metabolism influence cell migration in high grade brain tumors, we utilized brain tumor cell lines that had been genetically manipulated to express different levels of HFE. We observed that knocking down HFE in KR158 or LN229 glioma cell lines resulted in significantly decreased migratory capacity. Since HFE is known to inhibit transferrin mediated iron uptake, we studied how directly modulating the iron status of glioma cells impacted their ability to migrate. Treatment of a panel of glioma cell lines: LN229, T98G, U87, KR158, with iron in the form of hemin or ferric ammonium citrate resulted in significantly reduced migration. Furthermore, the iron-induced reduction in migration could be rescued by the addition of deferoxamine, an iron chelator. Cell viability in response to the iron treatments was assayed and found to not be significantly altered - suggesting that cellular iron status was influencing migratory capacity independent of cell viability. To gain mechanistic insights into HFE and iron-induced effects on cell migration, we analyzed the Chinese Glioma Genome Atlas (CGGA) for correlations between HFE and the Rho GTPases RHOA, RAC1, and CDC42 – genes which are known to play a crucial role in determining the migratory capacity of cancer cells. Interestingly, we found statistically significant correlations between the Rho GTPases RHOA, RAC1, CDC42 and HFE in both grade III astrocytoma and glioblastoma patient cohorts. Immunoblotting of iron treated glioma cell lines demonstrated that expression of RhoA and Cdc42 was reduced suggesting that alterations in Rho GTPase expression and signaling may play a role in iron-induced effects on cell migration. Our results demonstrate that targeting cancer cell iron metabolism as an addition to existing treatment regimens may be a promising avenue for further investigation. Citation Format: Ganesh Shenoy, Katie Troike, Kondaiah Palsa, Madison Kuhn, Quinn Wade, Becky Slagle-Webb, Amanda Snyder, Chachrit Khunsriraksakul, Justin D. Lathia, Dhimant Desai, Hong-Gang Wang, Elizabeth Proctor, James R. Connor. The role of cellular iron and the homeostatic iron regulator (HFE) in high-grade brain tumor cell migration [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2430.

Rho-GTPase dependent leukocyte interaction generates pro-inflammatory thymic Tregs and causes arthritis

Conditional mutation of protein geranylgeranyltransferase type I (GGTase-I) in macrophages (GLC) activates Rho-GTPases and causes arthritis in mice. Knocking out Rag1 in GLC mice alleviates arthritis which indicates that lymphocytes are required for arthritis development in those mice. To study GLC dependent changes in the adaptive immunity, we isolated CD4+ T cells from GLC mice (CD4+GLCs). Spleen and joint draining lymph nodes (dLN) CD4+GLCs exhibited high expression of Cdc42 and Rac1, which repressed the caudal HOXA proteins and activated the mechanosensory complex to facilitate migration. These CDC42/RAC1 rich CD4+GLCs presented a complete signature of GARP+NRP1+IKZF2+FOXP3+ regulatory T cells (Tregs) of thymic origin. Activation of the β-catenin/Lef1 axis promoted a pro-inflammatory Th1 phenotype of Tregs, which was strongly associated with arthritis severity. Knockout of Cdc42 in macrophages of GLC mice affected CD4+ cell biology and triggered development of non-thymic Tregs. Knockout of Rac1 and RhoA had no such effects on CD4+ cells although it alleviated arthritis in GLC mice. Disrupting macrophage and T cell interaction with CTLA4 fusion protein reduced the Th1-driven inflammation and enrichment of thymic Tregs into dLNs. Antigen challenge reinforced the CD4+GLC phenotype in non-arthritic heterozygote GLC mice and increased accumulation of Rho-GTPase expressing thymic Tregs in dLNs. Our study demonstrates an unexpected role of macrophages in stimulating the development of pro-inflammatory thymic Tregs and reveal activation of Rho-GTPases behind their arthritogenic phenotype.

A novel lncRNA-miRNA-mRNA triple network identifies lncRNA XIST as a biomarker for acute myocardial infarction.

Despite the well-established role of long non-coding RNAs (lncRNAs) across various biological processes, their mechanisms in acute myocardial infarction (AMI) are not fully elucidated. The GSE34198 dataset from the Gene Expression Omnibus (GEO) database, which comprised 49 specimens from individuals with AMI and 47 specimens from controls, was extracted and analysed using the weighted gene co-expression network analysis (WGCNA) package. Twenty-seven key genes were identified through a combination of the degree and gene significance (GS) values, and the CDC42 (degree = 64), JAK2 (degree = 41), and CHUK (degree = 30) genes were identified as having the top three-degree values among the 27 genes. Potential interactions between lncRNA, miRNAs and mRNAs were predicted using the starBase V3.0 database, and a lncRNA-miRNA-mRNA triple network containing the lncRNA XIST, twenty-one miRNAs and three hub genes (CDC42, JAK2 and CHUK) was identified. RT–qPCR validation showed that the expression of the JAK2 and CDC42 genes and the lncRNA XIST was noticeably increased in samples from patients with AMI compared to normal samples. Pearson’s correlation analysis also proved that JAK2 and CDC42 expression levels correlated positively with lncRNA XIST expression levels. The area under ROC curve (AUC) of lncRNA XIST was 0.886, and the diagnostic efficacy of the lncRNA XIST was significantly better than that of JAK2 and CDC42. The results suggested that the lncRNA XIST appears to be a risk factor for AMI likely through its ability to regulate JAK2 and CDC42 gene expressions, and it is expected to be a novel and reliable biomarker for the diagnosis of AMI.

TIP60 acts as a regulator of genes involved in filopodia formation and cell migration during wound healing

Wound healing is a complex phenomenon that requires coordination of numerous molecular and cellular changes to facilitate timely and efficient repair of the damaged tissue. Although many of these molecular pathways have been detailed, others remain to be elucidated. In the present work, we show for the first time, roles for the acetyltransferase TIP60 and nuclear receptor transcription factor PXR in this process, participating in wound healing by altering actin dynamics and cellular motility. We found that in response to wound-injury, TIP60 induces rapid formation of filopodia at the wounded cell front, leading to enhanced cell migration and faster closure of the wound. Further, qPCR analysis revealed heightened expression of Cdc42 and ROCK1 genes, key regulators involved in filopodia formation and actin reorganization, exclusively in TIP60-PXR-expressing cells upon wound-induction. We also performed ChIP assays to confirm the context-specific binding of TIP60 on the ROCK1 promoter and demonstrated that the TIP60 chromodomain is essential for loading of the TIP60–PXR complex onto the chromatin. Results from immunoprecipitation assays revealed that during the wounded condition, TIP60 alters the chromatin microenvironment by specifically acetylating histones H2B and H4, thereby modulating the expression of target genes. Overall, findings of this study show that TIP60 is a novel regulator of the wound healing process by regulating the expression of wound repair-related genes.

Bioinformatics Study Revealed Significance of Exosome Transcriptome in Hepatocellular Carcinoma Diagnosis.

Background: Hepatocellular carcinoma (HCC) is one of the fifty most common cancers globally, having a high mortality rate being the second most common cause of cancer-related deaths. However, little attention has been paid to the involvement of exosomes and ceRNA in HCC. Method: The study aimed to explore exosome data from exoRBase database and a free online database to estimate possible binding miRNA from mRNA, lncRNA, and circRNA and discover useful exosome biomarkers for HCC therapy. Results: The results indicated that a total of 159 mRNAs, 60 lncRNAs, and 13 circRNAs were differentially expressed, with HIST2H3C exhibiting the highest log2FC change, CTD-2031P19 exhibiting the most relevant lncRNA, and CTD-2031P19 exhibiting the most relevant lncRNA. MARCH8, SH3PXD2A, has-circ-0014088, hsa-miR-186-5p, and hsa-miR-613 were identified as hub biomarkers used by Cytoscape. According to the KEGG pathway analysis results, the differentially expressed proteins were primarily enriched in the MAPK signaling network, central carbon metabolism in cancer, the glucagon signaling pathway, glutamatergic synapse, and spliceosome. Furthermore, immunohistochemical images from the Human Protein Atlas (HPA) online tool were used to directly evaluate the protein expression of SMARCA5, CDC42, and UBC between normal and cancer tissues, and the results showed that these three gene expressions were significantly higher in tumor tissues. Conclusion: This study discovered atypical signature exosomes for HCC prognostic prediction based on an online database. The signals could mimic exosome microenvironmental disorders providing potential biomarkers for exosome treatment.

Longitudinal Variations of CDC42 in Patients With Acute Ischemic Stroke During 3-Year Period: Correlation With CD4+ T Cells, Disease Severity, and Prognosis.

ObjectiveCell division cycle 42 (CDC42) modulates CD4+ T-cell differentiation, blood lipids, and neuronal apoptosis and is involved in the pathogenesis of acute ischemic stroke (AIS); however, the clinical role of CDC42 in AIS remains unanswered. This study aimed to evaluate the expression of CDC42 in a 3-year follow-up and its correlation with disease severity, T helper (Th)1/2/17 cells, and the prognosis in patients with AIS.MethodsBlood CDC42 was detected in 143 patients with AIS at multiple time points during the 3-year follow-up period and in 70 controls at admission by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). In addition, blood Th1, Th2, and Th17 cells and their secreted cytokines (interferon-γ (IFN-γ), interleukin-4 (IL-4), and interleukin-17A (IL-17A)) in patients with AIS were detected by flow cytometry and enzyme-linked immunosorbent assay (ELISA), respectively.ResultsCompared with controls (p < 0.001), CDC42 was reduced in patients with AIS. CDC42 was negatively correlated with the National Institutes of Health Stroke Scale (NIHSS) score (p < 0.001), whereas, in patients with AIS (all p < 0.050), it was positively associated with Th2 cells and IL-4 but negatively correlated with Th17 cells and IL-17A. CDC42 was decreased from admission to 3 days and gradually increased from 3 days to 3 years in patients with AIS (P<0.001). In a 3-year follow-up, 24 patients with AIS recurred and 8 patients died. On the 3rd day, 7th day, 1st month, 3rd month, 6th month, 1st year, 2nd year, and 3rd year, CDC42 was decreased in recurrent patients than that in non-recurrent patients (all p < 0.050). CDC42 at 7 days (p = 0.033) and 3 months (p = 0.023) was declined in reported deceased patients than in survived patients.ConclusionCDC42 is used as a biomarker to constantly monitor disease progression and recurrence risk of patients with AIS.

CDC42 Regulates Cell Proliferation and Apoptosis in Bladder Cancer via the IQGAP3-Mediated Ras/ERK Pathway.

Bladder cancer (BC) is the most common malignant tumour of the urinary system. The current conventional treatments for BC have certain limitations. It is very urgent and necessary to find new treatment strategies for BC. Our study elucidated the underlying regulatory mechanisms of cell division control protein 42 homologue (CDC42) to regulate the development of BC. Quantitative real-time polymerase chain reaction, Western blot, immunofluorescence and immunohistochemistry were used to assess the expression of CDC42 and IQ motif-containing GTPase-activating protein 3 (IQGAP3) in BC tissues and BC cells. We induced the knockdown or overexpression by transfecting sh-CDC42 or oe-IQGAP3 into BC cells. In addition, cell proliferation and apoptosis were evaluated by cell counting kit-8 and flow cytometry assays, respectively. Moreover, proteins involved in the rat sarcoma (Ras)/extracellular regulated protein kinase (ERK) pathway were determined by Western blot. The expression of CDC42 and IQGAP3 was markedly upregulated in both BC tissues and BC cells. CDC42 silencing downregulated the expression of IQGAP3 and suppressed the Ras/ERK pathway. In addition, CDC42 silencing markedly promoted apoptosis and inhibited proliferation in BC cells. Further experiments showed that overexpression of IQGAP3 dramatically abolished the bioeffects mediated by CDC42 silencing on the proliferation and apoptosis of BC cells. All our results suggested that CDC42 promoted the Ras/ERK pathway by regulating IQGAP3, thus enhancing cell proliferation and suppressing cell apoptosis in BC cells and ultimately participating in the pathogenesis of BC.

MiR-185-5p Regulates Inflammation and Phagocytosis through CDC42/JNK Pathway in Macrophages.

Macrophage activation is an essential component of systemic chronic inflammation and chronic inflammatory diseases. Emerging evidence implicates miR-185-5p in chronic inflammation diseases. However, the regulatory role of miR-185-5p in macrophage pro-inflammatory activation has not been studied previously. Here, we identified that miR-185-5p was one of the top genes and effectively downregulated in two macrophage miRNA expression datasets from GEO. Under LPS stress, miR-185-5p overexpression reduced pro-inflammatory cytokine expression, suppressed phagocytosis in RAW264.7 macrophage. miR-185-5p inhibitors augmented pro-inflammatory effects of LPS in macrophage. Mechanically, miR-185-5p sponged and negatively regulated the protein expression of CDC42. Ablation of CDC42 with selective CDC42 inhibitor CASIN reversed the pro-inflammatory effect of miR-185-5p inhibitors through inhibiting MAPK/JNK pathways. Collectively, these data demonstrate that miR-185-5p exhibited anti-inflammatory functions in LPS-induced RAW264.7 macrophages at least partially through CDC42/JNK pathways. Our findings yield insights into the understanding of miR-185-5p-regulated network in macrophages inflammation, which is beneficial for exploring miRNA-protein interaction in atherosclerotic inflammation.

EIF3a Regulates Colorectal Cancer Metastasis via Translational Activation of RhoA and Cdc42

Tumor metastasis is the major cause of tumor relapse and cancer-associated mortality in colorectal cancer, leading to poor therapeutic responses and reduced survival. eIF3a was previously described as an oncogene. However, its role in colorectal cancer progression and metastasis has not yet been fully investigated. In this study, the expression specificity and predictive value of eIF3a were investigated in clinical samples. The effects of eIF3a on cell proliferation and migration were verified in vivo and in vitro, respectively. The underlying molecular mechanism was revealed by western blotting, immunofluorescence, RNA-binding protein immunoprecipitation, and dual-luciferase reporter gene assays. The results showed that eIF3a was significantly overexpressed in tumor tissues compared with adjacent normal tissues. High eIF3a expression was correlated with tumor metastasis and overall survival. Downregulation of eIF3a obviously inhibited the proliferation and motility of malignant cells in vitro and in vivo. Mechanistically, eIF3a regulates Cdc42 and RhoA expression at the translation level, which further affects pseudopodia formation and actin cytoskeleton remodeling. Taken together, eIF3a accelerates the acquisition of the migratory phenotype of cancer cells by activating Cdc42 and RhoA expression at the translational level. Our study identified eIF3a as a promising target for inhibiting colorectal cancer metastasis.