Tanshinone ⅡA improves Alzheimer’s disease via RNA nuclearenriched abundant transcript 1/microRNA-291a-3p/member RAS oncogene family Rab22a axis

BACKGROUND Alzheimer’s disease(AD)is a neurodegenerative condition characterized by oxidative stress and neuroinflammation.Tanshinone ⅡA(Tan-ⅡA),a bioactive compound isolated from Salvia miltiorrhiza plants,has shown potential neuroprotective effects;however,the mechanisms underlying such a function remain unclear.AIM To investigate potential Tan-ⅡA neuroprotective effects in AD and to elucidate their underlying mechanisms.METHODS Hematoxylin and eosin staining was utilized to analyze structural brain tissue morphology.To assess changes in oxidative stress and neuroinflammation,we performed enzyme-linked immunosorbent assay and western blotting.Additionally,the effect of Tan-ⅡA on AD cell models was evaluated in vitro using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay.Genetic changes related to the long non-coding RNA(lncRNA)nuclear-enriched abundant transcript 1(NEAT1)/microRNA(miRNA,miR)-291a-3p/member RAS oncogene family Rab22a axis were assessed through reverse transcription quantitative polymerase chain reaction.RESULTS In vivo,Tan-ⅡA treatment improved neuronal morphology and attenuated oxidative stress and neuroinflammation in the brain tissue of AD mice.In vitro experiments showed that Tan-ⅡA dose-dependently ameliorated the amyloid-beta 1-42-induced reduction of neural stem cell viability,apoptosis,oxidative stress,and neuroinflammation.In this process,the lncRNA NEAT1-a potential therapeutic target-is highly expressed in AD mice and downregulated via Tan-ⅡA treatment.Mechanistically,NEAT1 promotes the transcription and translation of Rab22a via miR-291a-3p,which activates nuclear factor kappa-B(NF-κB)signaling,leading to activation of the pro-apoptotic B-cell lymphoma 2-associated X protein and inhibition of the anti-apoptotic B-cell lymphoma 2 protein,which exacerbates AD.Tan-ⅡA intervention effectively blocked this process by inhibiting the NEAT1/miR-291a-3p/Rab22a axis and NF-κB signaling.CONCLUSION This study demonstrates that Tan-ⅡA exerts neuroprotective effects in AD by modulating the NEAT1/miR-291a-3p/Rab22a/NF-κB signaling pathway,serving as a foundation for the development of innovative approaches for AD therapy.

Mutations in Ras homolog family member A in patients with peripheral T-cell lymphoma and implications for personalized medicine

Genome sequencing has revealed frequent mutations in Ras homolog family member A(RHOA)among various cancers with unique aberrant profiles and pathogenic effects,especially in peripheral T-cell lymphoma(PTCL).The discrete positional distribution and types of RHOA amino acid substitutions vary according to the tumor type,thereby leading to different functional and biological properties,which provide new insight into the molecular pathogenesis and potential targeted therapies for various tumors.However,the similarities and discrepancies in characteristics of RHOA mutations among various histologic subtypes of PTCL have not been fully elucidated.Herein we highlight the inconsistencies and complexities of the type and location of RHOA mutations and demonstrate the contribution of RHOA variants to the pathogenesis of PTCL by combining epigenetic abnormalities and activating multiple downstream pathways.The promising potential of targeting RHOA as a therapeutic modality is also outlined.This review provides new insight in the field of personalized medicine to improve the clinical outcomes for patients.

5-Aza-2'-deoxycytidine inhibits retinoblastoma cell by reactivating epigenetically silenced RASSF1A gene

AIM: To investigate the effect of 5-Aza-2’-deoxycytidine(5-Aza-CdR),a DNA methyltransferase(DNMT) inhibitor,on the growth and survival of the Chinese retinoblastoma(RB) cell line HXO-RB44. ·METHODS: The DNA methylation status of the Ras association domain family(RASSF1A) promoter in the presence of 5-Aza-CdR at different concentrations was analyzed by methylation-specific polymerase chain reaction(MSP). RASSF1A mRNA and protein levels were measured by semiquantitative RT-PCR and immunohistochemistry staining,respectively,when cells were treated with 5.0μmol/L of 5-Aza-CdR. The effect of 5.0μmol/L 5-Aza-CdR on the proliferation and viability of HXO-RB44 cells was examined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assay and flow cytometry. ·RESULTS: 5-Aza-CdR efficiently induced cell cycle arrest at G0 /G1 and apoptotic death in HXO-RB44 cells. MSP analysis showed that unmethylated RASSF1A DNA increased and methylated RASSF1A decreased in a dose-dependent manner in a range of 0.5-5.0μmol/L 5-Aza-CdR. Accordingly,RASSF1A expression was reactivated at both mRNA and protein levels. Incubation time of 5-Aza-CdR treatment also functioned as a factor for the demethylation status of RASSF1A promoter DNA,with a plateau on day four. 5-Aza-CdR at 5.0μmol/L completely demethylated the RASSF1A promoter in HXORB44 cells on day four,and as a result,RASSF1A expression increased significantly from day 4 to day 7.·CONCLUSION: 5-Aza-CdR inhibits the growth of the HXO-RB44 RB cell line and induces apoptosis by demethylating the RASSF1A gene.

Expression and clinicopathologic significance of RASSF1A and WT1 in recurrent epithelial ovarian cancer

Objective To explore the expression and clinicopathological significance of RASSF1A and WT1 in recurrent epithelial ovarian cancer.Methods Sixty-three cases of patients with pathologically confirmed epithelial ovarian cancer were collected in the department of Gynaecology and Obsterics of the North China University of Technology from January 2013 to December 2018.The expression of RASSF1A and WT1 were measured by IHC staining.The relation of these proteins with ovarian cancer was also analyzed.Results Compared with non-recurrent group(46.4%),the positive expression rate of RASSF1A was 17.1%in recurrent group.The positive expression rate of WT1 was 74.3%and higher than the rate of 42.8%in non-recurrent group.The reducing expression of RASSF1A was related to clinical stage,differentiation,and with ascites(P<0.05).The increasing expression of WT1 was related to pathological type,clinical stage,histological differentiation,with ascites,and lymph node metastasis(P<0.05).Conclusion Low expression of RASSF1A and high expression of WT1 may be related to recurrent epithelial ovarian cancer.

Axonal growth inhibitors and their receptors in spinal cord injury:from biology to clinical translation

Axonal growth inhibitors are released during traumatic injuries to the adult mammalian central nervous system, including after spinal cord injury. These molecules accumulate at the injury site and form a highly inhibitory environment for axonal regeneration. Among these inhibitory molecules, myelinassociated inhibitors, including neurite outgrowth inhibitor A, oligodendrocyte myelin glycoprotein, myelin-associated glycoprotein, chondroitin sulfate proteoglycans and repulsive guidance molecule A are of particular importance. Due to their inhibitory nature, they represent exciting molecular targets to study axonal inhibition and regeneration after central injuries. These molecules are mainly produced by neurons, oligodendrocytes, and astrocytes within the scar and in its immediate vicinity. They exert their effects by binding to specific receptors, localized in the membranes of neurons. Receptors for these inhibitory cues include Nogo receptor 1, leucine-rich repeat, and Ig domain containing 1 and p75 neurotrophin receptor/tumor necrosis factor receptor superfamily member 19(that form a receptor complex that binds all myelin-associated inhibitors), and also paired immunoglobulin-like receptor B. Chondroitin sulfate proteoglycans and repulsive guidance molecule A bind to Nogo receptor 1, Nogo receptor 3, receptor protein tyrosine phosphatase σ and leucocyte common antigen related phosphatase, and neogenin, respectively. Once activated, these receptors initiate downstream signaling pathways, the most common amongst them being the Rho A/ROCK signaling pathway. These signaling cascades result in actin depolymerization, neurite outgrowth inhibition, and failure to regenerate after spinal cord injury. Currently, there are no approved pharmacological treatments to overcome spinal cord injuries other than physical rehabilitation and management of the array of symptoms brought on by spinal cord injuries. However, several novel therapies aiming to modulate these inhibitory proteins and/or their receptors are under investigation in ongoing clinical trials. Investigation has also been demonstrating that combinatorial therapies of growth inhibitors with other therapies, such as growth factors or stem-cell therapies, produce stronger results and their potential application in the clinics opens new venues in spinal cord injury treatment.

Pannexin 1,a large-pore membrane channel,contributes to hypotonicity-induced ATP release in Schwann cells

Pannexin 1(Panx 1),as a large-pore membrane channel,is highly permeable to ATP and other signaling molecules.Previous studies have demonstrated the expression of Panx 1 in the nervous system,including astrocytes,microglia,and neurons.However,the distribution and function of Panx 1 in the peripheral nervous system are not clear.Blocking the function of Panx 1 pharmacologically(carbenoxolone and probenecid)or with small interfering RNA targeting pannexins can greatly reduce hypotonicity-induced ATP release.Treatment of Schwann cells with a Ras homolog family member(Rho)GTPase inhibitor and small interfering RNA targeting Rho or cytoskeleton disrupting agents,such as nocodazole or cytochalasin D,revealed that hypotonicity-induced ATP release depended on intracellular RhoA and the cytoskeleton.These findings suggest that Panx 1 participates in ATP release in Schwann cells by regulating RhoA and the cytoskeleton arrangement.This study was approved by the Animal Ethics Committee of Nantong University,China(No.S20180806-002)on August 5,2018.

Upregulation of miR-345-5p suppresses cell growth of lung adenocarcinoma by regulating ras homolog family member A(RhoA)and Rho/Rho associated protein kinase(Rho/ROCK)pathway

Background:Microribose nucleic acids(miRNAs)are implicated in the progression of lung adenocarcinoma.MicroRNA-345-5p(miR-345-5p)is a recently identified anti-oncogene in some human cancers,but its functional role and possible molecular mechanism in lung adenocarcinoma remain unknown.This study aimed to identify the biological function and underlying mechanism of miR-345-5p in lung adenocarcinoma cells.Methods:In this study,lung adenocarcinoma tissues and adjacent tissues were collected in the First Affiliated Hospital of Anhui Medical University between April 2016 and February 2017.The expression of miR-345-5p and ras homolog family member A(RhoA)in lung adenocarcinoma tissues and human lung adenocarcinoma cell lines(A549,H1650,PC-9,and H441)was detected by reverse transcription quantitative polymerase chain reaction analysis.Functional assays including colony formation,flow cytometry analysis,wound healing,and transwell assays were performed to assess the proliferation,apoptosis,migration,and invasion of lung adenocarcinoma cells.In addition,RNA pulldown and luciferase reporter assays were conducted to evaluate the relationship between miR-345-5p and RhoA.Difference between the two groups was analyzed with Student’st test,while that among multiple groups was analyzed with one-way analysis of variance.Results:MiR-345-5p expression displayed lower level in lung adenocarcinoma tissues(0.241±0.095vs.1.000±0.233,t=19.247,P<0.001)and cell lines(F=56.992,P<0.001)than control tissues and cells.Functional experiments demonstrated that upregulation of miR-345-5p inhibited the malignant phenotypes of lung adenocarcinoma cells via suppressing cell proliferation,migration,invasion,and facilitating cell apoptosis.Additionally,RhoA was verified to be the downstream target of miR-345-5p.Expression of RhoA was downregulated by overexpression of miR-345-5p in PC-9(0.321±0.047vs.1.000±0.127,t=8.536,P<0.001)and H1650(0.398±0.054vs.1.000±0.156,t=4.429,P=0.011)cells.Rescue assays revealed that overexpression of RhoA rescued the suppressive effects of miR-345-5p upregulation on proliferation,migration,and invasion of lung adenocarcinoma cells.Further,miR-345-5p was found to regulate the Rho/Rho-associated protein kinase(ROCK)signaling pathway by downregulation of RhoA in lung adenocarcinoma cells.Conclusions:MiR-345-5p plays a tumor suppressor role in lung adenocarcinoma cells by downregulating RhoA to inactivate the Rho/ROCK pathway.

RASSF1A基因的甲基化检测对宫颈癌和宫颈上皮内瘤变患者的临床意义

目的探讨RASSF1A基因甲基化程度对宫颈癌和宫颈上皮内瘤变(CIN)患者的临床意义。方法选取2017年5月至2019年2月大同市第五人民医院的46例宫颈癌组织,30例CIN及10例正常宫颈组织作为对照组。采用甲基化特异PCR技术来检测RASSF1A启动子区的甲基化程度;分析RASSFIA基因启动子区甲基化程度与临床及病理特征(即肿瘤组织学分级、FIGO分期、淋巴结转移、宫颈间质的浸润深度及HPV感染)的关系。结果本研究检测了46例宫颈癌、30例CIN及10例正常宫颈中RASSF1A的甲基化程度,结果显示其在宫颈癌中的甲基化率为39.1%(18/46),未甲基化率为60.9%(28/46),CIN及正常宫颈组织未见有明显甲基化。RASSF1A基因的甲基化程度与宫颈癌的FIGO分期及肿瘤的组织学分级相关,而与宫颈间质侵犯深度、淋巴结是否转移及是否有HPV感染无关。结论RASSF1A的甲基化程度可能与宫颈癌的发生相关。RASSF1A基因启动子区在宫颈癌组织中高甲基化程度,其与宫颈癌的FIGO分期、肿瘤的组织学分级有关,使其有望成为宫颈癌检测的敏感标志物,并可用于早期宫颈癌的诊断及临床治疗的预后分析。

Frequent 3p21 Allelic Loss and Methylation-Associated RASSF1A Inactivation in Non-Small Cell Lung Cancer and Its Clinical Implication

A total of 110 primary NSCLCs （non-small cell lung cancers） were recruited in this study to characterize the pattern of 3p21 LOH together with the RASSF1A methylation status and their clinical implication. 3p21 LOH by 8 microsatellite markers, RASSF1A methylation status by methylation-specific PCR （MSPCR） as well as bisulfite genomic sequencing （BGS）, and RASSF1A expression level by real-time quantitative PCR was performed. 3p21 LOH is frequent in NSCLC with a mean frequency of （41.2±3.7）%. Significant associations between 3p21 LOH and gender, smoking history, histological type, and tumor size were observed. Cases with LOH have a slightly lower RASSF1A expression than cases without LOH but not statistically significant. Comparison of RASSF1A methylation that resulted from the three analyses shows significant correlations from one another. Higher frequency of methylation was observed in larger tumors and in smokers compared with smaller tumors and non-smokers, respectively. A significant correlation was also observed in extent between methylation and RASSF1A expression, illustrating that epigenetic mechanism could affect gene expression. The significant clinicopathological relations of 3p21 LOH may be of great use for both early detection and therapeutic interventions.