Histone demethylation in cardiac hypertrophy is poorly understood. This study aims to determine the role of the histone demethylase LSD1 in pathological cardiac hypertrophy. Both isoprenaline (ISO)-treated and transverse aortic constriction (TAC)-treated rats developed hypertrophic hearts. LSD1 was significantly decreased; the histone marks mono- and dimethyl H3K4 and H3K9 (H3K4me1/2 and H3K9me1/2) were significantly up-regulated in the hypertrophic heart tissue, as well as the expression of the ANP, α-HMC and MLV-2v genes. An LSD1 inhibitor, OG-L002 could also induce cardiac hypertrophy and enhance the induction of cardiac hypertrophy by ISO. Overexpressed LSD1 abolished ISO-induced cardiac hypertrophy and downregulated H3K4me1/2 and H3K9me1/2 expression. Overexpression of LSD1 also reduced the expression of ANP, α-HMC and MLV-2v. In addition, we have reported isoprenaline (ISO) as one of the histone demethylase LSD1 inhibitors. This was confirmed by molecular docking, molecular dynamic studies and a histone demethylation assay. The H3K4me1/2 expression increases with the incubation of ISO in HEK 293T and HELA cells. CaMKII could be significantly activated by the LSD1 inhibitor OG-L002 as well as by ISO in rats. In summary, we have identified a novel role for LSD1 in initiating and maintaining cardiac hypertrophy.

Background and Aims: Diabetic kidney disease (DKD) is a prevalent and intractable microvascular complication of diabetes mellitus (DM), the process of which is closely related to abnormal expression of angiogenesis-regulating factors (ARFs). Stem cell transplantation might be a novel strategy for treating DKD. This study aims to explore the effect of transplantation of human amniotic mesenchymal stem cells (hAMSCs) on renal microangiopathy in a type 1 DKD rat model (T1DRM).Methods: Seventy-two rats were randomly divided into three groups, including normal control group, DKD group, and hAMSCs transplantation group. T1DRM was established using a rat tail vein injection of streptozotocin (STZ) (55 mg/kg). hAMSCs were obtained from placental amniotic membranes during cesarean delivery and transplanted at 3 and 4 weeks through penile veins. At 6, 8, and 12 weeks following transplantation, blood glucose levels, renal function, pathological kidney alterations, and the expressions of ARFs' mRNA and protein were analyzed.Results: In T1DRM, transplanted hAMSCs that were homed at the injured site of kidneys increased ARFs' expression and decreased blood glucose levels. Compared to the DKD group, the levels of 24-h urinary protein, serum creatinine, urea, and kidney injury molecule-1 (KIM-1) were reduced in hAMSCs transplantation group. In terms of renal pathology such as the degree of basement membrane thickening, hAMSCs transplantation was also less severe than the DKD group, thereby alleviating kidney injury.Conclusion: hAMSCs transplantation might ameliorate STZ-induced chronic kidney injury through increasing ARFs' expression in kidneys and lowering blood glucose levels.

Background. Lung adenosquamous carcinoma (LASC) is a special type of lung cancer. LASC is a malignant tumor with strong aggressiveness and a poor prognosis. Previous studies have revealed that microRNAs (miRNAs) are widely involved in the development of tumors by targeting mRNA. This study is aimed at identifying the key mRNAs and miRNAs of LASC and constructing miRNA-mRNA networks for deeply comprehending the latent molecular mechanisms. Methods. mRNA dataset (GSE51852) and miRNA dataset (GSE51853) were extracted and downloaded from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) and differentially expressed miRNAs (DEMs) were picked out by the GEO2R web tool. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment analyses were conducted in the DAVID database. The protein-protein interaction (PPI) network was performed and analyzed by using the STRING database and Cytoscape software, respectively. TransmiR v2.0 was applied to predict potential transcription factors of miRNAs. The target genes of DEMs were predicted in the miRWalk database. Results. In comparison to normal tissues, a total of 1458 DEGs (511 upregulated and 947 downregulated) and 13 DEMs (5 upregulated and 8 downregulated) were screened out in LASC tissues. The PPI network of the DEGs displayed five key modules and seventeen hub genes. Six target genes of the DEMs were predicted, and five essential miRNA-mRNA regulatory pairs were established. Ensuingly, CENPF, one of the target genes, was also the hub genes of GSE51852, which was obtained from MCODE and cytoHubba and regulated by hsa-miR-205. Conclusions. We constructed the miRNA-mRNA regulatory pairs, which are helpful to study the potential regulatory mechanisms and find out promising diagnosis biomarkers and therapeutic targets for LASC.

系统生物学已成为揭示中医脾藏象科学内涵的有力工具,其主要技术包括基因组学、转录组学、蛋白质组学、代谢组学4个方面.通过梳理目前中医脾藏象系统生物学研究文献,发现基因组学可以阐释中医脾藏象的生物学基础,转录组学可以揭示中医脾藏象的内在调控机制,蛋白质组学可以解析中医脾藏象的物质基础和内在联系,代谢组学可以解释中医脾藏象的代谢机制.系统生物学为构建脾藏象学术发展体系从而实现脾藏象的继承与创新提供了可能,但相关研究仍然存在一些问题需要解决,如对病和证的混合差异、对脾虚证本质研究缺乏特异性、忽略没有差异表达的潜在生物标志物的生物学信息、相关组学临床数据的分析相对匮乏、"基因-蛋白质-代谢物分离"等.

The thiophene [2,3-d]pyrimidine structure-like small molecules were discovered from structure-based virtual screening of 1 billion compounds. Base on enzyme activity assay results, a SHP2-specific molecule inhibitor Comp#2 with IC50 of 1.174 mu M, 85-fold more selective for SHP2 than the highly related SHP1 (IC50 > 100 mu M). The compound can effectively inhibit SHP2-mediated cell signaling and cancer cell proliferation, including cervix cancer, human pancreatic cancer, large cell lung cancer, and mouse glioma cell. Moreover, the in vivo assay indicated that Comp#2 could inhibit cervix cancer tumors growth in BABL/c mice. This work has shown the specific SHP2 inhibitor can inhibit glioblastoma growth in vivo.

Abstract In vivo, stem cells reside in a three-dimensional (3D) extracellular microenvironment in which complicated biophysical and biochemical factors regulate their behaviors. Biomimicking of the stem cell−matrix interactions is an ideal approach for controlling the stem cell fate. This study investigates the effects of the incorporation of cell-adhesive ligands in 3D self-assembling peptide hydrogels to modulate stem cell survival, proliferation, maintenance of stemness, and osteogenic differentiation. The results show that the composite hydrogels were non-cytotoxic and effective for maintaining human amniotic mesenchymal stem cell (hAMSC) survival, proliferation and phenotypic characterization. The expression levels of pluripotent markers were also upregulated in the composite hydrogels. Under inductive media conditions, mineral deposition and mRNA expression levels of osteogenic genes of hAMSCs were enhanced. The increasing expression of integrin α- and β-subunits for hAMSCs indicates that the ligand−integrin interactions may modulate the cell fate for hAMSCs in composite hydrogels.

目的 观察心肌肥厚大鼠心肌组织及肥大心肌细胞中环状RNA rno_circRNA_016002的表达变化,并探讨其在心肌肥厚过程中的作用.方法 将12只SD大鼠随机分为Control组与异丙肾上腺素(ISO)组,每组6只.ISO组每天腹腔注射ISO 3 mg/kg,Control组注射同等体积的生理盐水,连续注射两周后采用心脏超声检测心肌以明确心肌肥厚大鼠模型建立,然后处死大鼠,取出心脏组织,利用实时定量PCR法检测心肌组织中环状RNA rno_circRNA_016002.将心肌细胞H9C2饥饿过夜后分为ISO 50μmol/L与ISO 100μmol/L组、细胞对照组,ISO 50μmol/L与ISO 100μmol/L组中分别添加50与100μmol/L的ISO进行心肌肥厚的诱导,细胞对照组不作处理,利用实时定量PCR法检测肥大心肌细胞H9C2中环状RNA rno_circRNA_016002;将心肌细胞H9C2分为siRNA+ISO 50μmol/L组、siRNA+ISO 100μmol/L组、siRNA组、阴性对照组,其中siRNA+ISO 50μmol/L与siRNA+ISO 100μmol/L组加入siRNA与转染试剂后,分别再加入50与100μmol/L的ISO处理饥饿过夜;阴性对照组中只加入转染试剂,siRNA组中加入siRNA与转染试剂.加入ISO 48 h后利用实时定量PCR法检测各组心肌细胞H9C2中ANP mRNA与BNP mRNA,采用Western blotting检测各组心肌细胞H9C2中ANP、BNP蛋白质.结果 与Control组比较,ISO组环状RNA rno_circRNA_016002的表达水平上调(P<0.01);与细胞对照组比较,ISO 50μmol/L、ISO 100μmol/L组环状RNA rno_circRNA_016002的表达水平均上调(P均<0.01).siRNA+ISO 50μmol/L与siRNA+ISO 100μmol/L组中ANP、BNP mRNA和蛋白质表达与阴性对照组相比,P均>0.05.结论 rno_circRNA_016002在ISO诱导的大鼠肥厚的心肌组织和肥大心肌细胞中表达均上调,并且促进了心肌肥厚.

Diabetes mellitus and the associated complications are metabolic diseases with high morbidity that result in poor quality of health and life. The lack of diagnostic methods for early detection results in patients losing the best treatment opportunity. Oral hypoglycemics and exogenous insulin replenishment are currently the most common therapeutic strategies, which only yield temporary glycemic control rather than curing the disease and its complications. Exosomes are nanoparticles containing bioactive molecules reflecting individual physiological status, regulating metabolism, and repairing damaged tissues. They function as biomarkers of diabetes mellitus and diabetic complications. Considering that exosomes are bioactive molecules, can be obtained from body fluid, and have cell-type specificity, in this review, we highlight the multifold effects of exosomes in the pathology and therapy of diabetes mellitus and diabetic complications.

For patients with nonvalvular atrial fibrillation (NVAF) following hemorrhagic infarction (HI)/hemorrhage transformation (HT) and complicated with venous thrombosis, the management of anticoagulation is controversial. Our study intends to explore the safety and effectiveness of using low-dose of low molecular weight heparin (LMWH) to treat NVAF patients with HI (or HT) and complicated with venous thrombosis. Between January 2018 and January 2019, NVAF related acute ischemic stroke patients with HT/HI, hospitalized in the department of neurology or rehabilitation in our hospital, are enrolled retrospectively. Among them, those who were found to have venous thrombosis and undergo anticoagulation (LMWH) during the treatment were extracted. We investigate the efficacy and safety in those patients who have been treated with anticoagulant of LMWH. Five cases accepted LMWH within 3 weeks attributed to the appearance of venous thrombosis, and all of them did not display new symptomatic bleeding or recurrent stroke. However, based on the results of a head computed tomography scan, there were 2 cases of slightly increased intracranial hemorrhage, and then we reduced the dose of anticoagulant. In addition, color ultrasound showed that venous thrombosis disappeared or became stable. Patients with NVAF following HI/HT have a higher risk of thromboembolism. Early acceptance of low-dose LMWH as an anticoagulant is relatively safe and may gain benefit. However, in the process of anticoagulant therapy, we should follow-up head computed tomography/magnetic resonance imaging frequently, as well as D-dimer values, limb vascular ultrasound. Besides, the changes of symptoms and signs should be focused to judge the symptomatic bleeding or recurrent stroke. Furthermore, it is better to adjust anticoagulant drug dosage according to specific conditions.

Previous studies have shown that lysine-specific demethylase 1 (LSD1) could regulate cell cycle progression through demethylation. The 3'domain of HOX transcript antisense RNA (Hotair) combined with the LSD1/CoREST/REST complex helps LSD1 target the corresponding gene. However, its role in mice's myocardial regeneration is still unclear. The heart from neonatal mice shows strong myocardial regeneration ability, but this ability disappears 7 days after birth. Our study shows that the myocardial tissue highly expresses Hotair and Lsd1 within 1 week after birth, consistent with the myocardial regeneration time window. Knockdown Lsd1 or Hotair expression by RNA interference could inhibit myocardial regeneration and cardiomyocyte proliferation. Our results suggest that Hotair-mediated demethylation of LSD1 may play an important role in myocardial regeneration in neonatal mice.

Mounting evidence shows that long noncoding RNAs play important roles in human diseases and radioresistance could be an important factor for tumor recurrence. We observed that HOX antisense intergenic RNA (HOTAIR) expression increased in invasive ductal carcinoma (IDC) tissue. The irradiation effect of HOTAIR was detected by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay, cell cycle and apoptosis analysis, and xenografts in nude mice. Western blot, RNA pulldown assay, and RNA immunoprecipitation were used for mechanistic studies. HOTAIR, upregulated in IDC of breast cancer tissue, could promote breast cancer cell proliferation by regulating cell cycle and apoptosis. Overexpression of HOTAIR promoted DNA damage repair factors KU70, KU80, DNA-PKs, and ATM expression, and these could be impeded by small molecular inhibitors of enhancer of zeste homolog 2 (EZH2). Meanwhile, we found that HOTAIR could facilitate recruitment of EZH2 to the Avian Myelocytomatosis Viral Oncogene Homolog (MYC) promoter. HOTAIR is an important modulator not only to the prognostic of breast cancer, but also a good marker for radiotherapy.

目的 探索芫花素对于MC3T3-E1细胞向成骨细胞分化的影响.方法 采用芫花素1、5、10、15 μmol/L处理MC3T3-E1细胞14 d,采用MTS法检测芫花素对MC3T3-E1细胞增殖的抑制作用;实时定量PCR和Western blotting法检测测芫花素对MC3T3-E1细胞中ALP、HDAC1和Runx2 mRNA和蛋白表达的影响.结果 芫花素5、10、15 μmol/L可以显著促进MC3T3-E1细胞中ALP、Runx2 mRNA和蛋白表达水平的提高(P＜0.05),HDAC1 mRNA和蛋白表达水平的下调(P＜0.05),表明芫花素可以促进MC3T3-E1向成骨细胞分化.结论 芫花素具有促进MC3T3-E1细胞可以促进成骨细胞分化,对成骨细胞的分化过程有一定的调节作用.

目的 建立一种心肌组织稳定过表达Hotair的新生小鼠模型并进行验证.方法 采用改良后的重叠延伸PCR法从小鼠基因组中获取目的基因Hotair,构建过表达Hotair的慢病毒质粒,并采用高分子聚合物转染法进行包装及浓缩纯化,测序结果显示构建的慢病毒质粒Hotair序列正确.将18只P0期(出生24 h内)小鼠随机分为Hotair过表达组、阴性对照组、空白对照组,每组6只.Hotair过表达组于左侧胸腔第五肋间隙注射浓缩后的Hotair慢病毒10μL,阴性对照组和空白对照组分别注射等量空载病毒和生理盐水;各组注射4 d后处死小鼠,取心脏组织.采用实时定量荧光PCR法检测Hotair及受Hotair协同调控的赖氨酸特异性去甲基化酶1(LSD1)mRNA表达,Western blotting法检测LSD1调控的下游蛋白即组蛋白H3二甲基化的赖氨酸4(H3K4me2)表达.结果 Hotair过表达组心肌组织Hotair表达高于阴性对照组和空白对照组,H3K4me2蛋白表达低于阴性对照组和空白对照组(P均<0.01).Hotair过表达组与阴性对照组心肌组织LSD1 mRNA相对比表达量比较差异无统计学意义(P>0.05).结论 成功应用慢病毒载体建立心肌组织稳定过表达Hotair的新生小鼠模型;该模型构建方法简便快捷、Hotair过表达效果明显,为后续研究Hotair参与心脏疾病的相关机制提供了可靠的动物模型.

Objective To research the effect of alkylation of glycerol phosphate synthase (AGPS) in isoproterenol (ISO) induced rat cardiac hypertrophy. Methods The pathological cardiac hypertrophy rat model was constructed by ISO intraperitoneal injection. Twelve healthy Sprague-Dawley rats (120～150 g) were divided into ISO group and control group randomly. In the ISO group, rats were injected with ISO (3 mg/kg) per day for two consecutive weeks. In the control group, rats were injected with normal saline (3 mg/kg) per day for two consecutive weeks. Changes of left ventricular diastolic diameter, left ventricular posterior wall thickness, left ventricular ejection fraction, left ventricular short-axis shortening rate and left ventricular mass were detected by echocardiography. The cross-sectional area of myocardial cells in rats was measured by hematoxylin-eosin staining. The expression of hypertrophic factors [atrial natriuretic peptide (ANP), myosin light chain-2V (MLC-2V), α-myosin heavy chain (α-MHC)] and AGPS were detected by Western Blot and real-time quantitative PCR (qPCR). Results The results of echocardiography showed that the cardiac hypertrophy rat model was successfully constructed. The results of hematoxylin-eosin staining showed that the myocardial cross-sectional area in the ISO group was significantly larger than that of the control group. The Western Blot and qPCR results indicated that the relative expression of protein and mRNA of hypertrophic factor and AGPS in the ISO group were both up-regulated comparing with that of the control group, and the differences were statistical significance (all P<0.05). Conclusions The rat model of pathological cardiac hypertrophy with up-regulated AGPS expression was successfully constructed providing a theoretical basis for further study on the role of AGPS in pathogenesis of pathological cardiac hypertrophy.

目的 探索木兰花碱对慢性不可预见性温和应激所致抑郁小鼠行为学的影响机制.方法 将ICR小鼠随机分为正常对照组(control组)、抑郁模型组(PG组)、木兰花碱高剂量组(MH组)和木兰花碱低剂量组(ML组).采用慢性不可预见性温和刺激方法建立抑郁小鼠模型,检测各组小鼠行为学的变化,并用实时定量PCR、Western blot检测小鼠脑部赖氨酸特异性组蛋白去甲基化酶1(LSD1)mRNA及蛋白表达水平的变化.免疫组化染色法检测小鼠脑部LSD1阳性细胞数量.结果 木兰花碱在行为学上改善了小鼠的抑郁状况,与PG组相比,MH组和ML组悬尾不动时间、强迫游泳不动时间明显缩短(P<0.05).实时定量PCR及Western blot结果显示,与control组相比,PG组LSD1 mRNA表达差异无统计学意义,蛋白表达明显降低(P<0.05),ML组LSD1 mRNA和蛋白表达明显升高(P<0.05);与PG组相比,ML组LSD1 mRNA和蛋白表达明显升高(P<0.05).免疫组化结果显示,与control组相比,PG组小鼠脑部LSD1阳性细胞数量减少,而ML组较PG组增加.结论 在木兰花碱治疗抑郁小鼠的过程中,LSD1可能起到了重要的调节作用.

Objective To construct a human renal epithelial cell line HEK293T by CRISPR-Cas9-based site-directed knock-in of vascular endothelial growth factor 165 (VEGF165) gene, and avoid the off-target effect caused by lentivirus infection. Methods The VEGF165 expression vector with homologous arm (pUCm-T-VEGF165 plasmid) and the sgRNA expression vector [pSpCas9(BB)-2A-Puro-sgRNA plasmid] were designed and constructed based on the DNA sequence of the EZH2 gene, and then co-transfected into HEK293T cells. The expression of VEGF165 mRNA was detected by qPCR and the expressions of VEGF165 proteins were detected by Western Blot. Results The qPCR and Western Blot results showed that, comparing with the control, the pUCm-T-VEGF165 plasmid and pSpCas9(BB)-2A-Puro-sgRNA plasmid, the expression of the co-transfection plasmid were significantly increased, i.e. 3.42±0.30 vs. 1.02±0.21, 1.13±0.16 and 0.98±0.18 for the VEGF165 mRNA level (all P<0.01), and 1.13±0.16 vs. 1.02±0.06, 0.88±0.03 and 0.80±0.05 for the VEGF165 protein level (all P<0.01), respectively. Besides, the expression of EZH2 was significantly down-regulated, i.e. 0.14±0.06 vs. 1.08±0.11, 1.02±0.12 and 1.13±0.16 for the EZH2 mRNA level (all P<0.01), and 0.23±0.03 vs. 1.05±0.13, 0.91±0.04 and 0.81±0.06 for the EZH2 protein level (all P<0.01), respectively. This result showed that the VEGF165 was successfully inserted into the EZH2 genome, interfering the EZH2 expression. Conclusions VEGF165 gene can be successfully knocked into HEK293T cells by CRISPR/Cas9 system.

本研究旨在探讨富含甘油三酯的脂蛋白(triglyceride-rich lipoproteins,TRLs)中载脂蛋白CⅢ(apolipoprotein CⅢ,ApoCⅢ)的高表达对人肾小球系膜细胞(human mesangial cells,HMCs)表型转分化的作用及其机制.从正常野生型小鼠(WT)及ApoCⅢ转基因小鼠血浆中提取TRLs,在体外培养的HMCs中分别给予相同甘油三酯浓度的正常TRLs (TRL-WT),或富含ApoCⅢ的TRLs(TRL-CⅢ)孵育12 h.结果 显示,TRL-CⅢ中ApoCⅢ蛋白质表达水平约为WT小鼠TRLs的4倍.与阴性对照(NC)组相比,两种TRLs均可诱导HMCs中转分化标志物α-SMA及纤维化相关因子纤连蛋白、TGF-β1、CTGF、Col Ⅰ、ColⅣ的mRNA及α-SMA和纤连蛋白的蛋白质表达水平升高,且TRL-CⅢ较TRL-WT作用更明显.TRL-CⅢ组中,α-SMA及纤连蛋白蛋白质表达水平分别为TRL-WT组的1.4倍(P＜0.05)和2.1倍(P＜0.01).与NC组相比,两种TRLs均可引起细胞内活性氧(reactive oxygen species,ROS)水平增加,且TRL-CⅢ组较TRL-WT组高1.5倍(P＜0.05);氧化应激相关因子Nox4、P22phox、PKC-β的mRNA及Nox4蛋白表达水平在两组均增高,且TRL-CⅢ较TRL-WT作用更为明显(P＜0.05).TRL-CⅢ组中,Nox4蛋白表达水平为TRL-WT组的1.6倍(P＜0.01).NADPH氧化酶抑制剂apocynin可使TRL-CⅢ引起的细胞内ROS水平下降约2倍(P＜0.01).上述转分化标志物及纤维化相关因子基因及蛋白质表达水平均明显下降.本研究表明,TRLs通过氧化应激通路促进HMCs表型转分化,ApoCⅢ的高表达进一步促进该作用.

OBJECTIVE:To investigate the therapeutic effect of splenic ischemic preconditioning (sIPC) on renal ischemia-reperfusion (IR) injury.MATERIALS AND METHODS:A total of 18 adult male Sprague Dawley (SD) rats were treated by 45 min renal ischemia and followed by 24 h reperfusion. In the sIPC group, three cycles of splenic ischemic preconditioning including 5 min ischemia and 5 min reperfusion were carried out before renal ischemia. The blood samples and kidney tissues were collected after 24 h. The levels of Cr and BUN in serum were measured to evaluate the kidney function. The morphological changes in ischemia-reperfusion kidneys were determined by hematoxylin-eosin (HE) staining. The levels of pro-inflammatory cytokines including TNF-α and IL-6 in serum, and renal tissues, were measured by ELISA and Western Blotting. Furthermore, the levels of IKK-β, intra-nuclear NF-κB, p65, and IL-10 in renal tissues were also measured.RESULTS:The results demonstrated that the level of Cr and BUN in the IR group were increased while decreased in the sIPC group. HE staining showed that the damage caused by renal ischemia-reperfusion was attenuated by sIPC with a low renal injury score in the sIPC group. ELISA and Western Blotting results showed that the production and secretion of TNF-α and IL-6 induced by IR were inhibited by sIPC. The expression level of IKK-β and intranuclear p65 in renal tissues were increased in the IR group while sIPC had exhibited the function of depressing the increased expression levels of IKK-β and intranuclear p65. Compared with the IR group, the expression level of IL-10 of serum and renal tissues in the sIPC group were increased.CONCLUSIONS:sIPC exhibited a potent anti-inflammatory capacity to attenuated renal IR injury.

Objective To construct rat lysine-specific histone demethylase 1 (LSD1) overexpression plasmids and LSD 1 demethylation fragment disfunction plasmids,and to evaluate their expression levels in HEK293T cells.Methods LSD1 fragments were amplified by PCR,and LSD1 demethylation disfunction fragments were amplified by overlap PCR.Rat-specific LSD1 overexpression plasmids and LSD1 demethylation disfunction plasmids were constructed and verified by agarose gel electrophoresis and sequencing.HEK293T cells were infected using the validated recombinant plasmids and blank vectors,and the stably expressed cells were selected by puromycin.The expression of LSD1 in the stably expressed HEK293T was detected by Western Blot and real-time quantitative PCR.Results The results of agarose gel electrophoresis and sequencing showed that LSD1 overexpression plasmids and LSD1 demethylation disfunction plasmids were successfully constructed.The Western Blot and real-time quantitative PCR results showed that compared with the blank group,the relative expression of LSD1 and mRNA in the LSD1 overexpression group and the LSD1 demethylation disfunction group were up-regulated,and the differences were statistically significant(all P＜0.01).Conclusions The constructed LSD1 overexpression plasmids and LSD1 demethylationi disfunction plasmids can achieve overexpression of LSD1 gene in HEK293T cells.This paper lays a foundation for further study of the relationship between LSD 1 gene and related diseases and demethylation of LSD 1.