Correlation research of Runt-related transcription factor 2 with proliferation genes, tumor suppressor genes and angiogenesis molecules in colon cancer lesions

Objective: To investigate the correlation of Runt-related transcription factor 2 (RunX2) with proliferation genes, tumor suppressor genes and angiogenesis molecules in colon cancer lesions. Methods: A total of 90 patients with primary colon cancer were enrolled in colon cancer group, 68 patients with benign colon polyps were enrolled in colon polyps group, the differences in the expression levels of RunX2, proliferation genes, tumor suppressor genes and angiogenesis molecules in the two groups of lesions were compared, and Pearson test was further used to evaluate the correlation of RunX2 expression level with proliferation gene, tumor suppressor gene and angiogenesis molecule expression levels in colon cancer tissues. Results: RunX2 mRNA expression level in the lesions of colon cancer group was higher than that of colon polyps group. Proliferation genes GTPBP4, HOXB7, ZNF331, ADAM17 and HSP60 mRNA expression levels in the lesions of colon cancer group were higher than those of colon polyps group;tumor suppressor genes ATF3, FOXN3, OTUD1 and NDRG2 mRNA expression levels were lower than those of colon polyps group;angiogenesis molecules Musashi 1, NF-κB, RegⅣ and STAT3 mRNA expression levels were higher than those of colon polyps group. RunX2 mRNA expression level in the colon cancer lesions was directly correlated with the expression levels of the above proliferation genes, tumor suppressor genes and angiogenesis molecules. Conclusion: RunX2 expression is abnormally high in colon cancer lesions, the specific expression level is positively correlated with cancer cell proliferation activity and angiogenesis activity, and it is an important molecular target that can lead to the occurrence and development of colon cancer.

Molecular and cellular changes in the post-traumatic spinal cord remodeling after autoinfusion of a genetically-enriched leucoconcentrate in a mini-pig model

Post-traumatic spinal cord remodeling includes both degenerating and regenerating processes,which affect the potency of the functional recovery after spinal cord injury(SCI).Gene therapy for spinal cord injury is proposed as a promising therapeutic strategy to induce positive changes in remodeling of the affected neural tissue.In our previous studies for delivering the therapeutic genes at the site of spinal cord injury,we developed a new approach using an autologous leucoconcentrate transduced ex vivo with chimeric adenoviruses(Ad5/35)carrying recombinant cDNA.In the present study,the efficacy of the intravenous infusion of an autologous genetically-enriched leucoconcentrate simultaneously producing recombinant vascular endothelial growth factor(VEGF),glial cell line-derived neurotrophic factor(GDNF),and neural cell adhesion molecule(NCAM)was evaluated with regard to the molecular and cellular changes in remodeling of the spinal cord tissue at the site of damage in a model of mini-pigs with moderate spinal cord injury.Experimental animals were randomly divided into two groups of 4 pigs each:the therapeutic(infused with the leucoconcentrate simultaneously transduced with a combination of the three chimeric adenoviral vectors Ad5/35‐VEGF165,Ad5/35‐GDNF,and Ad5/35‐NCAM1)and control groups(infused with intact leucoconcentrate).The morphometric and immunofluorescence analysis of the spinal cord regeneration in the rostral and caudal segments according to the epicenter of the injury in the treated animals compared to the control mini-pigs showed:(1)higher sparing of the grey matter and increased survivability of the spinal cord cells(lower number of Caspase-3-positive cells and decreased expression of Hsp27);(2)recovery of synaptophysin expression;(3)prevention of astrogliosis(lower area of glial fibrillary acidic protein-positive astrocytes and ionized calcium binding adaptor molecule 1-positive microglial cells);(4)higher growth rates of regeneratingβIII-tubulin-positive axons accompanied by a higher number of oligodendrocyte transcription factor 2-positive oligodendroglial cells in the lateral corticospinal tract region.These results revealed the efficacy of intravenous infusion of the autologous genetically-enriched leucoconcentrate producing recombinant VEGF,GDNF,and NCAM in the acute phase of spinal cord injury on the positive changes in the post-traumatic remodeling nervous tissue at the site of direct injury.Our data provide a solid platform for a new ex vivo gene therapy for spinal cord injury and will facilitate further translation of regenerative therapies in clinical neurology.

Adhesion molecule and proinflammatory cytokine gene expression in hepatic sinusoidal endothelial cells following cecal ligation and puncture

INTRODUCTIONMultiple organ dysfunction syndrome (MODS) isthought to be a frequent consequence of sepsis[1-3].Despite substantial advances in our knowledge and understanding of the basic pathophysiologic mechanisms[4-7], in critically ill patients infections and sepsis are still associated with a high mortality[8,9].

Real-Time PCR Technique and Its Application in Quantification of Plant Nucleic Acid Molecules

Real-time PCR is a closed DNA amplification system that skillfully integrates biochemical, photoelectric and computer techniques. Fluorescence data acquired once per cycle provides rapid absolute quantification of initial template copy numbers as PCR products are generated. This technique significantly simplifies and accelerates the process of producing reproducible quantification of nucleic acid molecules. It not only is a sensitive, accurate and rapid quantitative method, but it also provides an easier way to calculate the absolute starting copy number of nucleic acid molecules to be tested. Together with molecular bio-techniques, like microarray, real-time PCR will play a very important role in many aspects of molecular life science such as functional gene analysis and disease molecular diagnostics. This review introduces the detailed principles and application of the real-time PCR technique, describes a recently developed system for exact quantification of AUX/IAA genes In Arabidopsis, and discusses the problems with the real-time PCR process.

Aberrant expression of genes and proteins in pterygium and their implications in the pathogenesis

Pterygium is a common ocular surface disease induced by a variety of factors. The exact pathogenesis of pterygium remains unclear. Numbers of genes and proteins are discovered in pterygium and they function differently in the occurrence and development of this disease. We searched the Web of Science and PubMed throughout history for literatures about the subject. The keywords we used contain pterygium, gene, protein, angiogenesis, fibrosis, proliferation, inflammation, pathogenesis and therapy. In this review, we summarize the aberrant expression of a range of genes and proteins in pterygium compared with normal conjunctiva or cornea, including growth factors, matrix metalloproteinases and tissue inhibitors of mefalloproteinases, interleukins, tumor suppressor genes, proliferation related proteins, apoptosis related proteins, cell adhesion molecules, extracellular matrix proteins, heat shock proteins and tight junction proteins. We illustrate their possible mechanisms in the pathogenesis of pterygium as well as the related intervention based on them for pterygium therapy.

Re-expression of Cell Adhesion Molecule Inhibits Growth and Induces Apoptosis of Human Pancreatic Cancer Cell Line PANC-1

This study examined the expression of cell adhesion molecule 1 （CADM1） in pancreatic cancer and the possible mechanism. The expression of CADM 1 was detected by immunohistochemistry in tissues of pancreatic cancer, pancreatitis, and normal pancreas. The plasmid pcDNA3.1-Hy- gro（＋）/CADM1 was transfected into PANC-1 cells （a pancreatic cancer cell line）. The expression of CADM1 in the transfected cells was determined by RT-PCR and Western blotting. Cell growth was measured by the MTT method and cell apoptosis by flow cytometry. The results showed that CADM1 was weakly expressed in tissues of pancreatic cancer in contrast to its high expression in normal pancreatic and pancreatitis tissues. The expression level of CADM in pancreatic caner was intensely correlated with the differentiation degree, lymph node metastasis and TNM stages. The growth of CADMl-transfected PANC-1 cells was significantly suppressed in vitro by a G1 cell cycle arrest and apoptosis occurrence. It was concluded that re-expression of CADM1 inhibits the growth of pancreatic cancer cells and induces their apoptosis in vitro. As a tumor suppressor gene, CADM1 plays an important role in the occurrence, progression and metastasis of pancreatic cancer.

整合素β3基因多态性蛋白质变异及其与疾病关系研究进展

整合素(ITG)β3基因位于17号染色体长臂2区1号3带(17q21.3),基因区间在47253827~47313743,全长59917 kb,编码区间在47253862~47310204,全长56343 kb,有15个外显子、14个内含子;编码1648个氨基酸残基,分子量为105 kDa。ITGβ3蛋白是一种二价离子依赖性细胞黏附分子,广泛存在于细胞膜中。其主要功能是介导细胞之间,细胞和细胞外基质之间的识别、连接和黏附,主导细胞内外跨膜接头蛋白连接的作用。近年来,关于ITG在细胞识别与黏附、细胞活化与分化、免疫反应、细胞信号传导、炎症反应、创伤修复与愈合、凝血与血栓形成、肿瘤形成与转移等方面发挥重要作用的研究取得许多进展,本文综述如下。

Screening and Application of SSR Markers of Resistant Gene against Rice Stripe Virus

[Objective] New SSR primers were designed and screened to apply in the backcross breeding for modified resistance against rice stripe virus.[Method] The conventional late japonica rice varieties including 502 with high resistance to stripe virus,Xiushui 09 with high susceptibility to stripe virus and their derived strains were adopted as the test materials,SSR and SAPR markers were used to locate RSV1 gene with high resistance against stripe virus,and three pairs of SSR markers （M-11-1,M-11-2,M-11-3） were further designed.Through screening and analysis,M-11-3 was selected as the RSV1 detection marker gene for tracking RSV1 gene,thus RSV1 gene was successfully introduced to the backcross breeding of late japonica rice varieties such as Xiushui 09,and the resistance expression of different strains was identified.[Result]The resistance of improved strains against stripe virus was significantly higher than Xiushui 09,the resistance of most strains was close to the level of donor,and the expression of resistance among years was stable.Therefore,the resistance effect of RSV1 gene used in the test was very obvious,which was accurate with the assisted selection of RSV1 gene linked markers M-11-3.[Conclusion]The study certified the feasibility of molecular markers application in resistance improvement against rice stripe virus,which also showed that optimization and development of new marker genes could effectively improve the efficiency of marker-assisted selection.

SSCP和HMA方法在马MHC-I类分子基因多态性研究中的应用

文章使用SSCP和HMA两种基于聚丙烯酰胺凝胶电泳的方法对马MHC-I类分子基因多态性进行了分析。在应用SSCP法进行分析时,尽管经过实验条件优化,仍未得到对MHC-I基因理想的分离效果,提示该方法对分离多态性较高的基因有一定局限性。在对HMA法用参考标准DNA对影响DNA分子构象的温度和变性剂浓度等实验条件进行优化后,获得了对马MHC-I类分子基因较好的分离效果。6、7、8、9和10号马的样本在相对应泳道上分别出现了6、5、6、5和7个条带。从凝胶中进行DNA条带回收后克隆测序的结果表明,这一方法可以有效地分离高度多态性的MHC-I类分子基因。

重庆地区RhD变异型无偿献血者的基因多态性及其表型研究

目的对重庆地区22例RhD变异型无偿献血者标本进行Rh血型血清学检测和三代基因测序,了解重庆地区RhD变异型的表型分布及其基因分型。方法选择2023年1—8月本中心参与无偿献血的人群作为研究对象。使用传统血清学方法对其进行RhD表型鉴定,确定为RhD变异型后使用D-screen试剂盒对其进行RhD不同抗原表位的检测。此外,提取其基因组DNA,使用叠瓦式引物设计进行多段扩增、拼接获得RHD基因全长序列进行三代测序检测,并用SnapGene软件对序列结果进行分析。结果在22例RhD变异型中,8例为DVI 3型(36.36%),其存在RHD-CE(3-6)-D杂交等位基因;6例部分弱D15型(27.27%),主要发生的突变为c.845G>A;6例亚洲型Del(27.27%),主要发生的突变为c.1227G>A,还有1例弱D17型发生的突变为c.340C>T和1例推测为部分D(c.491A>T,p.Asp164Val,错义突变)。结论重庆地区无偿献血人群中最常见的RhD变异型为DVI 3型,使用SMRT三代测序技术可以获得RhD变异型单倍体全长。

细胞间黏附分子-1基因多态性与缺血性心肌病易感性及预后的关联

目的探讨细胞间黏附分子-1(intercellular adhesion molecule-1,ICAM-1)基因多态性与缺血性心肌病(ischemic cardiomyopathy,ICM)易感性及预后的关系。方法选取因ICM入院的患者64例为ICM组,另选取健康体检者138例为对照组,采集两组血液标本并提取DNA。应用SNaPshot基因分型技术检测ICAM-1基因rs12462944、rs281430、rs281434、rs3093030、rs5491、rs923366共6个位点单核苷酸多态性,分析其与ICM易感性及预后的相关性。结果ICM组rs5491位点变异基因型AT频率高于对照组(P<0.05),等位基因T频率高于对照组(P<0.05),显性遗传模型中,rs281434位点变异基因型(AG+GG)频率高于对照组(P<0.05)。多因素Logistic回归分析结果显示,rs5491位点AT型携带者ICM风险是AA型的3.428倍(95%CI:1.152~7.747)。多因素COX回归分析结果显示,rs5491位点AT基因型是ICM心源性死亡的危险因素(HR=5.075,P<0.05),Kaplan-Meier生存曲线分析结果显示,AT基因型ICM患者较AA基因型3年内死亡风险更高(P<0.05)。结论ICAM-1rs5491基因型的变异提高了ICM的患病风险,AT型ICM患者较AA型预后较差。ICAM-1rs281434多态性可能与ICM易感性相关。

Niemann-Pick C1蛋白在埃博拉病毒感染中的作用及其靶向药物研究进展

埃博拉病毒属丝状病毒科,具有高传染性,能引起人类和灵长类动物出现严重出血热等症状,病死率高达90%。Niemann-Pick C1(NPC1)蛋白是埃博拉病毒感染过程中表达于宿主细胞内体膜上的一个重要受体,其与埃博拉病毒被组织蛋白酶裂解的糖蛋白(GP)的相互作用是病毒感染宿主的关键环节,介导病毒囊膜与内体膜的融合,进而将病毒基因组释放到宿主细胞。近年来,将NPC1蛋白作为广谱抗丝状病毒药物靶点研发的小分子抑制剂、单克隆抗体和基因治疗药物均有突破性进展。本文介绍了NPC1的结构及其在埃博拉病毒感染中的作用,并对靶向NPC1的小分子抑制剂、单克隆抗体药物和基因治疗药物的研究现状进行总结。

干扰素刺激基因在肿瘤免疫中作用机制的研究进展

干扰素刺激基因(ISGs)是一种经干扰素诱导、刺激后显著表达的基因。可通过免疫抑制分子的浸润,免疫检查点抑制,免疫编辑等方式抑制或者促进肿瘤细胞的生长浸润。本文对ISGs在肿瘤免疫中的具体作用机制及其在肿瘤药物治疗中发挥的作用作一综述,有望为开发更为精准的治疗策略提供理论基础。

RPS6亚基肽段抑制S180肿瘤细胞的分子机制

为探究核糖体蛋白RPS6亚基肽段对S180肿瘤细胞基因表达的影响及抑制肿瘤细胞的关键分子和信号通路,以S180荷瘤小鼠为模型,用RPS6亚基肽段处理S180荷瘤小鼠,对S180肿瘤细胞进行Illumina测序获得差异表达的基因,并对这些差异基因进行GO和KEGG分析。基因表达结果显示,RPS6亚基肽段会导致mt-Cytb、mt-Nd1、Rpl13基因表达降低,阻碍S180肿瘤细胞的氧化磷酸化过程。GO分析结果显示,RPS6亚基肽段抑制肿瘤细胞关键门类集中于质膜和质膜外侧组成成分的变化及免疫反应调节和免疫细胞的激活。差异基因结果显示,RPS6亚基肽段通过增强PRL/PRLR信号,上调Uchl1基因表达,诱导肿瘤细胞周期停滞。KEGG通路富集分析结果显示,穿孔素(PRF1)和颗粒酶B(GZMB)表达诱导细胞凋亡,同时自然杀伤细胞(natural killer cell,NK)介导的细胞毒性与NF-κB信号通路信号增强,IFN-γ和TNF-α表达上调共同参与RPS6亚基肽段作用下的S180肿瘤细胞凋亡,抑制S180肿瘤细胞增殖。RPS6亚基肽段导致S180肿瘤细胞细胞周期停滞,并诱导自然杀伤细胞介导的细胞毒性及NF-κB信号通路的上调导致S180肿瘤细胞凋亡,为RPS6亚基肽段在抗肿瘤研究的应用提供一定的理论依据。

Msx homeobox gene family and craniofacial development

Vertebrate Msx genes are unlinked,homeobox-containing genes that bear homology to the Drosophila muscle segment homeobox gene.These genes are expressed at multiple sites of tissue-tissue interactions during vertebrate embryonic development.Inductive interactions mediated by the Msx genes are essential for normal craniofacial,limb and ectodermal organ morphogenesis,and are also essential to survival in mice,as manifested by the phenotypic abnormalities shown in knockout mice and in humans.This review summarizes studies on the expression,regulation,and functional analysis of Msx genes that bear relevance to craniofacial development in humans and mice.

Combination of epidural electrical stimulation with ex vivo triple gene therapy for spinal cord injury:a proof of principle study

Despite emerging contemporary biotechnological methods such as gene-and stem cell-based therapy,there are no clinically established therapeutic strategies for neural regeneration after spinal cord injury.Our previous studies have demonstrated that transplantation of genetically engineered human umbilical cord blood mononuclear cells producing three recombinant therapeutic molecules,including vascular endothelial growth factor(VEGF),glial cell-line derived neurotrophic factor(GDNF),and neural cell adhesion molecule(NCAM)can improve morpho-functional recovery of injured spinal cord in rats and mini-pigs.To investigate the efficacy of human umbilical cord blood mononuclear cells-mediated triple-gene therapy combined with epidural electrical stimulation in the treatment of spinal cord injury,in this study,rats with moderate spinal cord contusion injury were intrathecally infused with human umbilical cord blood mononuclear cells expressing recombinant genes VEGF165,GDNF,NCAM1 at 4 hours after spinal cord injury.Three days after injury,epidural stimulations were given simultaneously above the lesion site at C5(to stimulate the cervical network related to forelimb functions)and below the lesion site at L2(to activate the central pattern generators)every other day for 4 weeks.Rats subjected to the combined treatment showed a limited functional improvement of the knee joint,high preservation of muscle fiber area in tibialis anterior muscle and increased H/M ratio in gastrocnemius muscle 30 days after spinal cord injury.However,beneficial cellular outcomes such as reduced apoptosis and increased sparing of the gray and white matters,and enhanced expression of heat shock and synaptic proteins were found in rats with spinal cord injury subjected to the combined epidural electrical stimulation with gene therapy.This study presents the first proof of principle study of combination of the multisite epidural electrical stimulation with ex vivo triple gene therapy(VEGF,GDNF and NCAM)for treatment of spinal cord injury in rat models.The animal protocols were approved by the Kazan State Medical University Animal Care and Use Committee(approval No.2.20.02.18)on February 20,2018.

Establishment of hepatocarcinoma cell line transfected by the B7 gene and its biocharacteristics

OBJECTIVE: To study the function of costimulation sign in tumor immunology and construct the new cell line B7^+ Smmc7721. METHODS: The B7 gene was transfected into the hepatocarcinoma cell Smmc7721 by liposma. Polymerase chain reaction (PCR) and reverse transcriptase-PCR (RT-PCR) were applied to test the result. MTT colorimetric assay was used to value the killing effect of lymphokine activated killer cells (LAK) activated by IL-2 on the transfected cell line and the original line. RESULT: B7^+ Smmc7721 was improved to be steadily expressed B7 molecule and LAK cells could more effectively act on the B7^+ Smmc7721 cells. CONCLUSION: The B7 gene can be transfected to hepatocarcinoma cells and can be expressed steadily in vitro, thus increasing the efficiency of LAK cells activated by IL-2 on them.

ANTITUMOR EFFECT OF INTRATUMORAL INJECTION OF LIPOSOME-ENCAPSULATED G-CSF GENE AND IN SITU BIOLOGICAL CHARACTERISTICS OF THE TREATED TUMOR CELLS

This word was supported by grant from Military Medical Research Foundation of china (96z032). ** To whom correspondence and requests for reprints should be addressed. This is one of papers of the special issue on gene therapy research (Chin J Cancer Res Vol. 9 No. 4 December, 1997). In order to investigate the antitumor effects of the in vivo G CSF gene therapy mediated by liposome and its mechanisms, human G CSF gene was encapsulated into liposome and was directly injected into tumor mass of C 26 colon adenocarcinoma bearing mice. After direct intratumoral injection of liposome encapsulated G CSF DNA, the subcutaneous tumor growth was dramatically inhibited and the survival time was prolonged signifi cantly. Tumor regression could be observed in about 30% of C 26 bearing mice. By the analysis of the antitumor mechanisms, we found that anti G 418 (600ug/ml) clone could be selected from the tumor cells freshly separated from the treated C 26 tumor mass, and secretion of G CSF in the supernatant could be detected. Northern blot also confirmed the expression of hG CSF by the tumor cells. Higher expressions of MHC class I(H 2k d) molecule and ICAM 1 on the tumor cells could be observed. The results demonstrated that liposome can effectively transfect G CSF gene into tumor cells in situ , and then increase the immunogenicity of the tumor cells which may contribute to the activation of the local antitumor immune responses effectively.

IMMUNOLOGICAL CHARACTERISTICS OF THE LEUKEMIA CELLS TRANSFECTED WITH ONCOSTATIN M GENE BY RECOMBINANT ADENOVIRUS

In the present study, FBL3 murine erythroleukemia cells were transfected with human OSM(hOSM) gene by recombinant adenovirus, then the immunological properties of hOSM genetransfected FBL3 cells(FBL3OSM+) were investigated. 4 hours after transfection with hOSM gene, hOSM could be detected in the supernatant of FBL3OSM+ cells and hOSM secretion peaked at 24 h. The proliferation of FBL3OSM+ cells was inhibited markedly. The clonal formation of FBL3OSM+ cells was suppressed more obviously in comparison with wildtype FBL3 cells when analysed in clonal argar culture. Flow cytometry analysis showed that FBL3OSM+ cells expressed higher levels of Fas protein, B7 and ICAM1 molecules.FBL3OSM+ cells also expressed higher level of MHC class I molecules(H2Kb) but remained unchanged in expression of MHC class II molecules (Ia). CD14, which is a specific marker of monocyte/macrophage and not expressed on the wildtype FBL3 cells, was also detected on the surface of FBL3OSM+ cells. The results suggested that OSM gene transfer could increase the immunogenicity of FBL3 cells and promote their differentiation into macrophagelike cells. The data outline a promising approach to OSM gene therapy of leukemia mediated by recombinant adenovirus.

Correlation of contrast-enhanced ultrasound parameters with oncogene expression and cell proliferation activity in breast cancer

Objective: To study the correlation of contrast-enhanced ultrasound parameters with oncogene expression and cell proliferation activity in breast cancer. Methods: Breast cancer lesions and benign breast lesions surgically removed in Zigong Third People's Hospital between May 2014 and February 2017 were selected, contrast-enhanced ultrasound was done before operation to draw the time-intensity curve and calculate the area under the curve (AUC), and the expression of proliferation molecules and tumor suppressor genes were detected after operation. Results:The contrast-enhanced ultrasound parameter AUC of the breast cancer lesion was greatly higher than that of the benign breast lesion;ECT2, ZKSCAN3, USP39 and EphA2 mRNA expression in breast cancer lesions were obviously higher than those in benign breast lesions whereas HPK1, TCEAL17, CCN5, ATG2B and ATG4D mRNA expression were greatly lower than those in benign breast lesions;ECT2, ZKSCAN3, USP39 and EphA2 mRNA expression in breast cancer lesions with high AUC were greatly higher than those in breast cancer lesions with low AUC whereas HPK1, TCEAL17, CCN5, ATG2B and ATG4D mRNA expression were greatly lower than those in breast cancer lesions with low AUC. Conclusion: The contrast-enhanced ultrasound parameter AUC of breast cancer lesion significantly increases and is closely related to the higher expression of pro-proliferation molecules and the lower expression of tumor suppressor genes.