Mitochondrial DNA alterations and mitochondrial dysfunction in the progression of hepatocellular carcinoma

Hepatocellular carcinoma(HCC)is one of the most common malignancies and is ranked third in mortality among cancer-related diseases.Mitochondria are intracellular organelles that are responsible for energy metabolism and cellular homeostasis,and mitochondrial dysfunction has been regarded as a hallmark of cancer.Over the past decades,several types of mitochondrial DNA(mtDNA)alterations have been identified in human cancers,including HCC.However,the role of these mtDNA alterations in cancer progression is unclear.In this review,we summarize the recent findings on the somatic mtDNA alterations identified in HCC and their relationships with the clinicopathological features of HCC.Recent advances in understanding the potential roles of somatic mtDNA alterations in the progression of HCC are also discussed.We suggest that somatic mtDNA mutations and a decrease in the mtDNA copy number are common events in HCC and that a mitochondrial dysfunction-activated signaling cascade may play an important role in the progression of HCC.Elucidation of the retrograde signaling pathways in HCC and the quest for strategies to block some of these pathways will be instrumental for the development of novel treatments for this and other malignancies.

Somatic alterations in mitochondrial DNA and mitochondrial dysfunction in gastric cancer progression

Energy metabolism reprogramming was recently identified as one of the cancer hallmarks.One of the underlying mechanisms of energy metabolism reprogramming is mitochondrial dysfunction caused by mutations in nuclear genes or mitochondrial DNA(mtDNA).In the past decades,several types of somatic mtDNA alterations have been identified in gastric cancer.However,the role of these mtDNA alterations in gastric cancer progression remains unclear.In this review,we summarize recently identified somatic mtDNA alterations in gastric cancers as well as the relationship between these alterations and the clinicopathological features of gastric cancer.The causative factors and potential roles of the somatic mtDNA alterations in cancer progression are also discussed.We suggest that point mutations and mtDNA copy number decreases are the two most common mtDNA alterations that result in mitochondrial dysfunction in gastric cancers.The two primary mutation types(transition mutations and mononucleotide or dinucleotide repeat instability)imply potential causative factors.Mitochondrial dysfunction-generated reactive oxygen species may be involved in the malignant changes of gastric cancer.The search for strategies to prevent mtDNA alterations and inhibit the mitochondrial retrograde signaling will benefit the development of novel treatments for gastric cancer and other malignancies.

Nuclear and mitochondrial DNA microsatellite instability in hepatocellular carcinoma in Chinese

AIM：To study the nuclear microsatellite instability (nMSI) at BAT26 and mitochondral microsalellite instability (mtMSI) in the occurrence and development of hepatocellular carcinoma and the relationship between nMSI and mtMSI.METHODS: nMSI was observed with PCR and mtMSI with PCR-SSCP in 52 cases of hepatocellular carcinoma.RESULTS:mtMSI was detected in 11 out of the 52 cases of hepatocellular carcinoma (21.2%). Among the 11 cases of hepatocellular carcinoma with mtMSI, 7 occured in one locus and 4 in 2 loci. The frequency of mtMSI in the 52 cases of hepatocellular carcinoma showed no correlation to sex, age,infection of hepatitis B, liver cirrhosis as well as positive AFP of the patients (P>0.05). In addition, nMSI was detected in 3 out of 52 cases of hepatocellular carcinoma (5.8%) and there was no correlation of the incidence of mtMSI to that of nMSI (P>0.05).CONCLUSION:mtMSI may be involved in the coccurrence and development of hepatocellular carcinoma and it is independent of nMSI.

Mutations of mitochondrial 12S rRNA in gastric carcinoma and their significance

AIM: To detect the variations of mitochondrial 12S rRNA in patients with gastric carcinoma, and to study their significance and the relationship between these variations and the genesis of gastric carcinoma.METHODS: PCR amplified mitochondrial 12S rRNA of 44 samples including 22 from gastric carcinoma tissues and 22 from adjacent normal tissues, was detected by direct DNA sequencing. Then laser capture microdissection technique (LCM) was used to separate the cancerous cells and dysplasia cells with specific mutations. Denaturing high performance liquid chromatography (DHPLC) plus allele-specific PCR (ASPCR), nest-PCR and polyacrylamide gel electrophoresis (PAGE)were used to further evaluate this mutant property and quantitative difference of mutant type between cancerous and dysplasia cells. Finally, RNAdraw biosoft was used to analyze the RNA secondary structure of mutant-type 12S rRNA.RESULTS: Compared with Mitomap database, some new variations were found, among which np652 G insertion and np716 T-G transversion were found only in cancerous tissues.There was a statistic difference in the frequency of 12S rRNA variation between intestinal type (12/17, 70.59%) and diffusive type (5/17, 29.41%) of gastric carcinoma (P＜0.05).DHPLC analysis showed that 12S rRNA np652 G insertion and np716 T-G transversion were heteroplasmic mutations.The frequency of 12S rRNA variation in cancerous cells was higher than that in dysplasia cells (P＜0.01). 12S rRNA np652 G insertion showed obviously negative effects on the stability of 12S rRNA secondary structure, while others such as T-G transversion did not.CONCLUSION: The mutations of mitochondrial 12S rRNA may be associated with the occurrence of intestinal-type gastric carcinoma. Most variations exist both in gastric carcinomas and in normal tissues, and they might not be the characteristics of tumors. However, np652 G insertion and np716 T-G transversion may possess some molecular significance in gastric carcinogenesis. During the process from normality to dysplasia, then to carcinoma, 12S rRNA tends to convert from homoplasmy (wild type) to heteroplasmy,then to homoplasmy (mutant type, np717 T-G).

18β-glycyrrhetinic acid regulates mitochondrial ribosomal protein L35-associated apoptosis signaling pathways to inhibit proliferation of gastric carcinoma cells

BACKGROUND Gastric carcinoma(GC)is a common gastrointestinal malignancy worldwide.Based on the cancer-related mortality,the current prevention and treatment strategies for GC still show poor clinical results.Therefore,it is important to find effective drug treatment targets.AIM To explore the mechanism by which 18β-glycyrrhetinic acid(18β-GRA)regulates mitochondrial ribosomal protein L35(MRPL35)related signal proteins to inhibit the proliferation of GC cells.METHODS Cell counting kit-8 assay was used to detect the effects of 18β-GRA on the survival rate of human normal gastric mucosal cell line GES-1 and the proliferation of GC cell lines MGC80-3 and BGC-823.The apoptosis and cell cycle were assessed by flow cytometry.Cell invasion and migration were evaluated by Transwell assay,and cell scratch test was used to detect cell migration.Furthermore,a tumor model was established by hypodermic injection of 2.5×106 BGC-823 cells at the selected positions of BALB/c nude mice to determine the effect of 18β-GRA on GC cell proliferation,and quantitative reverse transcription-polymerase chain reaction(qRT-PCR)was used to detect MRPL35 expression in the engrafted tumors in mice.We used the term tandem mass tag(TMT)labeling combined with liquid chromatography–tandem mass spectrometry to screen for differentially expressed proteins(DEPs)extracted from GC cells and control cells after 18β-GRA intervention.A detailed bioinformatics analysis of these DEPs was performed,including Gene Ontology annotation and enrichment analysis,Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis,and so on.Moreover,STRING database(https://string-db.org/)was used to predict proteinprotein interaction(PPI)relationships and Western blot was used to detect the expression of proteins of interest in GC cells.RESULTS The results indicated that 18β-GRA could inhibit the proliferation of GC cells in a dose-and timedependent manner.It could induce GC cell apoptosis and arrest the cell cycle at G0/G1 phase.The proportion of cells arrested at S phase decreased with the increase of 18-GRA dose,and the migration and invasiveness of GC cells were inhibited.The results of animal experiments showed that 18β-GRA could inhibit tumor formation in BALB/c nude mice,and qRT-PCR results showed that MRPL35 expression level was significantly reduced in the engrafted tumors in mice.Using TMT technology,609 DEPs,among which 335 were up-regulated and 274 were down-regulated,were identified in 18β-GRA intervention compared with control.We found that the intervention of 18β-GRA in GC cells involved many important biological processes and signaling pathways,such as cellular processes,biological regulation,and TP53 signaling pathway.Notably,after the drug intervention,MRPL35 expression was significantly down-regulated(P=0.000247),TP53 expression was up-regulated(P=0.02676),and BCL2L1 was down-regulated(P=0.01699).Combined with the Retrieval of Interacting Genes/Proteins database,we analyzed the relationship between MRPL35,TP53,and BCL2L1 signaling proteins,and we found that COPS5,BAX,and BAD proteins can form a PPI network with MRPL35,TP53,and BCL2L1.Western blot analysis confirmed the intervention effect of 18β-GRA on GC cells,MRPL35,TP53,and BCL2L1 showed dose-dependent up/down-regulation,and the expression of COPS5,BAX,and BAD also increased/decreased with the change of 18β-GRA concentration.CONCLUSION 18β-GRA can inhibit the proliferation of GC cells by regulating MRPL35,COPS5,TP53,BCL2L1,BAX,and BAD.

A Study on the D-loop Region of Mitochondrial DNA (mtDNA) Mutation in Cervical Carcinomas

Objective Background-study on genesis and development of tumor is mainly concentrated on gene mutation in nucleus.In recent years,however,the role of mitochondrial DNA(mtDNA) mutation in tumor genesis has been given more and more attention,which is the only extra-nucleus DNA in cells of higher animals.Carcinoma of the uterine cervix is a common tumor in gynecology,but there are few reports of mtDNA mutation in this area.The focus of this study was to investigate the mtDNA mutation in tumor tissues of cervical carcinomas and their relationship to tumorigenesis and tumor development.Methods The D-loop region of 24 cervical carcinomas together with the adjacent normal tissues were amplified by PCR and sequenced.Results Among the 24 cervical carcinomas,30 mutations in 9 patients′ specimen were identified with the mutations rate of 37.5%(9/24).There were 8 microsatellite instabilities among the mutations and 13 new polymorphisms which were not reported previously in the Genbank.Conclusions The D-loop region of mitochondrial DNA is a highly polymorphoric and mutable region and the mutation rate is relatively high in patients with cervical carcinomas.

RESTRICTION ENDONUCLEASE ANALYSIS OF MITOCHONDRIALDNA FROM HUMAN LUNG ADENOCARCINOMA CELLLINE SPC-A-1

Objective: To understand the role of mitochondrial DNA (mtDNA) in carcinogenesis. Methods: single-step method was used to isolate the mtDNA from human lung adenocarcinoma cell line SPC-A-1. The mtDNA was analyzed by restriction fragment length polymorphism (RFLP) with 11 kinds of restriction endonuclease, which were Pvu II, Xho I, Pst I, EcoR I, BstE II, Hind III, Hpa I, Bc1 I, EcoR V, Sca I and Xba I. Restriction map of mtDNA from SPC-A-1 cell was obtained by the single and double-digestion method. Results: It was found that no variation at 32 restriction-sites could be detected in the coding region of mtDNA from SPC-A-1 cell line. But a new site was found at nucleotide 16276 (EcoR V) within the noncoding region. Conclusion: These results indicate that the primary structure of gene coding region of mtDNA isolated from SPC-A-1 cell is highly stable. While the major variation of nucleotide is probably located in the noncoding region.

Expression of Mitochondrial Transcripts in Gastric MGC803 Cell Line Subjected by Hypoxia

OBJECTIVE To determine the transcriptional expression of mitochondrial genome （mtDNA） in MGC803 cell lines subjected by various time-phase hypoxic dispositions, and further to discuss the influence of mtDNA transcripts on hypoxic resistance to irradiation. METHODS The MGC803 cells exposed to anoxic environment were divided into control group （0 h）, hypoxic group （2 h, 8 h, 16 h, 24 h） and irradiated group after exposing the hypoxia. RTPCR was applied to detect the transcripts of cytochrome oxidase subunit Ⅰ （COI）, NADH dehydrogenase subunit 4 （ND4）, ND5, cytochrome b （cyt-b） and ATPase6 （ATP-6） in MGC803 cell lines at various time-phases of hypoxic, and after X-ray irradiation. Flow cytometry and colony formation assay were conducted to evaluate the cell cycle phase and survival fraction. RESULTS COI and ND4 transcripts of MGC803 cell lines were influenced remarkably by hypoxia. COI transcripts were decreased remarkably with the elongation time of exposing the hypoxic, and reduced to one fourth of its original amount of prehypoxia 24 h after exposing the hypoxia. ND4 transcripts were increased initially, and elevated to two folds 8 h after exposing the hypoxia, and then reduced to one second 24 h after exposing the hypoxia. Hypoxia resulted in G1 phase blockage, especially after hypoxia for 16 h. The survival fraction of MGCS03 ceils exposing the hypoxia in irradiated group showed that as the time of exposing the hypoxic before irradiation is prolonged, the survival fraction of MGC803 cells may have an elevated tendency. CONCLUSION The tumor cells with lower expression levels of the COI and the ND4 after exposing the hypoxic have stronger resistance to the radiation, which indicates that increasing the expression levels of the COI and the ND4 might be advantageous to enhance the sensitivity of hypoxic tumor cells to the radiotherapy.

Change and Signif icance of Mitochondrial DNA Copy Number in Esophageal Squamous Cell Carcinoma

OBJECTIVE To compare the differences of mitochondrial DNA (mtDNA) copies among the tissues of esophageal squamous cell carcinoma (ESCC), para-neoplastic tissue and normal mucous membrane of the esophagus, and to study the relationship between the mtDNA and the occurrence and devel- opment of esophageal squamous cell carcinoma. METHODS The mtDNA copies of 42 specimens with the ESCC, paraneoplastic mucous tissue and normal mucous membrane of the esophagus were determined using real-time fluorescence quantitative PCR. The mtDNA was analyzed using agarose gel electrophoresis. RESULTS The mtDNA from all of the tissues (42/42) from the ESCC, para-neoplastic tissue and normal esophageal mucous membranes was analyzed, showing that there were an average mtDNA copy number of 27.1894×106 μg DNA, 9.4102×106 μg DNA and 5.9347×106 μg DNA, from the respective tissues. There were signifi cant differences (F=27.83, P<0.05) in mtDNA copy number among the three. A positive band was shown at 403 bp after gel electrophoresis of the PCR products, and the lane where the ESCC mtDNA located was rather bright, which was in accordance with the result of the real-time PCR determination. CONCLUSION An increase in the mtDNA copy number is related to the occurrence and development of ESCC.

转录因子MYB转录调控MTFR2通过DNA损伤修复促进胃癌细胞化疗耐药性

目的探究v-myb禽成髓细胞病病毒癌基因同源物(MYB)转录调控线粒体裂变调节因子2(MTFR2)对胃癌(GC)细胞顺铂(DDP)耐药性的影响及分子作用机制。方法TCGA数据库分析GC中差异mRNA并预测上游调控分子,qRT-PCR检测MTFR2和MYB的表达,双荧光素酶和染色质免疫共沉淀(ChIP)实验验证MTFR2和MYB的调控关系,细胞计数盒8(CCK-8)检测细胞活力并计算IC_(50)值,流式细胞术检测细胞周期和细胞凋亡,彗星实验检测DNA损伤,蛋白质免疫印迹法检测DNA损伤相关蛋白(γ-H2AX、ATM、p-ATM)的表达。结果MTFR2在GC组织和细胞中显著高表达,敲低MTFR2能够降低细胞增殖,阻滞S期,诱导细胞凋亡,促进DNA损伤和DDP敏感性。生信预测MTFR2存在上游转录因子MYB,MYB在GC组织和细胞中的表达显著上调,双荧光素酶和ChIP验证了MTFR2启动子区域与MYB的结合关系。回复实验发现进一步过表达MTFR2能够逆转敲低MYB对GC细胞增殖和DDP耐药性的抑制作用。结论MYB上调MTFR2的表达通过DNA损伤途径促进GC细胞增殖和DDP耐药,表明靶向MYB/MTFR2调控轴可能是克服GC DDP耐药性的潜在途径。

Modulation of mitochondrial bioenergetics as a therapeutic strategy in Alzheimer＇s disease

Alzheimer’s disease （AD） is an increasingly pressing worldwide public-health, social, political and economic concern. Despite significant investment in multiple traditional therapeutic strategies that have achieved success in preclinical models addressing the pathological hallmarks of the disease, these efforts have not translated into any effective disease-modifying therapies. This could be because interventions are being tested too late in the disease process. While existing therapies provide symptomatic and clinical benefit, they do not fully address the molecular abnormalities that occur in AD neurons. The pathophysiology of AD is complex; mitochondrial bioenergetic deficits and brain hypometabolism coupled with increased mitochondrial oxidative stress are antecedent and potentially play a causal role in the disease pathogenesis. Dysfunctional mitochondria accumulate from the combination of impaired mitophagy, which can also induce injurious inflammatory responses, and inadequate neuronal mitochondrial biogenesis. Altering the metabolic capacity of the brain by modulating/potentiating its mitochondrial bioenergetics may be a strategy for disease prevention and treatment. We present insights into the mechanisms of mitochondrial dysfunction in AD brain as well as an overview of emerging treatments with the potential to prevent, delay or reverse the neurodegenerative process by targeting mitochondria.

线粒体DNA转录表达与胃癌发生关系的研究

目的:检测胃癌组织线粒体DNA的转录表达水平变化,探讨mtDNA转录表达变化与胃癌发生的关系。方法:应用RT-PCR法检测42例配对的胃癌和癌旁正常胃粘膜组织的线粒体编码基因COⅠ、ND4、ND5、cyt-b和ATP-6的转录表达差异,并以β-actin作为定量标准物,最后行琼脂糖凝胶电泳。结果:胃癌组织线粒体COⅠ和ND4的转录水平显著高于远癌正常胃粘膜组织,P<0.01;胃癌COⅠ和ND4的表达水平与胃癌的组织分化程度呈负相关;肠型胃癌ND4和COⅠ的表达高于弥漫型胃癌,P<0.05。结论:胃癌的发生需要线粒体提供的某些转录物和蛋白以维持其恶性表型所需要的能量和(或)其他物质,而且分化程度愈低愈需要这种活动以增强其恶性生物学表型和行为。

口腔鳞状细胞癌线粒体DNA高变Ⅱ区及高变Ⅲ区突变的功能意义

目的检测口腔鳞状细胞癌线粒体DNA(mtDNA)复制控制区D环(D-loop)区的高变Ⅱ区(HVR Ⅱ)及高变Ⅲ区(HVR Ⅲ)的突变情况,并探讨其意义。方法以癌旁组织和正常组织为对照,对7例口腔鳞状细胞癌组织样本的mtDNA D-loop HVR Ⅱ区及HVR Ⅲ区进行聚合酶链反应(PCR)扩增和测序分析。结果在7例患者的癌组织、癌旁组织、正常组织样本中共发现82个(56种)核苷酸改变,其中51个(26种)为核苷酸多态性改变;3个肿瘤组织样本中共发现31个(30种)突变,其中21个位于HVR Ⅱ区及HVR Ⅲ区范围内;癌旁组织及正常组织未发现突变;口腔鳞状细胞癌的mtDNA D-loop HVR Ⅱ区及HVR Ⅲ区突变率为42.9%(3/7)。结论 mtDNA D-loop HVR Ⅱ区及HVR Ⅲ区的突变与口腔鳞状细胞癌的发生相关,为寻找新的肿瘤基因诊断和肿瘤遗传易感性的标志物提供了依据。

热休克蛋白10和热休克蛋白的60表达、纯化及与线粒体DNA转录因子A相互结合

目的:表达、纯化热休克蛋白10(Hsp10)和热休克蛋白60(Hsp60),研究人类线粒体DNA转录因子A(TFAM)在体外与Hsp10,Hsp60间的相互交联作用。方法:使用Escherichia.Coli(E.coli)诱导表达含有6个组氨酸标记的Hsp10和Hsp60,镍离子鳌合树脂分离纯化后,体外分别与纯化TFAM共育,抗-TFAM抗体免疫沉淀,SDS-PAGE分离蛋白,CBB探测分析。结果:Hsp10可在体外与TFAM发生免疫共沉淀,Hsp60不能。结论:Hsp10可能为TFAM-复合体组分。

食管鳞癌组织中线粒体DNA拷贝数量的变化及意义

目的:比较线粒体DNA(mitochondrial DNA,mtDNA)拷贝数量在食管鳞癌、癌旁粘膜和正常食管粘膜组织间的差异,探讨mtDNA与食管鳞癌发生、发展的关系。方法:应用荧光定量PCR(real-time PCR)技术,定量检测42例食管鳞癌、癌旁粘膜和正常食管粘膜组织中mtDNA拷贝数;并采用凝胶电泳对mtDNA进行定性检测。结果:食管鳞癌组织中mtDNA检测率为100%(42/42),平均拷贝数为27.1894×106μgDNA;正常食管粘膜组织mtDNA检测率100%(42/42),平均拷贝数为5.9347×106μgDNA;癌旁粘膜组织mtDNA检测率100%(42/42),平均拷贝数为9.4102×106μgDNA,三者之间有显著差异(F=27.83,P<0.05)。PCR产物经凝胶电泳后,在403bp处显示阳性条带,且食管鳞癌组织所在的泳道较亮,与real-timePCR检测结果一致。结论:mtDNA拷贝数量的增加与食管鳞癌的发生、发展有关。

人食管鳞癌EC9706细胞线粒体DNA与凋亡的关系

目的建立人食管鳞癌EC9706细胞的无线粒体DNA(ρ°)细胞,探讨食管癌线粒体DNA与凋亡的关系。方法在细胞培养液中加EB50μg/ml、尿嘧啶50μg/ml、丙酮酸100μg/ml,进行连续传代培养,获得完全缺失mtDNA的细胞(ρ°细胞);运用实时荧光定量PCR技术,检测EB处理后不同时间的人食管鳞癌细胞EC9706 mtDNA的拷贝数,并采用琼脂糖凝胶电泳对mtDNA进行定性检测;采用TUNEL染色和流式细胞技术,检测EB处理后不同时间人食管鳞癌细胞EC9706的凋亡情况。结果成功建立了人食管鳞癌细胞EC9706的ρ°细胞,经实时荧光定量PCR鉴定,发现在EB存在下,随着细胞分裂,mtDNA拷贝数进行性减少,直到12天,mtDNA完全丢失;流式细胞术检测结果显示,EC9706细胞EB处理后,第4天、8天及12天细胞凋亡率(%)分别为(2.78±1.04)、(11.68±1.85)、(26.62±1.06),与对照组相比,差异均有统计学意义(P<0.05);TUNEL检测结果与上述一致,从第4天到第12天凋亡也逐渐增加。结论成功建立了EC9706ρ°细胞。随着EC9706细胞mtDNA拷贝数量的逐渐减少,细胞凋亡率逐渐增加,表明mtDNA在诱导细胞凋亡中起着一定调控作用,提示选择性地诱导食管癌细胞mtDNA损伤,使食管癌细胞mtDNA拷贝数量明显减少,进而诱导细胞凋亡,可望成为食管癌生物治疗的一个新靶点。

肺鳞癌mtDNA D-环序列微卫星不稳的研究

目的分析肺鳞癌患者癌组织线粒体DNA(mitochondrial DNA,mtDNA)非编码D-环区序列的微卫星不稳(mitochondrial microsatellite instability,mtMSI)现象并探讨其意义。方法应用改良一步法制备15例肺鳞癌患者癌组织mtDNA,PCR产物直接测序,对比分析D-环区序列mtMSI情况。结果15例肺鳞癌患者癌组织mtDNAD-环区中,全部出现了mtMSI(100%),在3个不同位点上共发现30个mtMSI。结论肺鳞癌患者mtMSI发生率高,D环区碱基序列的mt-MSI可能与肿瘤的发生发展有关。

一氧化氮诱导食管癌细胞线粒体DNA编码基因过表达

以人食管癌细胞系EC10 9作为驱赶方 (driver) ,以被一氧化氮 (nitricoxid ,NO)诱导的EC10 9作为实验方 (tester) ,应用抑制消减杂交 (suppressionsubtractivehybridization ,SSH)、反向mRNA斑点印迹和RNA印迹等技术手段研究了NO诱导的食管癌细胞中基因的过表达情况 .然后对过表达基因的表达序列标签 (expressedsequencetag ,EST)实施序列测定 ,并与GenBank进行BLAST同源性比较和序列突变分析 .结果先后两次从 6 9个SSH阳性克隆中共鉴定出 6个线粒体DNA (mitochondrialDNA ,mtDNA)编码的基因 ,即ND 4L、ND 4、COX 2、Lys tRNA、ATP 8和ATP 6 .表明NO可以诱导食管癌细胞mtDNA编码的基因过表达 .另外 ,在ND 4L/ND 4基因的片段 (10 736～ 114 4 9)上发现了三处同型单核苷酸置换 (10 872T→C ,110 0 1A→G ,11346A→G) ,在COX 2 /Lys tRNA/ATP 8/ATP 6基因片段 (80 11～ 85 89)上发现了一处单核苷酸缺失(8380A) .氨基酸序列分析表明 ,在NO诱导的EC10 9中可能存在着一种结构异常的ATP 8肽链 (一条在N端被截短的只有 11个氨基酸残基的肽链 ,而正常的ATP 8肽链为 6 8个氨基酸残基 ) .

原发性肝癌组织线粒体DNA含量检测及意义

目的检测原发性肝癌及其癌旁组织中线粒体DNA含量,探讨线粒体DNA含量变化与肝癌发生的关系。方法荧光实时定量聚合酶链反应(real-time PCR)法扩增22例肝癌及相应癌旁组织中线粒体DNA编码基因细胞色素C氧化酶Ⅱ(COⅡ),以核编码基因磷酸甘油醛脱氢酶(GAPDH)作为内参照,比较两种组织中线粒体DNA含量的差异。结果肝癌组织中线粒体DNA相对含量为156.54±107.54,癌旁组织线粒体DNA相对含量为290.24±187.37,差异有统计学意义(P<0.01)。结论肝癌组织线粒体DNA的含量与肝癌发生有密切关系,有望成为一种新的肝癌标志物。

食管鳞状细胞癌组织中线粒体DNA含量及拷贝数量的变化

目的:比较食管鳞状细胞癌组织、癌旁组织和正常食管组织中线粒体DNA(mitochondrial DNA,mtDNA)含量及拷贝数量的差异,探讨mtDNA与食管鳞状细胞癌发生、发展的关系。方法:应用半定量PCR法分别扩增62例食管鳞状细胞癌组织及其相应的癌旁组织和正常食管组织中的mtDNA编码区,琼脂糖凝胶电泳比较3种组织中mtDNA含量的差异;利用荧光定量PCR技术定量检测上述3组样品中的mtDNA拷贝数,并采用琼脂糖凝胶电泳对mtDNA含量进行定性检测。结果:食管鳞状细胞癌组织、癌旁组织及正常食管组织中mtDNA的检出率均为100%(62/62),但食管鳞状细胞癌组织中mtDNA相对含量高于癌旁组织及正常食管组织(P<0.05)。食管鳞状细胞癌组织、癌旁组织和正常食管组织中mtDNA的平均拷贝数分别为(26.79±0.92)×106、(26.43±0.96)×106和(25.42±0.85)×106,3组间比较差异有统计学意义(F=37.532,P<0.05)。2种PCR结果一致。结论:mtDNA拷贝数量的增加与食管鳞状细胞癌的发生、发展密切相关。