γ-射线上调线粒体DNA损伤修复相关因子的表达

目的 研究γ-射线对口腔鳞状细胞癌(OSCC)细胞线粒体DNA (mtDNA)损伤相关修复因子——线粒体转录因子A (Tfam)、8-氧鸟嘌呤DNA糖基化酶(Ogg)-1和DNA聚合酶-γ(Polg)表达的影响.方法体外培养OSCC-2和-5细胞株,甲噻唑四唑氮比色检测其存活率,RNA干涉技术敲除相关基因,半定量反转录...     

口腔癌线粒体DNA D-loop环区HVRⅠ的功能突变

目的研究口腔颌面部鳞状细胞癌的线粒体DNA（mitochondria DNA,mtDNA）复制控制区（D-Loop）HVRⅠ的突变情况。方法7例口腔鳞状细胞癌患者,取癌组织、癌旁组织及正常粘膜,提取线粒体DNA（mtDNA）,对其D环区HVRⅠ进行PCR扩增和测序分析,与GENBANK人类同源线粒体序列对比,寻找突变位点。结果在7例鳞癌组织中共发现3例突变,突变率为43%,其中突变位点共51个,A/G或C/T为13个,A、G转换为C、T为39个;碱基缺失12个,插入突变为6个。癌组织和癌旁组织中共同存在19个突变位点。在癌旁组织中发现突变共28个,A/G或C/T为8个,A、G转换为C、T为16个;碱基缺失4个,插入突变为1个。结论口腔癌线粒体DNA D环区HVRⅠ突变率较高,可能与口腔癌的发生相关。     

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、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 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。     

线粒体功能与口腔鳞状细胞癌关系的研究进展

口腔鳞状细胞癌(oral squamous cell carcinoma,OSCC)是头颈部常见的恶性肿瘤,近来发病率呈逐年上升的趋势。线粒体是真核细胞中参与多种细胞行为的动态细胞器,线粒体功能障碍与肿瘤发展关系密切,作为决定癌细胞死亡的开关,靶向线粒体已成为OSCC治疗的重点。本文对线粒体与肿瘤发生发展、OSCC治疗以及顺铂耐药性OSCC的关系进行综述。目前研究发现:线粒体功能障碍促进细胞癌变,癌细胞的线粒体形态及功能均发生显著改变;线粒体裂变的增加提高癌细胞的侵袭性,线粒体自噬功能失调可诱导癌细胞凋亡;新型药物的出现以及定向药物递送系统纳米技术的发展开拓了靶向线粒体治疗OSCC的新方法,降低了全身用药的副作用;OSCC的顺铂耐药性通过线粒体途径产生,明确线粒体功能及线粒体DNA的突变机制,为靶向线粒体治疗顺铂耐药性OSCC提供新思路。     

线粒体DNA突变与头颈肿瘤发生的研究进展

线粒体DNA是核外唯一的遗传物质,较核DNA更易发生突变。在头颈肿瘤中,线粒体DNA所存在的点突变、片段缺失和微卫星不稳定性等现象可能与头颈肿瘤的发生相关,故线粒体DNA有望成为一种检测头颈肿瘤的新的生物标志物。笔者下面就线粒体DNA的结构和功能特点、头颈肿瘤中线粒体DNA的突变、线粒体DNA突变与头颈肿瘤发生的关系、线粒体DNA突变的临床价值等研究进展作一综述。     

Sirt1调控上皮细胞衰老的研究进展

在牙釉质发育过程中,成釉细胞过早衰老和凋亡是遗传性牙釉质发育不全的重要原因。沉默信息调节因子2相关酶1(silent matingtype information regulator 2 homolog 1,Sirt1)是一种依赖烟酰胺腺苷二核苷酸（nicotinamide adenine dinucleotide,NAD+）的脱乙酰酶,已被广泛报道参与调节细胞衰老。本文就Sirt1调控上皮细胞衰老研究进展作一综述,从Sirt1的结构特点入手,阐述Sirt1与衰老的关系。研究表明,当上皮细胞受到外界刺激时,Sirt1通过多种途径影响上皮细胞的衰老：Sirt1参与调节线粒体功能和代谢稳态,线粒体功能障碍会影响细胞衰老表型;端粒长度与衰老呈负相关,Sirt1调节端粒延伸所需的端粒逆转录酶的表达,从而正向调节端粒的稳态;DNA受损后会经历损伤修复,未修复的DNA损伤会引起细胞衰老,Sirt1/p53通路可通过减轻DNA损伤抑制上皮细胞衰老;衰老细胞是慢性炎症的来源,慢性炎症也可以多种方式促成衰老,Sirt1通过缓解炎症症状抑制上皮细胞衰老。未来可重点关注Sirt1对成釉细胞衰老的影响,探究其...     

表皮生长因子对人牙髓细胞DNA合成及细胞周期的影响

目的 研究表皮生长因子(EGF)对体外培养的人牙髓细胞(HDPCs)DNA合成及细胞周期的影响。方法 将培养的第5代HDPCs分成EGF(1ug/ml)实验组和对照组,分别以含5％FBS的DMEM培养48h;常规消化、固定细胞,调整细胞密度为2.O×10~6/ml,经DNA荧光染色后用流式细胞仪(FCM)测定DNA分子含量。结果与对照组相比,EGF实验组HDPCs的DNA合成前期细胞比例(G_1％)明显降低,而DNA合成期细胞比例(S％)及细胞增殖指数PrI值(S+G_2M)％均显著增高。结论 EGF具有促进HDPCs的DNA合成和分裂增殖的作用,可能主要是通过促使处于G_1期的细胞进入S期来实现的;提示EGF在牙髓组织的损伤修复过程中起着重要作用。     

Prrx1在骨发育和骨改建中作用的研究进展

转录因子配对相关同源框1(paired related homeobox 1,Prrx1)作为一种间充质细胞的标志物，不仅参与胚胎时期的骨发育过程，亦在机体成年后骨稳态的调节与损伤后的修复过程中发挥作用。Prrx1+细胞是间充质细胞的一种亚群，作为成骨的前体细胞参与骨骼的发育与形成。本文对Prrx1及Prrx1+细胞在骨发育及骨改建中的作用及影响进行综述。     

POLQ表达降低对SACC-83细胞DNA损伤敏感性的影响

目的 ：探讨DNA损伤环境中抑制POLQ对唾液腺腺样囊性癌（salivary adenoid cystic carcinoma,SACC）细胞SACC-83增殖及克隆形成能力、细胞周期及DNA损伤修复通路的影响。方法：应用shRNA(short hairpin RNA)瞬时转染方法构建POLQ敲减的SACC-83细胞模型，利用qRT-PCR及Western免疫印迹实验检测POLQ干扰效率。应用不同浓度DNA损伤剂依托泊苷（etoposide,ETP/VP-16-213）诱导SACC-83细胞发生DNA损伤，利用Western免疫印迹实验检测γH2AX的表达水平，评价DNA双链断裂水平。在不同浓度依托泊苷诱导形成的DNA损伤环境中，通过CCK-8细胞增殖实验检测抑制POLQ对SACC-83细胞增殖能力的影响。采用平板克隆实验观察抑制POLQ对SACC-83细胞克隆形成能力的影响，应用流式细胞仪分析抑制POLQ对SACC-83细胞周期的影响，应用Western免疫印迹实验检测抑制POLQ对SACC-83细胞γH2AX、RAD51及PARP1表达水平的影响。采用SPSS 20.0软件包对数据...     

不同烤瓷合金材料诱导细胞DNA损伤与细胞凋亡的研究

目的研究不同牙科烤瓷合金材料诱导细胞DNA损伤与细胞凋亡的情况,评价各种烤瓷合金材料的生物相容性。方法选择镍铬合金、钛合金、钴铬合金和金合金四种牙科烤瓷合金材料,制备四种合金材料的浸提液,用浸提液培养L929细胞24h。分别采用单细胞凝胶电泳(彗星试验)和流式细胞技术检测细胞DNA损伤与细胞凋亡的情况。结果镍铬合金、钛合金、钴铬合金可诱导不同程度的细胞DNA损伤和细胞凋亡(P<0.05),而金合金与对照组间无统计学差异(P>0.05)。结论不同烤瓷合金材料诱导细胞DNA损伤和细胞凋亡的情况存在差异性。金合金是种生物相容性优越的牙科烤瓷合金材料。     

New evidence of premature oxidative DNA damage: mitochondrial DNA deletion in gingival tissue of patients with periodontitis

BACKGROUND: Overproduction of reactive oxygen species (ROS) causes increased oxidative stress in gingival tissue. It has been generally accepted that increased oxidative stress might contribute to additional damage of lipids, proteins, and DNA molecules. The mitochondrial DNA (mtDNA) mutation is a superb biomarker of oxidative damage. The aim of the present study was to investigate the mtDNA deletions in the gingival tissue of patients with periodontitis and to explain the correlations between mtDNA deletion in gingival tissue and clinical parameters of periodontitis and age. METHODS: Gingival tissue and blood samples were collected from 30 patients with chronic periodontitis (CP group) and 30 healthy control subjects (H group). To determine the clinical condition of each subject, the plaque index, gingival index, clinical attachment level, and probing depth were measured. Using the polymerase chain reaction (PCR) method, we examined the 7.4- and 5-kbp mtDNA deletions in tissue and blood samples. Three different pairs of PCR primers were used in this study. RESULTS: In this study, we did not detect any deletions in blood DNA samples in either the CP or H group. Also, the 7.4-kbp mtDNA deletion was not detected in gingival tissues of subjects. However, the 5-kbp mtDNA deletion was detected in 24 of the 30 subjects (80%) in the CP group and was not detected in the H group (0%). Significant correlations were found between the occurrence of the 5-kbp mtDNA deletion and all clinical parameters (P <0.01). A similar correlation was found between the occurrence of the 5-kbp mtDNA deletion and age (P <0.05). CONCLUSIONS: The overproduction of ROS by activated polymorphonuclear leukocytes in chronic inflammation may lead to premature oxidative damage of the mtDNA. In this study, the occurrence of the 5-kbp mtDNA deletion in 24 periodontitis subjects may be evidence of premature oxidative DNA damage.     

The underlying mechanisms and strategies of DNA damage and repair in radiation sialadenitis

Radiation therapy is a critical strategy for the treatment of malignant tumors. X-ray external radiation has been successfully used to treat head and neck cancer. On the other hand, ¹³¹I internal radiation has been effective in managing differentiated thyroid cancer. However, these therapies cause radiation damage to salivary glands. Radiation sialadenitis is the most common complication associated with radiotherapy applied to the head and neck and it severely affects patients’ quality of life. Since DNA is the main intracellular target of radiation, and the integrity of the DNA structure is critical to genomic stability and the cellular survival of salivary glands, regulating radiation-induced DNA damage offers great promise in preventing and managing radiation sialadenitis. In this review, we summarize recent progress in DNA damage and repair in irradiated salivary glands.     

氟对大鼠氟斑牙形成和成釉细胞DNA损伤的研究

目的：研究在氟中毒引起氟斑牙时,氟对大鼠切牙成釉细胞DNA损伤的影响。方法：给雄性SD大鼠饮用含10、50、100mg/LNaF的高氟水120d,制备氟斑牙模型,用单细胞凝胶电泳检测DNA的损伤。结果：雄性SD大鼠饮用高氟水后,血清氟含量较对照组显著增高,P〈0.05。饮水氟含量与血清氟含量的相关系数为0.9153（P〈0.05）,具有较好的剂量-效应关系,大鼠切牙呈现典型的氟斑牙改变。在50mg/LNaF的剂量条件下,大鼠切牙成釉细胞彗星长与对照组比较,P〈0.05。在100mg/LNaF的剂量条件下,彗星长、Olive尾距、尾分布距与对照组比较,P〈0.05,而尾长值虽比对照组增加,但P〉0.05。低剂量染氟组（10mg/LNaF）大鼠切牙成釉细胞DNA损伤情况与对照组比较,没有显著性差异（P〉0.05）。结论：在氟中毒引起氟斑牙的过程中,大鼠切牙成釉细胞发生明显的DNA损伤。     

吸烟致口腔癌发生分子机制的研究

目的：研究吸烟对口腔黏膜细胞的遗传毒性，分析与口腔癌发生的关系。方法：采用单细胞凝胶电泳技术，检测24例吸烟者和17例非吸烟者（其中口腔恶性肿瘤12例，良性肿瘤19例，10例健康者）口腔黏膜脱落上皮细胞DNA损伤，按组织病理类型进行分析，观察指标为彗星尾长和彗星率。试验所得数据由SPSS、SAS软件进行统计检验和分析。结果-恶性肿瘤患者的彗星尾长和彗星率均明显高于良性肿瘤患者和健康人，有显著性差异（P〈0．01），良性肿瘤患者与健康者的彗星试验结果无显著差异（P〉0．05），吸烟者的彗星尾长和彗星率均明显高于非吸烟者（P〈0．01）。结论：吸烟可导致口腔黏膜细胞DNA损伤，随肿瘤组织病变程度的加重，DNA损伤程度亦加重。组织细胞DNA损伤与口腔恶性肿瘤的发生密切相关。     

Stimulatory effect of Aggregatibacter actinomycetemcomitans DNA on proinflammatory cytokine expression by human gingival fibroblasts

ObjectiveWhile different virulence factors have been reported of Aggregatibacter actinomycetemcomitans (Aa), there is little information about the stimulatory effect of its DNA. The main purpose of this study was to assess the inflammatory response of human gingival fibroblasts (HGFs) stimulated with A. actinomycetemcomitans DNA.DesignCytokine levels of IL-6, IL-1α and TNF-α were measured on the supernatant of HGFs activated with 10, 25, 50 and 100 μg/ml DNA of Aa during 24 h. Primary cultures of HGFs were infected with Aa and its DNA at different times and concentrations to compare its cytotoxic effect. Cell damage and adhesion of Aa to HGFs were evaluated under light microscopy and Scanning electron microscopy respectively.ResultsThere was a statistical difference (p < 0.05) in cytokine expression in HGFs activated by bacterial DNA with a dose dependent on IL-6 expression and a significantly elevated expression of IL-1α and TNF-α compared to Human DNA negative control. Substantial morphological alterations were observed after infection of A. actinomycetemcomitans in HGFs but not with bDNA exposure. Aggregatibacter actinomycetemcomitans showed a high rate of adhesion and cell damage to HGFs after 30 min.ConclusionsGenomic DNA of A. actinomycetemcomitans could be a factor in the pathogenesis of periodontitis that might play a major role in the inflammatory response.     

Human mitochondrial DNA and nuclear DNA isolation from food bite marks

ObjectiveBite mark analysis is used for comparison between bite marks on a bitten object and the suspects’ teeth. However, if it is not possible to obtain a correct match, it is important to recover salivary DNA. Previous studies have tried to isolate human nuclear DNA from bitten foods but were not completely successful. In the present work, we studied the efficiency of human nuclear and mitochondrial DNA isolation from bite marks in cheese, a donut and an apple.DesignUsing a double swab technique and silica-based DNA extraction kit, nuclear and mitochondrial DNA were isolated. Human housekeeping genes were amplified to analyse the efficiency of nuclear DNA profiling. mtDNA was sequencing and haplogroup assign.ResultsAlthough cheese and apple samples showed the highest concentration of DNA, the purity of DNA on the apple was low. Moreover, apple samples failed to amplify the two human housekeeping genes, GAPDH and RPL22. In contrast, cheese samples have high purity and amplification efficiency. Donut samples showed an intermediate value and low amplification efficiency. In spite of these results, isolation and characterization/sequencing of human mitochondrial DNA was completely successful in the three samples, which pointed out the possibility of identification through this type of DNA.ConclusionsThis research indicated that it is possible to recover and isolate human nuclear and mitochondrial DNA from bitten foods, although the quantity and purity of nuclear DNA depends on the type of food. That is of significance important in forensic sciences for the correct identification of a suspect.     

Identification of unknown body using DNA analysis and dental characteristics in chest X-ray photograph

An unknown skeletonized body was identified by DNA analysis and dental information. The body had already been cremated when a candidate for the unknown body was proposed. Therefore, for DNA analysis we used teeth that had been kept for a long time after use for serological examination. We also used a chest X-ray photograph of the candidate and photographs of dentition, as well as dental X-ray photographs taken when the unknown body was found. Because DNA obtained from teeth was highly degraded, we amplified three PCR fragments to determine the 766 bp mitochondrial DNA (mtDNA) sequence including HV1 and HV2. Polymorphism of the ABO locus was also analyzed using small PCR fragments. Although the isolated DNA was contaminated, probably with DNA from a different individual, DNA polymorphisms of mtDNA and the ABO locus could be analyzed. We discuss the reliability of our conclusions from the point of view of the necessity of constructing an accurate mtDNA database. Although a dentist who had treated the teeth of the unknown body could not be found, a chest X-ray photograph for medical diagnosis was very useful in comparing dental characteristics, as it included an image of the frontal part of the lower jaw and upper teeth.