Defective DNA repair replication in xeroderma pigmentosum fibroblasts and DNA repair of somatic cell hybrids after UV irradiation.

Primary fibroblast cultures were obtained from 9 patients with xeroderma pigmentosum of various clinical types. Repair replication of the UV-damaged DNA in fibroblasts was studied by means of 3H-thymidine labeling and radioautography. A DNA repair replication was found to be decreased in all xeroderma pigmentosum fibroblasts as compared with the control cells obtained from normal donors. The repair activities in cells from patients ranged from nearly 0% in two infant cases and one case of De Sanctis-Cacchione syndrome to approximately 100% in adult moderate case. There was, however, no correlation between the level of repair replication and the severity of clinical symptoms. Since two cases which showed a lack of repair DNA replication were infant, it is assumed that these cases may develop De Sanctis-Cacchione syndrome in future. On xeroderma pigmentosum cells, a genetic analysis was performed with cell fusion methods using irradiated HVJ virus in order to determine the type of the complementation group. XP-1, XP-4 and XP-9 may be classified into the group D; XP-2, XP-7 and XP-8 into the group A; and XP-5 into the group E.     

Complementation of the DNA repair deficiency in human xeroderma pigmentosum group a and C cells by recombinant adenovirus-mediated gene transfer

Nucleotide excision repair (NER) is one of the most versatile DNA repair mechanisms, ensuring the proper functioning and trustworthy transmission of genetic information in all living cells. The phenotypic consequences caused by NER defects in humans are autosomal recessive diseases such as xeroderma pigmentosum (XP). This syndrome is the most sun-sensitive disorder leading to a high frequency of skin cancer. The majority of patients with XP carry mutations in the XPA or XPC genes that encode proteins involved in recognition of DNA damage induced by UV light at the beginning of the NER process. Cells cultured from XPA and XPC patients are hypersensitive to UV light, as a result of malfunctioning DNA repair. So far there is no effective long-term treatment for these patients. Skin cancer prevention can only be achieved by strict avoidance of sunlight exposure or by the use of sunscreen agents. We have constructed recombinant adenoviruses carrying the XPA and XPC genes that were used to infect XP-A and XP-C immortalized and primary fibroblast cell lines. UV survival curves and unscheduled DNA synthesis confirmed complete phenotypic reversion in XP DNA repair deficient cells with no trace of cytotoxicity. Moreover, transgene expression is stable for at least 60 days after infection. This efficient adenovirus gene delivery approach may be an important tool to better understand XP deficiency and the causes of DNA damage induced skin cancer.     

DNA修复基因XPD多态性在中国汉族（四川泸州）和泰国人群中的分布特点

目的:分析DNA修复基因着色性干皮病基因DLys751Gln基因型频率和等位基因频率在中国汉族(四川泸州)和泰国人群中的分布特点,并比较其在两个群体中的分布有无差异。方法:实验于2006-01进行。选择150名无血缘关系的泸州医学院附属医院健康体检者,均为汉族,25～65岁,平均45岁;140名泰国无血缘关系个体,均来自曼谷地区,年龄25～63岁,平均46岁。采用聚合酶链反应-限制性片段长度多态性、聚丙烯酰胺凝胶电泳银染显色方法检测所有受试者的Lys751Gln等位基因频率和基因型频率。采用Promega公司的统计学软件PowerStats进行Hardy-Weinberg平衡检测,应用χ2检验或Fisher确切概率法分析Lys751Gln等位基因及基因型频率在中国汉族、泰国及其他人群中的分布是否有差异,采用SPSS11.5软件完成。结果:290名受试者全部进入结果分析。①基因型Lys/Lys,Lys/Gln和Gln/Gln在泸州汉族人群中的频率分别为72.67%,27.33%和0.00%,在泰国人群中的频率分别为87.86%,12.14%和0.00%。②基因型频率分布符合Hardy-Weinberg平衡定律,采用χ2检验或Fisher确切概率法进行统计分析,两个人群中基因型频率和等位基因频率分布有显著性差异(P<0.05)。结论:Lys751Gln基因多态性在中国汉族(四川泸州)和泰国人群中的分布有显著性差异,其多态性分布存在地域和种族差异。     

Genetic correction of DNA repair-deficient/cancer-prone xeroderma pigmentosum group C keratinocytes

Xeroderma pigmentosum (XP) is a rare photosensitive and cancer-prone syndrome transmitted as an autosomal recessive trait. Most cancers developed by XP patients are basal and squamous cell carcinoma strikingly restricted to sun-exposed parts of the skin. Cells from patients with classic XP are deficient in nucleotide excision repair, a versatile biochemical mechanism for removal of ultraviolet-induced DNA lesions. Among the seven classic XP complementation groups known to date (XP-A to XP-G), XP-C is the most common one in Europe and North Africa and XP-C patients remain free of neurologic problems often seen in other XP complementation groups. This has prompted us to undertake genetic correction of XP-C fibroblasts and particularly keratinocytes, which are the most relevant cells in relation to skin cancer and have proven recently to be capable of reconstructing XP-C skin in vitro. In this study, we demonstrate that DNA repair capacity, cell survival properties, and transition from proliferative to abortive keratinocyte colonies toward UVB irradiation can be fully recovered in keratinocytes from patients with XPC transduced with a retroviral vector stably driving the expression of the wild-type XPC protein. In addition, we show that in the absence of UV, XP-C keratinocytes exhibit intrinsic cell cycle abnormalities, and beta(1)-integrin overexpression, defects that are also both fully reversed after genetic correction. Together with full correction of the defects in XP-C corrected keratinocytes, in vitro reconstruction of skin from these cells open a rational perspective to XP tissue therapy.     

Impaired interferon production and natural killer cell activation in patients with the skin cancer-prone disorder, xeroderma pigmentosum.

Xeroderma pigmentosum (XP) is a rare autosomal recessive disorder with sun sensitivity, markedly increased skin cancer susceptibility, and defective DNA repair without consistently identified symptoms of immune deficiency. We examined natural killer (NK) cell activity and interferon production in peripheral blood lymphocytes (PBL) of eight XP patients who had multiple primary skin cancers. The XP patients had normal numbers of T cells and NK cells, as well as normal lymphokine-activated killer cell activity and normal tumor necrosis factor-alpha production. Unstimulated NK cell function was 40% of normal controls in five XP patients, but was normal in three other XP patients. However, PBL from all the XP patients tested showed no enhancement of NK activity by the interferon inducer, polyinosinic acid:polycytidilic acid (polyIC) but enhancement by interferon-alpha was normal, suggesting an impairment in interferon production. Parallel studies in non-XP skin cancer patients revealed that both unstimulated and polyIC-enhanced NK activity were normal. Further investigation using PBL from XP patients revealed that the production of interferon-gamma after stimulation with interferon inducers (polyIC, interleukin 2, or K562 tumor cells) was 13-43% of normals. These data indicate that XP lymphocytes have a defect in production of interferons and suggest that defective interferon production, as well as DNA repair defects, may play an important role in the susceptibility of XP patients to skin cancer.     

Abnormal ultraviolet mutagenic spectrum in plasmid DNA replicated in cultured fibroblasts from a patient with the skin cancer-prone disease, xeroderma pigmentosum.

A shuttle vector plasmid, pZ189, was utilized to assess the types of mutations that cells from a patient with xeroderma pigmentosum, complementation group D, introduce into ultraviolet (UV) damaged, replicating DNA. Patients with xeroderma pigmentosum have clinical and cellular UV hypersensitivity, increased frequency of sun-induced skin cancer, and deficient DNA repair. In comparison to UV-treated pZ189 replicated in DNA repair-proficient cells, there were fewer surviving plasmids, a higher frequency of plasmids with mutations, fewer plasmids with two or more mutations in the marker gene, and a new mutagenic hotspot. The major type of base substitution mutation was the G:C to A:T transition with both cell lines. These results, together with similar findings published earlier with cells from a xeroderma pigmentosum patient in complementation group A, suggest that isolated G:C to A:T somatic mutations may be particularly important in generation of human skin cancer by UV radiation.     

Idiopathic intracranial hypertension with and without papilloedema in a consecutive series of patients with chronic migraine

Vieira DSS, Masruha MR, Gonçalves AL, Zukerman E, Senne Soares CA, da Graça Naffah-Mazzacoratti M & Peres MFP. Idiopathic intracranial hypertension with and without papilloedema in a consecutive series of patients with chronic migraine. Cephalalgia 2008; 609–613. London. ISSN 0333-1024
Chronic migraine (CM) has been associated with idiopathic intracranial hypertension without papilloedema (IIHWOP), a significant percentage of these cases occurring in obese patients with intractable headache. A prospective study from February 2005 to June 2006 was made of 62 CM patients who fulfilled International Headache Society diagnostic criteria and had cerebral magnetic resonance venography (MRV) and lumbar puncture (LP) done. Two patients were excluded, six (10%) with elevated cerebrospinal fluid (CSF) open pressure (OP), five with body mass index (BMI) > 25. None of the patients had papilloedema or abnormal MRV. BMI and CSF OP were significantly correlated ( r  = 0.476, P  < 0.001, Pearson's correlation test). Obesity (defined as BMI > 30) was a predictor of increase in intracranial pressure (defined as OP > 200 mmH₂O) (f = 17.26, 95% confidence interval 6.0, 8.6; P  < 0.001). From our study we strongly recommend that not only intractable CM patients with high BMI, but also first diagnosed patients with BMI > 30 should be systematically evaluated by a LP to rule out IIHWOP.     

Clinical,angiographic and cardiovascular magnetic resonance findings in consecutive patients with Takotsubo cardiomyopathy

BACKGROUND: The aim was to assess clinical, angiographic and cardiovascular magnetic resonance (CMR) findings in patients with Takotsubo cardiomyopathy. METHODS: Between 2003 and 2007, 20 consecutive patients admitted to our hospital with suspected acute myocardial infarction and presenting with apical ballooning in the left ventricular (LV) angiogram in the absence of a significant coronary artery disease, were included in the study. Echocardiography and CMR was performed in all patients. RESULTS: The mean age of patients with Takotsubo cardiomyopathy was 62 +/- 8 years (range 43-78 years). Eighteen (90%) were female. Clinical presentations included chest pain (95%) and cardiogenic shock (5%). The mean angiographic LV ejection fraction on admission was 45% +/- 9% (range 26%-60%) and resolved rapidly in all cases. Mean time delay between presentation CMR was 2 +/- 1 days (range 1-6 days). Mean ejection fraction was 51% +/- 15% (range 25%-81%). While 19 (95%) patients showed no evidence of late enhancement or signs of myocarditis in the CMR, 1 (5%) patient who was resuscitated showed hyperenhancement confined to the apex. CONCLUSION: In patients showing the clinical picture of an acute myocardial syndrome and angiographic picture of a TakoTsubo cardiomyopathy, CMR might be helpful in confirming the diagnosis through the exclusion of other causes for the acute LV dysfunction.     

Endovascular repair of abdominal aortic aneurysms: initial experience with 100 consecutive patients

OBJECTIVE: To review early results of endovascular repair of abdominal aortic aneurysms (AAAs). PATIENTS AND METHODS: The first 100 patients who underwent endovascular repair of AAA (EVAR) between June 26,1996, and October 31, 2001, at the Mayo Clinic in Rochester, Minn, were studied retrospectively to evaluate technical success, freedom from reinterventions, and early clinical outcome. RESULTS: A total of 89 men and 11 women (mean +/- SD age, 76 +/- 7 years; range, 47-92 years) underwent EVAR. The procedure was successful in 97 patients. There was no early death. Major complications occurred in 25 patients. The 30-day technical success rate was 86% (95% confidence interval [CI], 77%-92%). The median intensive care unit stay was 1 day (range, 1-15 days), and the median hospital stay was 3 days (range, 1-35 days). Median follow-up was 7 months (range, 1-60 months). Endoleak (incomplete seal of the endovascular graft) at discharge was observed in 14 patients; 13 developed endoleak during follow-up. There were 23 reinterventions, 65% of which were percutaneous procedures. One aneurysm ruptured at 5 months, but the patient was successfully treated by open repair. Primary and secondary graft patency rates at 1 year were 83% (95% CI, 74%-93%) and 94% (95% CI, 87 %-99%), respectively. The freedom from reintervention rate at 1 year was 71% (95% CI, 59%-84%), with an overall success rate from EVAR of 92% (95% CI, 84%-100%). There were no differences in early patency, reinterventions, and success rates between unibody and modular devices. CONCLUSION: EVAR can be performed with high technical success and low mortality rates; however, nonfatal complications and catheter-based reinterventions are frequent, and EVAR may not prevent aneurysm rupture. Although stent graft repair for high-risk patients is appealing, current data are insufficient to support EVAR as the preferred treatment of AAAs.     

DNA修复基因多态性与胃癌易感性的关系

目的分析中国西南地区人群DNA修复基因XRCCl和XPD基因多态性分布,探讨XRCCl和XPD基因多态性与胃癌发病风险的关系。方法采用1：1病例对照研究方法,采用基因测序法检测160例胃癌患者及160例正常人外周血GSTPl基因XPDAsp312Asn,XRCClArgl94Trp,和XRCClArg280His多态性,进行胃癌风险分析。结果胃癌组与正常对照组相比,XPD312、XRCCl280位点的多态性分布频率无明显差异,而XRCCl194Trp的分布频率在病例组和对照组中分别为17．2％和7．3％,具有统计学差异。XRCCl194Trp等位基因的个体其胃癌风险增高（OR=2．72,95％CI=1．04~7．24,P=0．027）。结论XRCClArgl94Trp基因多态性可增加人群患胃癌的风险,XPDAsp312Asn、XRCClArg280His基因多态性与胃癌易感性无关联。     

Cytogenetic studies in 174 consecutive patients with preleukemic or myelodysplastic syndromes

Routine cytogenetic studies were done in 174 consecutive patients with preleukemic or myelodysplastic syndromes (PL/MDS): 5 had the 5q - syndrome, 2 had refractory cytopenia, 43 had refractory anemia, 38 had refractory anemia with ringed sideroblasts, 69 had refractory anemia with excess blasts, 6 had refractory anemia with excess blasts in transition, and 11 had chronic myelomonocytic leukemia. Successful chromosome studies were accomplished in 167 patients (96%); 64 (37%) had a chromosomally abnormal clone. Abnormal clones were most common among patients who had refractory anemia with excess blasts (45%), refractory anemia with excess blasts in transition (60%), and chronic myelomonocytic leukemia (45%); they were least common among patients with refractory anemia (32%) and refractory anemia with ringed sideroblasts (21%). The two patients with refractory cytopenia had normal cytogenetic results. Each patient with the 5q - syndrome had a 5q-chromosome, as this is a prerequisite for the diagnosis. The two most common structural abnormalities were deletion of part of a chromosome 5 long arm (17 patients) and deletion of part of a chromosome 20 long arm (8 patients). Nonspecific structural abnormalities of chromosomes 1, 3, 6, and 17 were also common. The most common numeric abnormalities were monosomy 5 (7 patients), monosomy 7 (4 patients), loss of the Y chromosome (9 patients), and trisomy 8 (20 patients). No chromosome abnormalities were specifically associated with any PL/MDS classification.     

Therapeutic results of 5-fluorouracil in multiple and unresectable facial carcinoma secondary to xeroderma pigmentosum

A prospective study was done to test the efficacy of 5-fluorouracil (topical and systemic) in multiple and unresectable histologically proven facial squamous cell carcinomas (SCC) secondary to XP. Twelve patients (7M/5F, mean age 19.8 years) with multiple facial SCC were treated between 1994 and 1997. 5-FU was used as a twice-a-day local application in the documented areas, by continuous infusion associated with cisplatin (2 patients) and short infusion combined with folic acid (3 patients). Evaluation was done by clinical examination every two months for topical therapy and after every cycle for systemic treatment. Median treatment duration was 12 months (2 to 36 months). Treatment was well tolerated excluding episodes of pruritus in the treated areas. We observed mainly superficial tumour regression followed by dryness and crusting. In 5 cases, we performed biopsies after treatment showing in one case an extensive fibrosis with absence of tumour. However in the remaining 4 cases, despite a superficial reduction of tumour and a reconstitution of the epidermis, viable and unmodified squamous cell carcinoma remained in the deeper dermis. In the 5 patients treated by systemic 5-FU, we observed 1 complete response and 3 partial responses. Despite a dissociation between a good cosmetic result and a relatively superficial effect, topical 5-FU represents a useful therapeutic option in multiple unresectable facial SCC in patients with XP. Systemic chemotherapy is recommended in the event of more extended or profound lesions.     

Clinical and molecular genetic studies of Machado-Joseph disease

Studies on clinical and molecular genetic aspects of Machado-Joseph disease (MJD) are reviewed. MJD is now regarded as the most common autosomal dominant form of ataxia. Analyses of haplotypes and an intragenic polymorphism in the MJD1 gene, however, ruled out the possibility that founder chromosome is present among worldwide MJD patients. The expanded CAG repeats become unstable during parent-offspring transmission and the mechanism is being studied. Two factors contributing to the intergenerational instability have sofar been identified: 1) paternal transmission and 2) a intragenic polymorphism flanking the CAG repeats. Single sperm analysis of MJD patients, however, revealed paradoxical contraction of the CAG repeat size in sperm. Correlations between the size of the CAG repeats and genetic anticipation, clinical manifestation and morphologic changes are also discussed.     

Diagnostic and genetic studies in 43 patients with classic cystinuria

We present results for laboratory screening and diagnostic tests--cyanide-nitroprusside test, semi-quantitative thin-layer chromatography, and quantitative amino acid column chromatography--of 43 patients with classic cystinuria. We report the efficaciousness of the cyanide-nitroprusside test and of thin-layer chromatography, as compared with quantitative amino acid chromatography, for detecting heterozygotes for type II or III cystinuria. The quantitative results for aminoaciduria in 57 blood relatives in 23 families were used to categorize the index patients with classic cystinuria. By column chromatography the ranges of excretion rates (mumol/24 h per 1.73 m2 body surface area) of diagnostic amino acids in the index patients were as follows: cystine 556-54044 (normal 20-128), arginine 131-11543 (10-80), lysine 768-21848 (51-514), ornithine 185-5685 (0-80). Also by column chromatography the median values for arginine and ornithine excretion in cystine-lysinuric heterozygotes (among the 57 blood relatives) were significantly higher (P less than 0.01) than in controls but never approached the values for homozygotes. The cyanide-nitroprusside test results were positive in urine samples of 41 of 43 index patients and in 16 (51.6%) of the urine samples of 31 obligate heterozygotes with column chromatographically proven cystine-lysinuria. Thin-layer chromatography detected all of the homozygotes, all the compound heterozygotes, and 54.8% of the carriers. According to the type of aminoaciduria in their relatives, 11 patients with classic cystinuria could be classified as having classic cystinuria type I, 11 as having type II or III, and three as being compound heterozygotes. We discuss the implications of these results for correct diagnoses and for genetic studies in classic cystinuria.