110例视网膜色素变性患者RP1基因突变的检测分析

Mutations analysis of RP1 gene in 110 Chinese with retinitis pigmentosa

背景视网膜色素变性（RP）是一组常见的单基因遗传性、致盲性眼底病变，目前尚无有效的治疗方法。目的研究RP1基因在宁夏地区RP患者中的突变频率及特征，进一步探讨其在RP发病机制中的潜在作用。方法收集宁夏回族自治区110例RP患者，其中包含35例常染色体显性遗传性RP（ADRP）患者和75例散发患者为RP患者组，同时收集100例健康成年人为正常对照组，2组均采集外周静脉血3～5ml，应用聚合酶链反应（PCR）和直接测序法进行RP1基因全编码区及邻近剪切位点内含子区域序列突变的检测。运用多因素Logistic回归方法和基于网络的评分软件PolyPhen、Sorting Intolerantfrom Tolerant（SIFT）对研究结果进行分析。结果RP患者组和正常对照组RPl基因共检测出11个变异位点，P．Lysll52Lys和C．＊247A〉C为新发现的突变。RP患者组第3内含子C．788—92T〉C的突变频率与正常对照组比较差异有统计学意义（χ2=9．12，P〈0．01）；3’-侧翼序列区c．＊247A〉C的突变频率高于正常对照组，差异有统计学意义（χ2=12．77，P〈0．01），且与RP发生呈显著正相关（r=1．11，P〈0．05），其余位点突变均证实为RPl基因的多态性。RP患者组P．Gln1725Gin的突变率显著高于正常对照组（χ2=42．09，P〈0．01），但其与RP的发生无关（r=1．74，P〉0．05）。1例散发患者发现了P．Lysll52Lys的突变。在83例RP患者中发现有P．Arg872His、P．Alal670Thr和P．Serl69lPro突变的同时出现，PolyPhen分析显示，三者均为保护性突变，评分分别为1．11、0．74、0．22；SIFT分析显示，这3个突变均对RP表现为保护作用，评分分别为0．07、0．60、0．22；携带这3个突变的RP患者夜盲出现的平均年龄为30．54岁，27例未携带这3个突变的RP患者夜盲出现的平均年龄为21．06岁；携带这3个突变的RP患者平均最佳矫正视力为0．50±0．38，未携带者的为0．40±0．33，差异有统计学意义（t=2．11，P〈0．05）。结论在宁夏回族自治区RP患者中，RPl基因的致病突变率低于中国其他地区的人群。P．Arg872His、P．Alal670Thr和P．Serl691Pro突变的协同出现可能对RP起到一定的保护作用，同时也可能降低了RPl基因致病突变的发生率。

Background Retinitis pigmentosa （RP） is a monogenic inheritance and blinding disease of fundus oculi. There is not an effective therapeutic method now. Objective This work was to identify the mutations of RP1 gene in Chinese RP patients in Ningxia area and to explore the potential interactions in the pathogenesis of RP. Methods The periphery blood of 3-5 ml was collected from 110 individuals with RP（35 ADRP and 75SRP） and 100 normal controls in Ningxia area. Polymerase chain reaction（PCR） and direct DNA sequencing were used to screening the sequence alterations in the entire coding region and splice sites of RP1 gene. Multivariate analysis and two web-based programs（ PolyPhen and SIFT） were used to analyze the results. Results Eleven mutation locus were detected in the exon 4 of RP1 gene including two novel sequence variants： p. Lysl152Lys without a higher mutation rate in comparison with normal control group（χ2 =9. 12 P〈0.01 ） ,but c. ＊ 247A〉E with a higher mutation rate in comparison with normal control group （χ2= 12.77, P〈0.01 ） and c. ＊ 247A 〉 C mutation was thought to be correlated with RP（ r = 1. 1,1, P〈0. 05 ）. The other ten mutation locus were reported as single nucleotide polymorphisms （SNP）. The mutation rate of p. Gln1725Gln was found to be higher in the RP patients than the normal controls （χ2 =42. 09,P〈0.01 ）, but no the significant correlation was seen between the pathogenesis of RP and mutation of p. Gln1725Gln（r=1.74,P〉0.05）.p. Lysl152Lys mutation was found in only 1 patient. Three SNPs（p. Arg872His, Ala1670Thr,Scr1691 Pro） were always occurred in the same 83 RP patient and the relevance ratio was higher than controls （P〈0.01）. The age of night blindness on patients with concurrent three mutations was （30.54 ± 13.68 ） years, and the best corrected visual acuity （BCVA） was 0.50 ± 0.38. The age of night blindness on patients without concurrent three mutations was （21.06± 16.24） years, and the BCVA was 0.40 ±0.33 and were higher than controls （ t = 2. 11, P 〈 0.05 ）. Conclusions In this study, the prevalence of RP1 mutations among the RP patients in Ningxia population was lower than other populations （ 〈 1% ）. The alliance of SNPs （ p. Arg872His, p. Ala1670Thr, p. Ser1691Pro） may play a protective role on RP patients and reduce the frequency of mutatiaon in RP1 gene.