本文選題：維生素D依賴性佝僂病IA型 + CYP27B1�。� 參考：《福建醫(yī)科大學(xué)》2015年碩士論文

【摘要】：[目的]分析2例維生素D依賴性佝僂病IA型(Vitamin D-dependent rickets type IA,VDDR-IA)患者的臨床特點(diǎn)、個(gè)體化治療及隨訪資料,以提高對(duì)本病的認(rèn)識(shí)和診治水平,并通過(guò)2個(gè)VDDR-IA患者家系CYP27B1基因突變分析,為VDDR-IA的臨床診斷和治療提供依據(jù),同時(shí)也可豐富中國(guó)病例基因突變類型數(shù)據(jù)庫(kù)。[研究對(duì)象]2例VDDR-IA患者及家系成員17人,另收集健康志愿者101人作為對(duì)照,在獲得知情同意的情況下,由福建省福州兒童醫(yī)院內(nèi)分泌科專人采集外周血樣及收集相關(guān)病歷資料。所有受檢人員在此前均未接受過(guò)任何遺傳學(xué)相關(guān)檢測(cè)。[研究方法]1.臨床資料與方法1.1收集臨床相關(guān)資料:詳細(xì)記錄2例VDDR-IA患者及其家系成員的病史,進(jìn)行體格檢查,并完成相關(guān)的實(shí)驗(yàn)室檢查及影像學(xué)檢查,對(duì)病歷資料進(jìn)行回顧性總結(jié)分析。1.2血清1,25-(OH)2D3水平測(cè)定:抽取2例患者及其家系成員外周靜脈血,分離血清后應(yīng)用人1,25-(OH)2D3 ELISA檢測(cè)試劑盒進(jìn)行血清1,25-(OH)2D3水平測(cè)定。2.CYP27B1基因測(cè)序分析2.1外周靜脈血基因組DNA提取:抽取2例患者及其家系成員和101例健康對(duì)照組的外周靜脈血,采用TIANGEN血液基因組DNA提取試劑盒提取基因組DNA。2.2 PCR擴(kuò)增CYP27B1基因片段:參考文獻(xiàn)資料,應(yīng)用Oligo6軟件進(jìn)行引物驗(yàn)證及PCR條件分析后對(duì)CYP27B1基因9個(gè)外顯子及其內(nèi)含子相鄰區(qū)進(jìn)行PCR擴(kuò)增,通過(guò)預(yù)實(shí)驗(yàn)確定PCR擴(kuò)增最佳條件。2.3 PCR產(chǎn)物測(cè)序和分析:所有PCR擴(kuò)增后的產(chǎn)物均經(jīng)過(guò)1.5%瓊脂糖電泳檢測(cè)后,送測(cè)序公司切膠純化后直接測(cè)序,測(cè)序結(jié)果與Blastn中CYP27B1基因參考序列(NC_000012.11與NM_000785.3)進(jìn)行對(duì)比分析,發(fā)現(xiàn)存在的堿基變異。[結(jié)果]1.臨床資料:2例患者均有嚴(yán)重的佝僂病癥狀、體征及影像學(xué)表現(xiàn),實(shí)驗(yàn)室檢查提示低鈣血癥、低磷血癥、血清堿性磷酸酶及甲狀旁腺激素水平顯著升高、血清25-(OH)D3水平升高,1,25-(OH)2D3水平明顯降低。兩家系其他成員無(wú)相關(guān)臨床表現(xiàn),實(shí)驗(yàn)室檢查結(jié)果均正常。2.CYP27B1基因測(cè)序結(jié)果:在2例VDDR-IA患者CYP27B1基因8號(hào)外顯子檢測(cè)到同一種純合子突變c.1319_1325dup CCCACCC,通過(guò)對(duì)其正常表型的父系及母系成員的CYP27B1基因測(cè)序證實(shí)多人為雜合突變,101名正常對(duì)照個(gè)體CYP27B1基因相同位點(diǎn)測(cè)序未發(fā)現(xiàn)此突變。3.患者的治療隨訪:予以骨化三醇和鈣劑個(gè)體化治療,2例患者癥狀明顯改善,實(shí)驗(yàn)室檢查提示生化指標(biāo)恢復(fù)正常,影像學(xué)檢查顯示骨骼損害明顯改善。[結(jié)論]本研究首次對(duì)來(lái)自中國(guó)同一小城市的2個(gè)完整大家系進(jìn)行了基因突變分析。兩例患者均為CYP27B1基因8號(hào)外顯子c.1319_1325dup CCCACCC純合突變發(fā)病,在中國(guó)VDDR-IA病例中為首次報(bào)道。治療隨訪提示VDDR-IA藥物應(yīng)用需要個(gè)體化,對(duì)于病情嚴(yán)重者大劑量骨化三醇治療是必要的,且長(zhǎng)期應(yīng)用尚未發(fā)現(xiàn)副作用。
[Abstract]:[objective] to analyze the clinical characteristics, individualized treatment and follow-up data of 2 patients with vitamin D-dependent rickets type IADDR-IA (Vitamin D-dependent rickets type IADDR-IA) in order to improve the understanding and diagnosis of the disease, and to analyze the CYP27B1 gene mutation in two families of VDDR-IA patients. It can provide evidence for clinical diagnosis and treatment of VDDR-IA and enrich the database of gene mutation types in Chinese cases. [participants] two VDDR-IA patients and 17 family members, and 101 healthy volunteers were collected as control. With informed consent, peripheral blood samples were collected and relevant medical records were collected by the Endocrinology Department of Fuzhou Children's Hospital in Fujian Province. None of the subjects had previously undergone any genetically-related tests. [research methods] 1. Clinical data and methods 1.1 Clinical data were collected: the history of 2 VDDR-IA patients and their family members were recorded in detail, the medical examination was performed, and the relevant laboratory and imaging examinations were completed. Retrospective analysis of medical records. 1.2 determination of serum 1O25- (OH) 2D3 levels: peripheral venous blood samples were drawn from 2 patients and their family members. After isolation of serum, Elisa kit was used to detect the level of 1m25- (OH) 2D3 in serum. 2.CYP27B1 gene sequencing analysis 2.1 peripheral venous blood genomic DNA extraction: extraction of peripheral venous blood from 2 patients and their family members and 101 healthy controls. The CYP27B1 gene fragment was amplified by TIANGEN blood genomic DNA extraction kit. 2.2 PCR. The CYP27B1 gene fragment was amplified by PCR using Oligo6 software after primer verification and PCR condition analysis, and the adjacent regions of 9 exons and introns of CYP27B1 gene were amplified by PCR. The best conditions for PCR amplification were determined by pre-experiment. 2.3 PCR products were sequenced and analyzed. All PCR products were detected by 1.5% agarose gel electrophoresis, and then sequenced directly after being purified by gumming by sequencing company. The results of sequencing were compared with the reference sequence of CYP27B1 gene in Blastn (NCSCS 000012.11 and NMSTX 000785.3). [result] 1. Clinical data: two patients had severe rickets symptoms, signs and imaging findings. Laboratory examination showed hypocalcemia, hypophosphatemia, serum alkaline phosphatase and parathyroid hormone levels increased significantly. The serum 25- (OH) D _ 3 level increased and the serum 25-(OH) _ 2D _ 3 level decreased significantly. Other members of the two families had no related clinical manifestations, Laboratory results were normal. 2. Sequencing of CYP27B1 gene: the same homozygous mutation was detected in exon 8 of CYP27B1 gene in 2 patients with VDDR-IA. The sequence of CYP27B1 gene of normal phenotype and maternal member of CYP27B1 gene was confirmed by sequencing the homozygous mutation of CYP27B1 gene. The mutation of CYP27B1 gene was not found by sequencing the same locus of CYP27B1 gene in 101 normal controls. Follow up: two patients were treated with ossification triol and calcium individualized treatment. The results of laboratory examination showed that the biochemical indexes returned to normal and the bone damage was obviously improved by imaging examination. [conclusion] for the first time, gene mutation analysis was carried out in two complete lines from the same small city in China. The homozygous mutation of exon 8 of CYP27B1 gene c.1319_1325dup CCCACCC was reported for the first time in Chinese VDDR-IA patients. The follow-up indicated that the drug use of VDDR-IA should be individualized, and it was necessary to treat severe patients with high dose of oscitic triol, and no side effects had been found in the long term use of VDDR-IA.
【學(xué)位授予單位】：福建醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R591.44

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 曾國(guó)章;;佝僂病兒童血礦物質(zhì)元素測(cè)定及相關(guān)性分析[J];臨床和實(shí)驗(yàn)醫(yī)學(xué)雜志;2007年10期

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本文編號(hào)：2064126