【摘要】：背景及目的 顱內(nèi)出血（Intracranial hemorrhage，ICH）是新生兒常見的一種嚴(yán)重顱腦損傷，早產(chǎn)兒多見，胎齡越小，發(fā)病率越高，嚴(yán)重者可留有神經(jīng)系統(tǒng)后遺癥，甚至死亡。近年來，隨著醫(yī)學(xué)技術(shù)的不斷發(fā)展，早產(chǎn)兒存活率大大提高，ICH的發(fā)生率有增無減。 ICH的發(fā)生與自身的解剖生理特點(diǎn)和多種圍產(chǎn)期高危因素有關(guān)，此外，許多炎性反應(yīng)和內(nèi)源性介質(zhì)等也可誘發(fā)，其中內(nèi)源性神經(jīng)介質(zhì)特別是內(nèi)源性阿片肽的釋放增加能加重這種損傷。近年來，神經(jīng)肽與腦血管疾病機(jī)理研究日益受到關(guān)注,研究發(fā)現(xiàn)阿片肽及阿片受體系統(tǒng)在成人缺血性腦卒中的病理生理發(fā)展過程中具有非常重要的作用。 臨床研究發(fā)現(xiàn)有著相同危險(xiǎn)因素的早產(chǎn)兒ICH的發(fā)生存在明顯的個(gè)體差異，基因遺傳學(xué)背景是造成個(gè)體差異性的重要原因之一。前期研究發(fā)現(xiàn)血小板活化因子PAF-AH基因Val279Phe單核苷酸多態(tài)性與早產(chǎn)兒ICH具有相關(guān)性，我們?cè)诖嘶A(chǔ)上探索ICH發(fā)生的其他易感基因。 μ阿片受體(μ opioid receptor，OPRMI)A118G基因多態(tài)性與多種疾病的易感性及藥物反應(yīng)的差異性有關(guān)，而其與新生兒疾病的相關(guān)性研究目前尚未見報(bào)道。本研究擬選擇被大家廣泛認(rèn)可的與精神神經(jīng)因素密切相關(guān)的OPRMIA118G單核苷酸基因多態(tài)性，研究其與早產(chǎn)兒顱內(nèi)出血的關(guān)聯(lián)性，探討ICH發(fā)生的分子遺傳學(xué)機(jī)制，為臨床有效防治ICH提供參考依據(jù)。 材料和方法 1研究對(duì)象與分組 選取2011.7至2013.3鄭州大學(xué)第一附屬醫(yī)院新生兒重癥監(jiān)護(hù)室(NICU)住院的漢族早產(chǎn)兒作為研究對(duì)象。將顱內(nèi)出血早產(chǎn)兒視為顱內(nèi)出血組，同期因早產(chǎn)要求住院觀察的非顱內(nèi)出血早產(chǎn)兒視為非顱內(nèi)出血組。并對(duì)顱內(nèi)出血進(jìn)行分度。2基因多態(tài)性檢測(cè) 采用聚合酶鏈?zhǔn)椒磻?yīng)一限制性片段長(zhǎng)度多態(tài)性分析(Polymerase chainreaction restriction fragment length polymorphisms，PCR-RFLP)技術(shù)，對(duì)OPRMI基因Al18G多態(tài)性位點(diǎn)進(jìn)行檢測(cè)分析， PCR產(chǎn)物直接測(cè)序驗(yàn)證基因型檢測(cè)方法的可靠性。3統(tǒng)計(jì)學(xué)分析 采用SPSS19.0統(tǒng)計(jì)軟件進(jìn)行數(shù)據(jù)統(tǒng)計(jì)分析，兩組研究對(duì)象胎齡、出生體質(zhì)量之間的比較采用獨(dú)立樣本t檢驗(yàn)，性別構(gòu)成的比較檢驗(yàn)、Hard-Weinberg平衡法則檢驗(yàn)、各組基因型及等位基因分布的比較、性別及出血程度與顱內(nèi)出血的關(guān)聯(lián)性比較檢驗(yàn)均采用卡方檢驗(yàn)，檢驗(yàn)水準(zhǔn)α=0.05。 結(jié)果 1一般情況 顱內(nèi)出血組167例，其中男性99例，女性68例，平均胎齡（33.59±1.95）周，平均出生體質(zhì)量（1849±578）g；非顱內(nèi)出血組163例，男性91例，女性72例，平均胎齡（33.98±1.63）周，平均出生體質(zhì)量（1939±472）g。兩組性別、出生體質(zhì)量、胎齡之間差異無統(tǒng)計(jì)學(xué)意義，兩組具有可比性。 2兩組OPRMI基因A118G基因型、等位基因頻率 OPRMI基因A118G位點(diǎn)共有兩種等位基因：等位基因A、G，三種基因型：A/A型、A/G型和G/G型。 顱內(nèi)出血組：野生型純合子A/A，73例(43.7%)，突變雜合子A/G，82例(49.1%)，突變純合子G/G，12例(7.1%)；等位基因A、G頻率分別為68.3%、31.7%；非顱內(nèi)出血組：野生型純合子A/A，89例(54.6%)，突變雜合子A/G，，68例(41.7%)，突變純合子G/G，6例(3.7%)；等位基因A、G頻率分別為75.5%、24.5%；兩組基因型的分布沒有統(tǒng)計(jì)學(xué)意義(χ2=4.839，P=0.089)，但是比較顱內(nèi)出血組與非顱內(nèi)出血組野生型(A/A)和突變型(A/G+G/G)的陽性率，二者差異有統(tǒng)計(jì)學(xué)意義（χ2=3.913，P=0.048），兩組等位基因差異有統(tǒng)計(jì)學(xué)意義(χ2=4.222，P=0.04，OR=1.549，95%CI:1.003～2.391)，提示OPRMI基因118G等位基因的攜帶與ICH的發(fā)生呈正相關(guān),該突變可能會(huì)增加ICH的發(fā)病風(fēng)險(xiǎn)。顱內(nèi)出血組男、女性別OPRMI基因A118G多態(tài)位點(diǎn)基因型、等位基因比較，差異無統(tǒng)計(jì)學(xué)意義(χ2=0.300，P=0.58；χ2=0.843，P=0.358)；顱內(nèi)出血組不同出血程度OPRMI基因A118G多態(tài)位點(diǎn)基因型、等位基因比較，差異無統(tǒng)計(jì)學(xué)意義(χ2=2.418，P=0.342；χ2=0.160，P=0.689)。結(jié)論 1. OPRMI基因A118G的單核苷酸基因多態(tài)性與早產(chǎn)兒顱內(nèi)出血發(fā)生具有相關(guān)性。 2. OPRMI基因A118G多態(tài)性可能是ICH發(fā)病的一個(gè)潛在的易感位點(diǎn)，等位基因G的攜帶與ICH的發(fā)生呈正相關(guān)，該突變可能會(huì)增加ICH的發(fā)病風(fēng)險(xiǎn)。 3. OPRMI基因A118G單核苷酸基因多態(tài)性在顱內(nèi)出血發(fā)生中無性別差異性。 4. OPRMI基因A118G單核苷酸基因多態(tài)性對(duì)顱內(nèi)出血程度無影響。
[Abstract]:Background and purpose
Intracranial hemorrhage (ICH) is a kind of serious craniocerebral injury common in newborns. Preterm infants are more common, the younger the gestational age is, the higher the incidence is, the serious patients can have the sequelae of the nervous system and even death. In recent years, with the continuous development of medical technology, the survival rate of premature infants has been greatly improved, and the incidence of ICH has increased unabated.
The occurrence of ICH is related to its own anatomical and physiological characteristics and high risk factors for perinatal period. In addition, many inflammatory reactions and endogenous mediators can also be induced. The release of endogenous neuropeptides, especially endogenous opioid peptides, can aggravate this damage. In recent years, the research of neuropeptides and cerebrovascular disease mechanism has attracted more and more attention. It has been found that opioid peptide and opioid receptor system play an important role in the pathophysiological development of adult ischemic stroke.
Clinical studies have found that there are obvious individual differences in the occurrence of ICH in preterm infants with the same risk factors. The genetic background is one of the important reasons for the individual difference. The previous study found that the single nucleotide polymorphism of the platelet activating factor PAF-AH gene Val279Phe is related to the preterm infant ICH. Other susceptible genes of cord ICH.
The polymorphism of the micron opioid receptor (OPRMI) A118G gene is related to the susceptibility to a variety of diseases and the difference in the drug response. However, the study of the correlation with neonatal diseases has not yet been reported. This study is to choose the OPRMIA118G single nucleotide polymorphism, which is closely related to the mental and neurologic factors. Objective:To study the relationship between ICH and intracranial hemorrhage in premature infants and to explore the molecular genetic mechanism of ICH.
Materials and methods
1 research objects and groups
The preterm infants in the neonatal intensive care unit (NICU) of the First Affiliated Hospital of Zhengzhou University (NICU) were selected as the subjects. The intracranial hemorrhage preterm infants were treated as intracranial hemorrhage group. The non intracranial hemorrhage premature infants who were hospitalized for premature delivery were considered as non intracranial hemorrhage group, and the.2 gene polymorphism of intracranial hemorrhage was detected. Sex detection
The polymerase chain reaction restriction fragment length polymorphism analysis (Polymerase chainreaction restriction fragment length polymorphisms, PCR-RFLP) was used to detect the Al18G polymorphic loci of the OPRMI gene. The reliability.3 statistical analysis of the PCR product direct sequencing verification method of genotyping was carried out.
SPSS19.0 statistical software was used to carry out data statistics and analysis. Two groups of subjects were compared by independent sample t test, comparative test of sex composition, Hard-Weinberg balance test, comparison of genotype and allele distribution in each group, and the correlation of sex and bleeding degree with intracranial hemorrhage. Chi square test was used to test the level of alpha =0.05.
Result
1 general situation
167 cases of intracranial hemorrhage, including 99 male and 68 female, average fetal age (33.59 + 1.95) weeks, average birth weight (1849 + 578) g, non intracranial hemorrhage group 163 cases, male 91 cases, 72 cases, average gestational age (33.98 + 1.63) weeks, average birth weight (1939 + 68) g. group sex, birth body mass, birth body mass, there was no statistical difference between gestational age The group has comparability.
2 the A118G genotype and allele frequency of the two groups of OPRMI genes.
There are two alleles at the A118G locus of OPRMI gene: allele A, G, and three genotypes: A/A, A/G and G/G.
Intracerebral hemorrhage group: wild type homozygote A/A, 73 (43.7%), mutant heterozygote A/G, 82 cases (49.1%), mutant homozygote G/G, 12 cases (7.1%); allele A, G frequency 68.3%, 31.7%; non intracranial hemorrhage group: wild type homozygote A/A, 89 cases (54.6%), mutation heterozygote A/G, 68 (41.7%), mutant homozygote G/G, 6 case (3.7%); allele A, G The frequencies of the two groups were 75.5% and 24.5% respectively. The distribution of genotypes in two groups was not statistically significant (x 2=4.839, P=0.089), but the positive rates of wild type (A/A) and mutant type (A/G+G/G) in the group of intracranial hemorrhage and non intracranial hemorrhage group were statistically significant (x 2=3.913, P=0.048), and the difference between the two groups was statistically significant (x 2=4.222, P=0.04). OR=1.549,95%CI:1.003 ~ 2.391), suggesting that the 118G allele of the OPRMI gene is positively related to the occurrence of ICH. This mutation may increase the risk of ICH. The genotype of A118G polymorphic loci in the intracranial hemorrhage group, the A118G polymorphism of the female OPRMI gene, and the allele comparison of the allele, and the difference of the allele difference (x 2=0.300, P=0.58; Chi 2=0.843, P=0.358); There was no significant difference in allele genotype of OPRMI gene A118G polymorphic loci with different bleeding degree in internal hemorrhage group (x 2=2.418, P=0.342; X 2=0.160, P=0.689).
1. the single nucleotide polymorphism of OPRMI gene A118G is associated with intracranial hemorrhage in premature infants.
The 2. OPRMI gene A118G polymorphism may be a potential susceptible locus for the pathogenesis of ICH, and the carrying of the allele G is positively related to the occurrence of ICH, which may increase the risk of ICH.
3. the single nucleotide polymorphism of OPRMI A118G gene is not sex difference in intracranial hemorrhage.
4. the single nucleotide polymorphism of OPRMI A118G gene had no effect on the degree of intracranial hemorrhage.
【學(xué)位授予單位】：鄭州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R722.6

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