中国人21-羟化酶缺乏症基因型和临床表型特点研究

利用快速可靠的基因突变检测方法研究中国人21-羟化酶缺乏症(21-hydroxylase deficiency ,21-OHD)基因突变特点及基因型和临床表型的关系.用8例非经典型患者、35例经典型患者及34例正常对照者基因组DNA作模板,用特定引物特异性扩增CYP21的两个片段,片段1从Exon1→Exon3,片段2从Exon3→Exon10.用片段1和片段2为模板进行第二轮PCR,用特定限制性内切酶消化后经琼脂糖凝胶电泳鉴定9种突变,包括Del、Exon3 Del8 bp、Q318X、 R356W、Exon6Cluster、i2g、I172N、P30L和 V281L.结果表明,43例患者的86个等位基因中的79个(91.9%)检测出突变,最常见的是I172N(36.0%),其次为i2g(20.9%)、Del(8.6%)、P30L(7.0%)、Q318X(7.0%)、V281L(4.7%)、R356W(2.3%)、E6Cluster(2.3%)和Exon3 Del8 bp(1.2%).失盐型、单纯男性化型和非经典型21-羟化酶缺乏症各临床分型中最常见的突变分别是 Del(44.4%)、I172N(44.2%)和 P30L(37.5%).另外根据对21-羟化酶活性影响程度将基因型分为轻、中、重3组,3组间初诊年龄、17-羟孕酮(17-OHP)、该病亚型构成均存在显著差异(P<0.05),提示基因型决定临床表型.研究结果表明,中国人21-OHD最常见的突变为I172N、i2g和Del,中国人21-OHD基因型和临床表型密切相关.     

None of the Six SNPs of IL28B Could Predict Treatment Responses in Genotype 2 Chronic HCV Infected Patients by Propensity Score Matching Analysis

Background & Aims

A combination of pegylated interferon-alpha and ribavirin (PR) is the standard therapy for patients with chronic hepatitis C. The impact of polymorphism of interleukin-28B (IL28B) on sustained virological response (SVR) to PR has been well documented in patients with CHC genotype-1 (GT1), but it is controversial in genotype-2 (GT2) CHC patients. This study investigated the predictability of six single nucleotide polymorphisms (SNP) of IL28B on the treatment responses of PR in patients with CHC GT2.

Method

197 CHC GT2 consecutive patients who received PR treatment in our prospective cohort were enrolled. Hepatitis C virus (HCV) genotyping, quantification of HCV-RNA and genotyping of the ten SNPs of IL28B were performed. Six SNPs of IL28B were chosen for analysis. The propensity score matching (PSM) analysis was applied using patients with CHC GT1 in another prospective cohort as a positive comparison to avoid covariate bias.

Results

The distribution of the six SNPs was similar in GT1 and GT2 patients. Five of these SNPs had strong association with treatment responses in GT1 but not in GT2 patients. After PSM analysis, these five SNPs still showed strong association with rapid virological response (RVR), cEVR and SVR in GT1 and had no influence in GT2 patients. Furthermore, rs12979860 and baseline viral load were the predictors for both RVR and SVR in GT1 patients. However, only baseline viral load could predict RVR and SVR in GT2 patients. In addition, in patients without RVR, rs12979860 was the only predictor for SVR in GT1 but no predictor for SVR was found in GT2.

Conclusions

The genetic polymorphisms of IL28B had no impact on treatment responses in GT2 patients.     

Phenotype and Genotype Analysis of Chinese Patients with Osteogenesis Imperfecta Type V

Osteogenesis imperfecta (OI) type V is an autosomal-dominant disease characterized by calcification of the forearm interosseous membrane, radial head dislocation, a subphyseal metaphyseal radiodense line, and hyperplastic callus formation. The causative mutation, c.-14C>T in the 5''-untranslated region of IFITM5, was recently discovered to be involved in this disease. However, in spite of the little genotypic variability, considerable phenotypic variability has been recognized in two cohorts of patients, the majority of whom were Caucasians. Using exome sequencing, we identified the same heterozygous mutation in four Chinese families with OI type V. This study confirms the molecular cause of OI type V and describes the phenotype of Chinese patients with this disorder. In conclusion, the phenotype of Chinese patients was generally similar to that of Caucasian patients.     

Hypoxanthine-Guanine Phosphoribosyltransferase Deficiency: Biochemical and Molecular Findings in Six Argentine Patients

Hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficiency is an inborn error of purine metabolism responsible for Lesch-Nyhan Disease (LND) and its partial phenotypes, HPRT-related hyperuricemia with neurologic dysfunction (HRND) and hyperuricemia alone. We report here the recognition of six Argentine patients, two with LND and four with HRND. All patients presented elevated excretion of uric acid, hypoxanthine, and xanthine and decreased HPRT enzyme activities <1 nmol/h/mg Hb. The molecular analysis demonstrated in the two LND patients a novel inherited transition mutation, c.203T >C (L68P), in one subject and a germline transition mutation, c.209G >A (G70E), in the other. In the HRND patients a novel transversion mutation, c.584 A >C (Y195S), was found in three related patients and an inherited transition mutation, c.143G >A (R48H), in the fourth subject.     

Interleukin-12B rs3212227 polymorphism and cancer risk: a meta-analysis

IL-12 plays a very important role in the development and progress of cancer. IL-12B rs3212227 polymorphism has been reported and many studies have focused on the role of this polymorphism in various cancers. However, the association between IL-12B rs3212227 polymorphism and cancer risk remains controversial. Therefore, we performed a systematic meta-analysis to estimate the overall cancer risk associated with this gene polymorphism and to quantify any potential between-study heterogeneity. PubMed and Embase databases were searched for case–control studies published up to April 1, 2012 that investigated IL-12B rs3212227 polymorphism and cancer risk. Odds ratios (OR) with 95?% confidence intervals (95?% CI) were used to access the strength of this association. Heterogeneity among articles and publication bias were also verified. Ten studies with 2,954 cancer patients and 3,276 healthy controls were included. This meta-analysis showed that there was a significant association between IL-12B rs3212227 polymorphism and overall cancer risk (CC/AC vs AA: OR?=?1.32, 95?% CI?=?1.06–1.63). When stratified by cancer type, we found a significant increased risk in cervical and nasopharyngeal cancer (OR?=?1.34, 95?% CI?=?1.04–1.73; OR?=?2.03, 95?% CI?=?1.57–2.63, respectively). In the stratified analysis, we also observed a similar association in population-based studies (OR?=?1.34, 95?% CI?=?1.00–1.80), Asian populations (OR?=?1.33, 95 % CI?=?1.06–1.67) and European populations (OR?=?1.54, 95 % CI?=?1.04–2.28). According to the results of our meta-analysis, IL-12B rs3212227 polymorphism probably is associated with a high risk of cancer.     

VITAMIN B12 AND THE MACROMOLECULAR COMPOSITION OF EUGLENA : II. Recovery from Unbalanced Growth Induced by Vitamin B12 Deficiency

When vitamin B₁₂ is added to B₁₂-deficient cultures of Euglena gracilis, the cells undergo two relatively synchronous cell divisions within a shorter than usual period of time, apparently as a result of a transitory shortening of the cell cycle. The first cell division pulse, occurring 4.5 h after addition of B₁₂, is preceded by the completion of DNA duplication, but appears to involve no net synthesis of RNA or protein. Before the second round of cell division at about 11 h, a significant amount of DNA synthesis is observed. This time it is accompanied by a minor increase in the RNA and protein content of the culture. The cellular contents of RNA and protein were observed to decrease steadily after the resumption of cell division in B₁₂-depleted cultures receiving the vitamin. Ultimately all three macromolecules returned to their nondeficient, plateau stage levels; by this time, cell division had ceased.     

肠易激综合征患者血清IL-10、IL-12及皮质醇变化及其意义

目的:探讨肠易激综合征(IBS)患者血清白细胞介素10(IL-10)白细胞介素12(IL-12)和皮质醇(CO)浓度变化及它们在IBS中的可能作用和临床意义.方法:随机抽取来源于门诊和住院IBS患者和健康志愿者血清,采用放射免疫法测量血清IL-10、IL-12和CO浓度.结果:腹泻型IBS病人血清IL-10和CO含量显著高于正常(P＜0.05),便秘型IBS病人血清内IL-10和CO水平也高于正常对照(P＜0.05).腹泻型和便秘型IBS病人血清内IL-10和CO水平之间差别无统计学意义(P＞0.05).腹泻型IBS组血清IL-12含量比正常组低(P＜0.05),便秘型IBS组血清内IL-12水平也低于正常对照组(P＜0.05).腹泻型和便秘型组之间血清内IL-12水平差别无统计学意义(P＞0.05).结论:炎症因子和应激可能参与IBS发病.     

Interleukin 12B (IL12B) genetic variation and pulmonary tuberculosis: a study of cohorts from The Gambia, Guinea-Bissau, United States and Argentina

We examined whether polymorphisms in interleukin-12B (IL12B) associate with susceptibility to pulmonary tuberculosis (PTB) in two West African populations (from The Gambia and Guinea-Bissau) and in two independent populations from North and South America. Nine polymorphisms (seven SNPs, one insertion/deletion, one microsatellite) were analyzed in 321 PTB cases and 346 controls from Guinea-Bissau and 280 PTB cases and 286 controls from The Gambia. For replication we studied 281 case and 179 control African-American samples and 221 cases and 144 controls of European ancestry from the US and Argentina. First-stage single locus analyses revealed signals of association at IL12B 3′ UTR SNP rs3212227 (unadjusted allelic p = 0.04; additive genotypic p = 0.05, OR = 0.78, 95% CI [0.61–0.99]) in Guinea-Bissau and rs11574790 (unadjusted allelic p = 0.05; additive genotypic p = 0.05, OR = 0.76, 95% CI [0.58–1.00]) in The Gambia. Association of rs3212227 was then replicated in African-Americans (rs3212227 allelic p = 0.002; additive genotypic p = 0.05, OR = 0.78, 95% CI [0.61–1.00]); most importantly, in the African-American cohort, multiple significant signals of association (seven of the nine polymorphisms tested) were detected throughout the gene. These data suggest that genetic variation in IL12B, a highly relevant candidate gene, is a risk factor for PTB in populations of African ancestry, although further studies will be required to confirm this association and identify the precise mechanism underlying it.     

Genotype and Phenotype of Echinococcus granulosus Derived from Wild Sheep (Ovis orientalis) in Iran

The aim of the present study is to determine the characteristics of genotype and phenotype of Echinococcus granulosus derived from wild sheep and to compare them with the strains of E. granulosus sensu stricto (sheep-dog) and E. granulosus camel strain (camel-dog) in Iran. In Khojir National Park, near Tehran, Iran, a fertile hydatid cyst was recently found in the liver of a dead wild sheep (Ovis orientalis). The number of protoscolices (n=6,000) proved enough for an experimental infection in a dog. The characteristics of large and small hooks of metacestode were statistically determined as the sensu stricto strain but not the camel strain (P=0.5). To determine E. granulosus genotype, 20 adult worms of this type were collected from the infected dog. The second internal transcribed spacer (ITS2) of the nuclear ribosomal DNA (rDNA) and cytochrome c oxidase 1 subunit (COX1) of the mitochondrial DNA were amplified from individual adult worm by PCR. Subsequently, the PCR product was sequenced by Sanger method. The lengths of ITS2 and COX1 sequences were 378 and 857 bp, respectively, for all the sequenced samples. The amplified DNA sequences from both ribosomal and mitochondrial genes were highly similar (99% and 98%, respectively) to that of the ovine strain in the GenBank database. The results of the present study indicate that the morpho-molecular features and characteristics of E. granulosus in the Iranian wild sheep are the same as those of the sheep-dog E. granulosus sensu stricto strain.     

KIR,HLA, and IL28B Variant Predict Response to Antiviral Therapy in Genotype 1 Chronic Hepatitis C Patients in Japan

Natural killer cell responses play a crucial role in virus clearance by the innate immune system. Although the killer immunoglobulin-like receptor (KIR) in combination with its cognate human leukocyte antigen (HLA) ligand, especially KIR2DL3-HLA-C1, is associated with both treatment-induced and spontaneous clearance of hepatitis C virus (HCV) infection in Caucasians, these innate immunity genes have not been fully clarified in Japanese patients. We therefore investigated 16 KIR genotypes along with HLA-B and -C ligands and a genetic variant of interleukin (IL) 28B (rs8099917) in 115 chronic hepatitis C genotype 1 patients who underwent pegylated-interferon-α2b (PEG-IFN) and ribavirin therapy. HLA-Bw4 was significantly associated with a sustained virological response (SVR) to treatment (P = 0.017; odds ratio [OR] = 2.50, ), as was the centromeric A/A haplotype of KIR (P = 0.015; OR 3.37). In contrast, SVR rates were significantly decreased in patients with KIR2DL2 or KIR2DS2 (P = 0.015; OR = 0.30, and P = 0.025; OR = 0.32, respectively). Multivariate logistic regression analysis subsequently identified the IL28B TT genotype (P = 0.00009; OR = 6.87, 95% confidence interval [CI] = 2.62 - 18.01), KIR2DL2/HLA-C1 (P = 0.014; OR = 0.24, 95% CI = 0.08 - 0.75), KIR3DL1/HLA-Bw4 (P = 0.008, OR = 3.32, 95% CI = 1.37 - 8.05), and white blood cell count at baseline (P = 0.009; OR = 3.32, 95% CI = 1.35 - 8.16) as independent predictive factors of an SVR. We observed a significant association between the combination of IL28B TT genotype and KIR3DL1-HLA-Bw4 in responders (P = 0.0019), whereas IL28B TT along with KIR2DL2-HLA-C1 was related to a non-response (P = 0.0067). In conclusion, combinations of KIR3DL1/HLA-Bw4, KIR2DL2/HLA-C1, and a genetic variant of the IL28B gene are predictive of the response to PEG-IFN and ribavirin therapy in Japanese patients infected with genotype 1b HCV.     

Role of IL1A rs1800587, IL1B rs1143627 and IL1RN rs2234677 Genotype Regarding Development of Chronic Lumbar Radicular Pain; a Prospective One-Year Study

Previous studies indicate that lumbar radicular pain following disc herniation may be associated with release of several pro-inflammatory mediators, including interleukin-1 (IL1). In the present study, we examined how genetic variability in IL1A (rs1800587 C>T), IL1B (rs1143627 T>C) and IL1RN (rs2234677 G>A) influenced the clinical outcome the first year after disc herniation. Patients (n = 258) with lumbar radicular pain due to disc herniation were recruited from two hospitals in Norway. Pain and disability were measured by visual analogue scale (VAS) and Oswestry Disability Index (ODI) over a 12 month period. The result showed that patients with the IL1A T allele, in combination with the IL1RN A allele had more pain and a slower recovery than other patients (VAS p = 0.049, ODI p = 0.059 rmANOVA; VAS p = 0.003, ODI p = 0.050 one-way ANOVA at 12 months). However, regarding the IL1B/IL1RN genotype, no clear effect on recovery was observed (VAS p = 0.175, ODI p = 0.055 rmANOVA; VAS p = 0.105, ODI p = 0.214 one-way ANOVA at 12 months). The data suggest that the IL1A T/IL1RN A genotype, but not the IL1B T/IL1RN A genotype, may increase the risk of a chronic outcome in patients following disc herniation.     

A large-scale genetic association study confirms IL12B and leads to the identification of IL23R as psoriasis-risk genes

We performed a multitiered, case-control association study of psoriasis in three independent sample sets of white North American individuals (1,446 cases and 1,432 controls) with 25,215 genecentric single-nucleotide polymorphisms (SNPs) and found a highly significant association with an IL12B 3'-untranslated-region SNP (rs3212227), confirming the results of a small Japanese study. This SNP was significant in all three sample sets (odds ratio [OR](common) 0.64, combined P [Pcomb]=7.85x10(-10)). A Monte Carlo simulation to address multiple testing suggests that this association is not a type I error. The coding regions of IL12B were resequenced in 96 individuals with psoriasis, and 30 additional IL12B-region SNPs were genotyped. Haplotypes were estimated, and genotype-conditioned analyses identified a second risk allele (rs6887695) located approximately 60 kb upstream of the IL12B coding region that exhibited association with psoriasis after adjustment for rs3212227. Together, these two SNPs mark a common IL12B risk haplotype (OR(common) 1.40, Pcomb=8.11x10(-9)) and a less frequent protective haplotype (OR(common) 0.58, Pcomb=5.65x10(-12)), which were statistically significant in all three studies. Since IL12B encodes the common IL-12p40 subunit of IL-12 and IL-23, we individually genotyped 17 SNPs in the genes encoding the other chains of these cytokines (IL12A and IL23A) and their receptors (IL12RB1, IL12RB2, and IL23R). Haplotype analyses identified two IL23R missense SNPs that together mark a common psoriasis-associated haplotype in all three studies (OR(common) 1.44, Pcomb=3.13x10(-6)). Individuals homozygous for both the IL12B and the IL23R predisposing haplotypes have an increased risk of disease (OR(common) 1.66, Pcomb=1.33x10(-8)). These data, and the previous observation that administration of an antibody specific for the IL-12p40 subunit to patients with psoriasis is highly efficacious, suggest that these genes play a fundamental role in psoriasis pathogenesis.     

Association of Interleukin-6, Interleukin-12, and Interleukin-10 Gene Polymorphisms with Essential Hypertension in Tatars from Russia

Essential hypertension is a common disease with fatal clinical complications. Epidemiological and family studies have confirmed the role of genetic predisposition in its development. Hypertensive patients have been shown to have an altered profile of pro- and anti-inflammatory cytokines. The aim of our investigation was to reveal the association of interleukin-6, interleukin-12, and interleukin-10 gene polymorphisms with essential hypertension and its clinical complications in a Tatar ethnic group from Bashkortostan, Russia. The study involved 362 hypertensive patients and 244 healthy subjects from this Tatar ethnic group (Bashkortostan, Russia). DNA was isolated from whole venous blood using phenol–chloroform extraction by the standard method. IL6 −572 G/C, IL12B 1159 C/A, and IL10 –627 C/A genotypes were typed using polymerase chain reaction followed by restriction enzyme digestion. We found that the IL10 −627 *C/*C genotype was associated with decreased risk of hypertension (OR = 0.64, P = 0.035). IL6 genotypes and allele distribution did not differ significantly between subjects with and without hypertension, but the IL6 −572 *G/*G genotype frequency was found to be significantly higher among those patients who had stroke, compared with normotensive control subjects (P = 0.036). Carriers of the IL12B 1159 *A/*A genotype had a lower risk of stroke (OR = 0.38, P = 0.028). Our study has shown the association between IL10 −627 C/A polymorphism and essential hypertension in the Tatar ethnic group from Bashkortostan, Russia. The IL10 −627*C/*C genotype was found to be protective against hypertension. We also demonstrated that hypertensive patients with the IL12B *A/*A and IL6 *G/*G genotypes had increased risk of stroke. Our results suggest a role for cytokines in cardiovascular disease development in the Tatar ethnic group, but further investigation is needed.     

Chronic Proliferative Dermatitis in Sharpin Null Mice: Development of an Autoinflammatory Disease in the Absence of B and T Lymphocytes and IL4/IL13 Signaling

SHARPIN is a key regulator of NFKB and integrin signaling. Mice lacking Sharpin develop a phenotype known as chronic proliferative dermatitis (CPDM), typified by progressive epidermal hyperplasia, apoptosis of keratinocytes, cutaneous and systemic eosinophilic inflammation, and hypoplasia of secondary lymphoid organs. Rag1^−/− mice, which lack mature B and T cells, were crossed with Sharpin^−/− mice to examine the role of lymphocytes in CDPM. Although inflammation in the lungs, liver, and joints was reduced in these double mutant mice, dermatitis was not reduced in the absence of functional lymphocytes, suggesting that lymphocytes are not primary drivers of the inflammation in the skin. Type 2 cytokine expression is increased in CPDM. In an attempt to reduce this aspect of the phenotype, Il4ra^−/− mice, unresponsive to both IL4 and IL13, were crossed with Sharpin^−/− mice. Double homozygous Sharpin^−/−, Il4ra^−/− mice developed an exacerbated granulocytic dermatitis, acute system inflammation, as well as hepatic necrosis and mineralization. High expression of CHI3L4, normally seen in CPDM skin, was abolished in Sharpin^−/−, Il4ra^−/− double mutant mice indicating the crucial role of IL4 and IL13 in the expression of this protein. Cutaneous eosinophilia persisted in Sharpin^−/−, Il4ra^−/− mice, although expression of Il5 mRNA was reduced and the expression of Ccl11 and Ccl24 was completely abolished. TSLP and IL33 were both increased in the skin of Sharpin^−/− mice and this was maintained in Sharpin^−/−, Il4ra^−/− mice suggesting a role for TSLP and IL33 in the eosinophilic dermatitis in SHARPIN-deficient mice. These studies indicate that cutaneous inflammation in SHARPIN-deficient mice is autoinflammatory in nature developing independently of B and T lymphocytes, while the systemic inflammation seen in CPDM has a strong lymphocyte-dependent component. Both the cutaneous and systemic inflammation is enhanced by loss of IL4 and IL13 signaling indicating that these cytokines normally play an anti-inflammatory role in SHARPIN-deficient mice.     

Difference of Phenotype and Genotype Between Human and Environmental: Isolated Vibrio cholerae in Surabaya,Indonesia

Cholera due to Vibrio cholerae has been spreading worldwide, although the reports focusing on Indonesian V. cholerae are few. In this study, in order to investigate how V. cholerae transmitted to human from environment. We extended an epidemiological report that had investigated the genotype of V. cholerae isolated from human pediatric samples and environmental samples. We examined 44 strains of V. cholerae isolated from pediatric diarrhea patients and the environment such as shrimps or oysters collected in three adjacent towns in Surabaya, Indonesia. Susceptibilities were examined for 11 antibiotics. Serotype O1 or O139 genes and pathogenic genes including cholera toxin were detected. Multi-locus sequence typing (MLST) and enterobacterial repetitive intergenic consensus (ERIC)-PCR were also performed to determine genetic diversity of those isolates. Serotype O1 was seen in 17 strains (38.6%) with all pathogenic genes among 44 isolates. Other isolates were non-O1/non-O139 V. cholerae. Regarding antibiotic susceptibilities, those isolates from environmental samples showed resistance to ampicillin (11.4%), streptomycin (9.1%) and nalidixic acid (2.3%) but those isolates from pediatric stools showed no resistance to those 3 kinds of antibiotics. MLST revealed sequence type (ST) 69 in 17 strains (38.6%), ST198 in 3 strains (6.8%) and non-types in 24 strains (54.5%). All the ST69 strains were classified to O1 type with more than 95% similarity by ERIC-PCR, including all 6 (13.6%) isolates from environmental samples with resistance to streptomycin. In conclusion, V. cholerae O1 ST69 strains has been clonally spreading in Surabaya, exhibiting pathogenic factors and antibiotic resistance to streptomycin, especially in the isolates from environment.