Hydrogen sulfide as a novel biomarker of asthma and chronic obstructive pulmonary disease

Hydrogen sulfide (H₂S) has recently been recognised as the third important gas-signalling molecule, besides nitric oxide and carbon monoxide. H₂S has been reported to be produced by many cell types in mammalian tissues and organs throughout the actions of H₂S-generating enzymes or redox reactions between the oxidation of glucose and element of sulfur. Although the pathological role of H₂S has not yet been fully elucidated, accumulative data suggest that H₂S may have biphasic effects. Briefly, it mainly has anti-inflammatory and antioxidant roles, although it can also have pro-inflammatory effects under certain conditions where rapid release of H₂S in tissues occur, such as sepsis. To date, there have been several clinical studies published on H₂S in respiratory disorders, including asthma and chronic obstructive pulmonary disease (COPD). According to previous studies, H₂S is detectable in serum, sputum, and exhaled breath, although a gold standard method for detection has not yet been established. In asthma and COPD, H₂S levels in serum and sputum can vary depending on the underlying conditions such as an acute exacerbation. Furthermore, sputum H₂S in particular correlates with sputum neutrophils and the degree of airflow limitation, indicating that H₂S has potential as a novel promising biomarker for neutrophilic airway inflammation for predicting current control state as well as future risks of asthma. In the future, concurrent measures of H₂S with conventional inflammatory biomarkers (fractional exhaled nitric oxide, eosinophils etc) may provide more useful information regarding the identification of inflammatory phenotypes of asthma and COPD for personalised treatment.     

单纯收缩期高血压患者同型半胱氨酸代谢关键酶基因多态性相关因子研究

目的　研究同型半胱氨酸代谢关键酶亚甲基 4 氢叶酸还原酶 (MTHFR)及胱硫醚 β 合成酶 (CBS)基因多态性及其代谢相关因子在收缩期高血压 (ISH)发病中的作用 ,寻找ISH的易感基因及中间表型。方法　从天津高血压防治区选取ISH患者 5 5例 ,对照组 46例 ,应用分子生物学方法对其进行MTHFRC6 77,CBSG919,T833,C341位点基因多态性检测 ,并同时测定血中同型半胱氨酸(Hcy)、叶酸、维生素B6 、维生素B1 2 浓度 ,并对结果进行相关性分析。结果　ISH组中MTHFR及CBS基因构成与对照组不同存在统计学差异 (P <0 0 5 )。ISH组中MTHFRT等位基因为 6 9 1% ,其发生率高于对照组 (P <0 0 1) ;CBS阳性等位基因发生率为 15 2 % ,与对照组不同。ISH组Hcy水平 [( 17 70± 8 5 0 ) μmol L]高于对照组 [( 10 6 5± 3 10 ) μmol L](P <0 0 5 )。而叶酸、维生素B6 、维生素B1 2 浓度与对照组相比明显降低 (P <0 0 5 )。ISH组中高同型半胱氨酸血症的发生率高于对照组 ,具有统计学差异 (P <0 0 5 ) ,ISH组及对照组三种基因型血浆Hcy水平的改变存在着相同的趋势 ,即突变型高于杂合型与野生型。但血压升高的水平与Hcy浓度之间并无同向性变化。Hcy水平的增高可以增加ISH的发病 ,叶酸在ISH的发病中具有负性作用。结论　Hcy代谢     

杏仁核脑区硫化氢对创伤后应激障碍模型大鼠抑郁样行为的影响

目的:探讨创伤后应激障碍(posttraumatic stress disorder,PTSD)大鼠抑郁样行为改变及杏仁核内胱硫醚β-合成酶(cystathionineβ-synthase,CBS)/硫化氢(hydrogen sulfide,H_2S)水平的变化,并研究补充H_2S对大鼠抑郁样行为的作用及机制。方法:应用连续单一应激方案制备PTSD动物模型,用强迫游泳实验和糖水偏好实验检测大鼠抑郁样行为,用Western blot技术和亚甲基蓝法检测杏仁核CBS/H_2S体系含量,用整体细胞外电生理方法记录杏仁核神经元放电情况。结果:与正常对照组比较,PTSD组大鼠强迫游泳不动时间显著延长(P0.01),糖水偏好率显著降低(P0.01);杏仁核脑区的CBS/H_2S体系含量显著降低(P0.01)。与PTSD组比较,PTSD+硫氢化钠组大鼠强迫游泳不动时间显著减少(P0.01),糖水偏好率显著升高(P0.01)。微量压力注射L-半胱氨酸使杏仁核神经元自发放电频率显著升高(P0.01)。结论:PTSD大鼠通过抑制杏仁核CBS/H_2S体系导致抑郁样行为加重。H_2S拮抗PTSD大鼠抑郁样行为的机制可能与其增加杏仁核神经元放电频率和兴奋性有关。     

血浆Hcy及其代谢酶基因多态性与Alzheimer病的关系

目的探讨血浆同型半胱氨酸(homocysteine,Hcy)及其相关代谢酶N5,N10-亚甲基四氢叶酸还原酶(N5,N10-methylenetetrahydrofolatereductase,MTHFR)胱硫醚β-合成酶(cystathionineβ-synthase,CBS)基因多态性与阿尔茨海默病(Alzheimersdisease,AD)的关系。方法对104例AD患者、100例健康对照组运用高压毛细管电泳紫外检测法测定血浆Hcy水平,采用多聚酶链反应-限制性片段长度多态性(PCR-RELP)方法检测MTHFR基因第4外显子C677T、CBS基因第8外显子844ins68的多态性。结果AD患者血浆Hcy(16.18±3.35)μmol/L非常显著高于健康对照者Hcy(12.24±3.96)μmol/L。AD组和对照组受试者间MTHFRC677T和CBS基因844ins68的各基因型分布和等位基因频率均无显著性差异,仅MTHFR的T等位基因与受试者血浆Hcy水平升高有关。结论AD患者的血浆Hcy水平高于健康人群,可能与疾病的发生发展有关;并可能主要由环境因素所致,而非MTHFR和CBS基因变异引起。补充VitB12和叶酸有助于AD的治疗。     

Urinary homocystine levels in a newborn infant with cystathionine synthase deficiency

A boy with homocystinuria due to cystathionine synthase deficiency was found to have hypermethioninaemia by neonatal blood screening, but was not diagnosed as homocystinuric until 3 months of age because urinary homocystine was not detected by the cyanidenitroprusside test or on two examinations with a sensitive amino acid autoanalyser. These findings indicate that tests for urinary homocystine should be made repeatedly with an amino acid auto-analyser in newborn infants with hypermethioninaemia until the enzyme defect is identified.     

全脑缺血-再灌注大鼠脑组织内源性硫化氢的动态变化

目的探讨大鼠全脑缺血-再灌注过程中,不同时点内源性硫化氢（H2S）含量、胱硫醚β合酶（cystathionine beta synthase,CBS）活性及其mRNA表达的变化。方法取雄性SD大鼠56只,随机分为正常组、假手术组和脑缺血-再灌注（cerebral ischemia-reperfusion;I/R）12、24、48、72h及7d组,每组8只大鼠。应用四血管阻断法制作大鼠全脑I/R模型。采用全自动酶标仪（Bio-TEK ELx800,美国）测定各组大鼠双侧前脑皮质中H2S含量、CBS活性;用逆转录聚合酶链反应（RT-PCR）检测CBS mRNA表达的变化。结果正常组和假手术组大鼠前脑皮质组织H2S含量分别为（12.5±1.3）和（11.7±1.4）nmol/g,CBS酶活性分别为（44.5±2.3）和（44.1±2.8）nmol·g^-1·h^-1,CBS mRNA灰度值为3.62±0.23和3.94±0.40,两组比较差异无统计学意义（P〉0.05）。I/R12h组H2S含量为（18.3±1.2）nmol/g,CBS酶活性为（66.7±3.1）nmol·g^-1·h^-1,CBS mRNA灰度值为7.20±1.25;与假手术组比较均增高,差异均有统计学意义（P〈0.01）。I/R24h组以上指标为（8.3±1）nmol/g、（30.8±3.5）nmol·g^-1·h^-1及0.90±0.16,与假手术组比较均降低,差异均有统计学意义（P〈0.01）。I/R48、72h组以上指标逐渐恢复正常水平,与假手术组比较差异无统计学意义（P〉0.05）。I/R7d组CBS mRNA灰度值为5.19±0.01,与假手术组比较差异有统计学意义（P〈0.01）,其余指标差异均无统计学意义。结论内源性H2S在全脑缺血-再灌注的发生、发展过程中呈动态变化,并可能在缺血-再灌注早期发挥作用。     

蛋氨酸合成酶及胱硫醚β合成酶基因多态性与冠状动脉粥样硬化发生的关系

目的:探讨蛋氨酸合成酶(MS)和胱硫醚β合成酶(CBS)的基因多态性对同型半胱氨酸代谢的影响及与动脉粥样硬化的关系。方法:共入选研究对象153例(对照组40例,冠心病组113例)。采用高效液相色谱法测定血清同型半胱氨酸浓度。采用聚合酶链式反应扩增及限制性内切酶技术检测CBS基因1011A→G突变型和MS基因D919G突变型。结果:在40例对照组中,MS基因纯合子突变型(GG)为0,杂合子突变型(AG)占7.5%(3/40),总突变率占7.5%;在113例冠心病组中,纯合子突变型(GG)占2.7%(3/113),杂合子突变型(AG)占21.2%(24/113),总突变率为23.9%。χ2检验表明,突变型和非突变型在两组间有显著性差异(χ2=5.16,P<0.025)。G等位基因对冠心病的比值比(OR值)为4.39(95%的可信限为1.31~14.73,P<0.05)。突变部分血清同型半胱氨酸水平显著高于非突变部分[(16.83±3.08)mmol/Lvs(9.86±3.18)mmol/L]。CBS基因仅扩增出等位基因A,没有扩增出突变型G等位基因。结论:MS基因D919G突变型可导致血清同型半胱氨酸水平升高,并促进动脉粥样硬化的发生。     

Expression study of mutant cystathionine beta-synthase found in Japanese patients with homocystinuria

Cystathionine beta-synthase (CBS) deficiency is the most common cause of homocystinuria. More than 130 pathogenic mutations, mostly in the Caucasian populations, have been described. Recently, our group reported a mutation analysis of Japanese homocystinuric patients. In the present paper, we report an expression study of several mutant CBS enzymes in Escherichia coli, i.e., R121H, G148R, G151R, S217F, H232D, R266G, 1591delTTCG, and K441X. All of the mutants except K441X exhibited severely decreased activity, and the capability to form tetramers of most mutants was severely impaired. The K441X mutant, on the other hand, exhibited relatively high activity (63% of the wild type activity). This was probably due to two factors. First, the high abundance of the full-length CBS protein, a likely K441Q mutant, which was produced through suppression of the amber termination codon by glutamine tRNA in E. coli. And second, the presence of a C-terminally truncated protein, which was previously shown to be constitutively activated. Patient-derived lymphocytes, however, showed no detectable CBS subunits. As previously hypothesized, the increased aggregation of mutant CBS subunits might be a common pathogenic mechanism in CBS deficiency.     

Cerebral edema associated with betaine treatment in classical homocystinuria

A child with cystathionine beta-synthase deficiency developed cerebral edema 4 to 6 weeks after starting betaine therapy. There was no evidence of intracranial thrombosis, but there was widespread edema of the white matter. He recovered fully after emergency decompressive craniotomy and withdrawal of betaine.