In vivo alterations in lactate dehydrogenase (LDH) and LDH isoenzymes patterns by acute carbofuran intoxication

Intoxication with an acute dose of carbofuran (1.5 mg/kg, sc) in male Sprague-Dawley rats evoked severe toxic manifestations characteristic of hypercholinergic preponderance with profound muscle fasciculations and convulsions during 30–60 min, lasting for about 2 h. Lactate dehydrogenase, a biomarker cytoplasmic enzyme catalyzing the reversible reaction of lactate-pyruvate, was represented by five electrophoretically distinct isoenzymes in the serum and tissues. The amounts of each isoenzyme in different tissues were widely varied and consequently the patterns were tissue specific. A 24-h time-course following carbofuran administration indicated a two-fold increase in the activity of total LDH in serum and more than 30% in hemidiaphragm and liver. The patterns of LDH isoenzymes in serum revealed a significant (P < 0.01) increase in all the isoenzymes except LDH-4 (64% decrease). Analysis of each tissue revealed characteristic changes in LDH isoenzyme patterns indicating organ-specific tissue damage. These alerations in LDH and its isoenzymes, in addition to acetylcholinesterase inhibition, may be directly or indirectly related to the mechanism(s) of the toxic action, and also provide insight into the site/organ(s) of toxic injury, thus providing an early prognostic indicator.Presented in part at the 74th Annual Meeting of the Federation of American Societies for Experimental Biology, April 1–5, 1990 Washington, DC     

Role of lactate dehydrogenase (LDH) level in the aqueous humor in the diagnosis of retinoblastoma (RB)

In spite of the progress of the imagery in ophthalmology, the retinoblastoma remains in some particular cases, difficult to make. Moreover, the therapy sanction is often heavy. The lactate dehydrogenase (LDH) dosage in aqueous humor is an invasive technique conceivable when there are diagnosis problems particularly in retino-blastoma of old children and in affections simulating retino-blastoma. The purpose of this work is to lay out three observations where the LDH dosage enabled to keep the retinoblastoma diagnosis in one 8-year old child and eliminate it in two children having pseudoglioma with spontaneous evolution allowing to confirm the non-tumoral nature of the affection. Thus, the technique of LDH dosage has been reliable for the 3 cases which we have introduced. The authors compare the reliability of the different cytochemical techniques during the retinoblastoma diagnosis.     

乳酸脱氢酶同功酶活性测定应用研究

我国卫生部１９８５年已将小鼠精子畸形试验作为可选择的致突变试验方法，但是精子发生是一个多时相的复杂过程，采样时错过了精子发育的敏感阶段，或因感染，天气炎热体温变化，致使实验得不到预期结果。为了探讨血浆中ＬＤＨ－Ｘ酶活性测定与精子畸形率的关系设计了本实验研究。三种阳性物（ＭＭＣ、ＣＰ、Ｃｄ）均对睾丸细胞产生明显的损伤作用，睾丸匀浆及血浆中ＬＤＨ－Ｘ酶活性、精子畸形率均显著增高，表明三者结果基本一致。本文还对血浆中ＬＤＨ－Ｘ酶活性测定的应用进行了讨论。     

Evaluation of a new Plasmodium lactate dehydrogenase assay (OptiMAL-IT) for the detection of malaria

The new OptiMAL-IT(R) rapid diagnostic test for malaria was evaluated in 271 patients in Thailand with uncomplicated malaria between June and July 2002. The sensitivity and specificity for the diagnosis of Plasmodium falciparum parasites were 88% and 92%, respectively. For species other than P. falciparum, the sensitivity was 65% and specificity was 99%. The performance of the new test decreased markedly at low levels of parasitaemia.     

Assessment of three new parasite lactate dehydrogenase (pan-pLDH) tests for diagnosis of uncomplicated malaria

A study to assess the diagnostic capabilities of three parasite lactate dehydrogenase (pan-pLDH) tests, Vistapan), Carestart and Parabank), was conducted in Uganda. An HRP2 test, Paracheck-Pf), and a Giemsa-stained blood film were performed with the pLDH tests for outpatients with suspected malaria. In total, 460 subjects were recruited: 248 with positive blood films and 212 with negative blood films. Plasmodium falciparum was present in 95% of infections. Sensitivity above 90% was shown by two pLDH tests, Carestart (95.6%) and Vistapan (91.9%), and specificity above 90% by Parabank (94.3%) and Carestart (91.5%). Sensitivity decreased with low parasitaemia (chi(2) trend, P<0.001); however, all tests achieved sensitivity >90% with parasitaemia > or =100/microl. All tests had good inter-reader reliability (kappa>0.95). Two weeks after diagnosis, 4-10% of pLDH tests were still positive compared with 69.7% of the HRP2 tests. All tests had similar ease of use. In conclusion, two pLDH tests performed well in diagnosing P. falciparum malaria, and all pLDH tests became negative after treatment more quickly than the HRP2. Therefore the rapid test of choice for use with artemisinin-combination therapies in this area would be one of these new pLDH tests.     

牛带绦虫乳酸脱氢酶基因原核表达及免疫学特征

目的 克隆牛带绦虫乳酸脱氢酶(lactate dehydrogenase,LDH)基因在大肠杆菌中表达,研究其免疫学特征。方法 将牛带绦虫成虫LDH基因与质粒pET-28a(+)连接构建原核重组质粒,在大肠埃希菌BL21/DE3中用异丙硫代-β-D半乳糖苷诱导表达,表达产物通过十二烷基-聚丙烯酰胺凝胶电泳(SDS-PAGE)和蛋白印迹(Westernblotting)进行分析。结果 牛带绦虫乳酸脱氢酶基因编码332个氨基酸,理论分子量为35.4kDa,PCR、双酶切及DNA测序的结果均证明pET-28a(+)-LDH构建成功;SDS-PAGE结果表明,牛带LDH基因在大肠埃希菌高效表达,经过亲和层析高纯度蛋白,且该重组蛋白可以被亚洲带绦虫、牛带绦虫及猪带绦虫感染病人血清识别,表明其具有免疫反应性。结论 牛带绦虫LDH基因可在原核系统中有免疫活性的高效表达。     

日本血吸虫乳酸脱氢酶原核表达、纯化及鉴定

目的原核表达日本血吸虫乳酸脱氢酶（SjLDH）,对表达产物进行纯化及生物活性鉴定。方法将SjLDH编码序列克隆入pET-28a原核表达载体并转染大肠埃希菌,异丙基-β-D-硫代半乳糖苷（IPTG）诱导表达后亲和层析法对表达产物进行纯化,并用聚丙烯酰胺变性凝胶电泳（SDS-PAGE）、蛋白印迹（Western blot）、酶联免疫吸附实验（ELISA）、酶活性染色及活性测定进行鉴定。结果成功构建pET28a-SjLDH重组质粒并在大肠埃希菌中获得可溶性表达,亲和层析纯化表达产物。SDS-PAGE及Western Blot结果显示,表达及纯化产物分子量约36kDa,与SjLDH理论分子量相符。ELISA结果表明,重组蛋白具有良好的免疫活性,酶活性染色表明重组蛋白具有乳酸脱氢酶活性,活性单位为379U/mg。结论成功进行SjLDH的原核表达并获得纯化的重组蛋白,重组蛋白具有良好的免疫原性及酶活性,为研究SjLDH的功能奠定了基础。     

Human erythrocyte lactate dehydrogenase activity changes in the presence of some alcohol dehydrogenase inhibitors

The aim of the study was to evaluate the effect of alcohol dehydrogenase (ADH; E.C. 1.1.1.1) inhibitors and substrates: cimetidine, 4-methylpyrazole (4MP), EDTA, ethanol and methanol on lactate dehydrogenase (LDH; E.C. 1.1.1.27) activity. The activity of LDH was spectrophotometrically determined in in-vitro prepared diluted hemolysates obtained from human erythrocytes with mentioned compounds at the concentrations 0.01, 0.1, 1.0 mM of cimetidine, EDTA, 4MP and 12.5, 25.0, 50.0 mM of ethanol and methanol. The reaction was conducted at 37 degrees C in pH 7.5 and changes of optical density was measured at lambda = 340nm. LDH activity was significantly inhibited by 0.10 mM (p < 0.05) and 1.0 mM (p < 0.01) of cimetidine and EDTA. There were no observed any significant changes vs. control in LDH activity when 4MP, ethanol or methanol was added to environment of reaction.     

脑脊液乳酸脱氢酶检查在小儿颅内感染中的意义

目的 研究和分析脑脊液乳酸脱氢酶检查对小儿颅内感染中鉴定小儿化脓性脑膜炎以及病毒性脑膜炎的意义.方法 分析201 3年1-6月进入该院治疗化脓性脑膜炎患儿52例,病毒性脑膜炎患儿48例,以及对照组患儿50例,检测脑脊液乳酸脱氢酶的浓度.结果 脑脊液乳酸脱氢酶浓度在病毒性脑炎组（24.5±3.7）U/L,对照组病例（2.2±3.56）U/L,化脓性脑膜炎组结果（54.6±3.4）U/L与其他两组检测结果相比显著增高（P＜0.05）.结论 脑脊液乳酸脱氢酶检测能够作为鉴别小儿化脓性脑膜炎和病毒性脑炎更为快速和准确的诊断标准,对临床上小儿颅内感染疾病的诊断治疗提供更为准确的实验室数据指标,有助于提高小儿颅内疾病的治疗效率.     

Binding of titanium dioxide nanoparticles to lactate dehydrogenase

Objective

Measurement of released lactate dehydrogenase (LDH) activity, a commonly used marker of lethal cell injury in both in vitro and in vivo screenings, has been used to assess the cytotoxicity of nanoparticles (NPs), chemical compounds, and environmental factors. We have recently demonstrated that titanium dioxide (TiO₂) particles bind to several serum proteins. In the present study we investigated the binding of TiO₂ NPs to LDH.

Methods

Purified LDH was incubated with TiO₂ NPs at 37°C for 1 h. The particles were then sedimented by centrifugation, and the activity and quantity of LDH in the supernatant and precipitated fraction were analyzed.

Results

Incubation with TiO₂ reduced the LDH activity in the supernatant in a dose-dependent manner, while LDH activity in the precipitated fraction increased in a dose-dependent manner. Moreover, sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed a TiO₂ dose-dependent reduction in the quantity of LDH protein in the supernatant and an increase of LDH in particulate re-suspensions.

Conclusions

These findings, although based on a purified form of LDH, suggest that TiO₂ NPs bind to LDH, and consequently, TiO₂ NP-induced toxicity could be underestimated by the LDH activity assay.