Enantioselective analysis of N‐hydroxymexiletine glucuronide in human plasma for pharmacokinetic studies

Enzymatic hydrolysis with β‐glucuronidase/sulfatase was used for the enantioselective determination of N‐hydroxymexiletine glucuronide in plasma for pharmacokinetic studies. N‐Hydroxymexiletine glucuronide was determined as the quantity of mexiletine released by hydrolysis (difference between the enantiomeric concentrations of mexiletine obtained with and without hydrolysis). Plasma samples (100 μl) were treated at pH 5.0 with 10 mg of the enzyme (Limpet Acetone Powder type I) for 16 hr at 37°C and extracted at pH 10.4 with diisopropyl ether. Chiral mexiletine discrimination was obtained by reaction with o‐phthalaldehyde/N‐acetyl‐L ‐cysteine, separation of the resulting diastereomers on a C‐18 reversed‐phase column with a mobile phase of methanol–0.05 N acetate buffer, pH 5.5 (6.5:3.5, v/v), and fluorescence detection (λ_ex 350 nm, λ_em 455 nm). The performance characteristics for the enantioselective analysis of mexiletine preceded by enzymatic hydrolysis were recovery ∼90%, quantification limit 1 ng/ml, and linearity up to 1000 ng/ml plasma for both enantiomers. The coefficients of variation obtained in the study of intra‐ and inter‐day precision were respectively 5% and 7% for both enantiomers. The assay was shown to be suitable for a pharmacokinetic study performed in a patient with the arrhythmic form of chronic Chagas' heart disease treated with 200 mg t.i.d. of racemic mexiletine hydrochloride. The high sensitivity of the method allows analysis of only 100 μl plasma. Chirality 11:85–90, 1999. © 1999 Wiley‐Liss, Inc.     

Molecular phylogeography of the Chagas' disease vector Triatoma infestans in Argentina

Triatoma infestans is the main vector of Chagas' disease in South America between latitudes 10°S and 46°S. A multilocus microsatellite data set of 836 individuals from 27 populations of T. infestans, from all its range of distribution in Argentina, was analyzed. Our results favor the hypothesis of two independent migration events of colonization in Argentina and secondary contacts. The majority of the populations of the western provinces of Catamarca, La Rioja, San Juan and the west of Cordoba province, had almost no shared ancestry with the rest of the populations analyzed. Probably those populations, belonging to localities close to the Andean region, could have been established by the dispersal line of T. infestans that would have arrived to Argentina through the Andes, whereas most of the rest of the populations analyzed may have derived from the dispersal line of T. infestans in non-Andean lowlands. Among them, those from the provinces of Formosa, Chaco, Santiago del Estero and Santa Fe shared different percentages of ancestry and presented lower degree of genetic differentiation. The migratory movement linked to regional economies and possibly associated with passive dispersal, would allow a higher genetic exchange among these populations of T. infestans. This study, using microsatellite markers, provides a new approach for evaluating the validity of the different hypotheses concerning the evolutionary history of this species. Two major lineages of T. infestans, an Andean and non-Andean, are suggested.     

Trypanosoma cruzi: 4-aminopyrazolopyrimidine in the treatment of experimental Chagas' disease

An allopurinol metabolite, 4-aminopyrazolopyrimidine, was tested on two different strains of mice (NMRI-IVIC and C57Bl/6J) that had been infected 4 days earlier with the virulent Ya strain of Trypanosoma cruzi. Low doses of 4-aminopyrazolopyrimidine (0.125-0.500 mg/kg body wt/day) for 10 days induced a significant reduction in parasitemia (direct counts and subinoculation experiments) and increased survival time (without any evidence of toxicity) compared with untreated animals. When tested in vitro, 4-aminopyrazolopyrimidine was sixfold more active than allopurinol as a trypanostatic drug. The low therapeutic doses of 4-aminopyrazolopyrimidine suggest that this drug may be useful in the treatment of acute Chagas' disease.     

Cardiovascular risk factors in chronic Chagas' disease are associated with a different profile of putative heart-pathogenic antibodies

Given that cardiovascular risk factors (CRF), such as smoking, alcoholism and hypertension, may contribute to the development of heart lesions, chronically Trypanosoma cruzi-infected individuals were studied to explore the relationship between the presence of such CRF, cardiomyopathy and antibodies that have been proposed to play a pathogenetic role in Chagas' disease. The targets of these antibodies were T. cruzi antigens such as cruzipain (Cz), a P ribosomal antigen (P2), and a component of myelin sheaths also present in T. cruzi (sulphatide). Individuals were classified into four groups on the basis of specific serology and presence of CRF: subjects with T. cruzi infection and CRF; those with positive serology and no CRF; seronegatives with CRF; and seronegatives without CRF, were analysed. Seronegatives or seropositives with CRF showed a greater occurrence of heart involvement (chest X-ray and/or electrocardiogram abnormalities). Seropositives with CRF displayed significantly higher levels of antisulphatide antibodies than the three remaining groups and higher levels of antibodies against Cz and P2 compared to the seropositives without CRF. Increased amounts of anti-P2 and antisulphatide antibodies were also found in seropositives with marked heart involvement. The presence of CRF is associated with a different profile of antibody responses and degree of cardiac effects.     

Blood pressure and heart rate variability are linked with hyperphosphatemia in chronic kidney disease patients

ABSTRACT

Hyperphosphatemia is a common complication of chronic kidney disease (CKD) and is associated with cardiovascular disease (CVD), which has contributed to an increase in mortality of CKD patients. The onset of CVD often varies by time-of-day. Acute myocardial infarction or ventricular arrhythmia occurs most frequently during early morning. Blood pressure (BP) and heart rate circadian rhythms account for the diurnal variations in CVD. Preservation of normal circadian time structure from the cardiomyocyte level to the whole organ system is essential for cardiovascular health and CVD prevention. Independent risk factors, such as reduced heart rate variability (HRV) and increased BP variability (BPV), are particularly prevalent in patients with CKD. Analysis of HRV is an important clinical tool for characterizing cardiac autonomic status, and reduced HRV has prognostic significance for various types of CVD. Circadian BP rhythms are classified as extreme dipper, dipper, non-dipper or riser. It has been reported that nocturnal riser BP pattern contributes to cardiovascular threats. Previous studies have indicated that the circadian rhythm of serum phosphate in CKD patients is consistent with the general population, with the highest diurnal value observed in the early morning hours, followed by a progressive decrease to the lowest value of the day, which occurs around 11:00 am. Rhythm abnormalities have become the main therapeutic target for treating CVD in CKD patients. It has been reported that high levels of serum phosphate are associated with reduced HRV and increased BPV in CKD patients. However, the mechanisms related to interactions between hyperphosphatemia, HRV and BPV have not been fully elucidated. This review focuses on the evidence and discusses the potential mechanisms related to the effects of hyperphosphatemia on HRV and BPV.     

血浆N端脑钠肽前体与COPD合并慢性肺源性心脏病患者肺动脉压的相关性分析

目的探讨血浆N端脑钠肽前体与COPD合并慢性肺源性心脏病患者肺动脉压的相关性。方法将我院收治的94例COPD稳定期患者,根据心脏彩超检查明确合并慢性肺心病患者分为观察组46例,单纯COPD患者分为对照组48例,采用化学发光法测定两组患者血浆NT-proBNP水平,心脏彩超检查两组患者右心室前壁厚度、右心室舒张末期内径、左室射血分数、主肺动脉宽度、肺动脉压,并进行比较。结果与对照组比较,观察组血浆NT-proBNP水平明显升高(P0.05),右心室前壁厚度、右心室舒张末期内径、主肺动脉宽度、肺动脉压明显升高(P0.05)。血浆NT-proBNP水平与肺动脉压成明显正相关(r=-0.284,P0.01)。结论 COPD合并慢性肺心病患者血浆NT-proBNP水平升高,有助于评估肺动脉高压的严重程度。     

Prosthetic valves in adult patients with congenital heart disease: Rationale and design of the Dutch PROSTAVA study

Background

Data on long-term complications in adult patients with congenital heart disease (ACHD) and a prosthetic valve are scarce. Moreover, the influence of prosthetic valves on quality of life (QoL) and functional outcome in ACHD patients with prosthetic valves has not been studied.

Objectives

The primary objective of the PROSTAVA study is to investigate the relation between prosthetic valve characteristics (type, size and location) and functional outcome as well as QoL in ACHD patients. The secondary objectives are to investigate the prevalence and predictors of prosthesis-related complications including prosthesis-patient mismatch.

Methods

The PROSTAVA study, a multicentre cross-sectional observational study, will include approximately 550 ACHD patients with prosthetic valves. Primary outcome measures are maximum oxygen uptake during cardiopulmonary exercise testing and QoL. Secondary outcomes are the prevalence and incidence of valve-related complications including prosthesis-patient mismatch. Other evaluations are medical history, physical examination, echocardiography, MRI, rhythm monitoring and laboratory evaluation (including NT-proBNP).

Implications

Identification of the relation between prosthetic valve characteristics in ACHD patients on one hand and functional outcome, QoL, the prevalence and predictors of prosthesis-related complications on the other hand may influence the choice of valve prosthesis, the indication for more extensive surgery and the indication for re-operation.     

Proteomic surveillance of autoantigens in patients with Behcet's disease by a proteomic approach

To promote an understanding of autoimmunity in BD, we surveyed autoAgs in patients with BD and investigated the prevalence and clinical significance of the identified autoAbs. Specifically, proteins, extracted from peripheral blood mononuclear cells and separated by 2DE, were subjected to WB, using five serum samples from patients with BD. The detected candidate autoAgs were identified by mass spectrometry. As a result, 17 autoantigenic spots were detected by the 2DE‐WB, out of which eight spots were identified. They are enolase‐1, cofilin‐1, vimentin, Rho‐GDI β protein, tubulin‐like protein, and actin‐like proteins. The autoAbs to one of the identified proteins, cofilin‐1, were investigated by WB using a recombinant protein in 30 patients with BD, 35 patients with RA, 32 patients with SLE, and 16 patients with PM/DM. The autoAbs to cofilin‐1 were detected by WB in four (13.3%) of the 30 patients with BD, five (14.3%) of the 35 patients with RA, two (6.3%) of the 32 patients with SLE, and eight (24.2%) of the 33 patients with PM/DM. Our data indicate that the generation of autoAbs to cofilin‐1 may reflect common immunological disorders in BD, RA, and PM/DM. Our data would help understanding of the immunopathology of BD. In addition, the proteomic approach would be a useful way to investigate autoAgs.     

Significance of serum trace element status in patients with rheumatic heart disease

It is known that certain trace elements can affect various heart diseases. In this study, we aimed to evaluate the changes in concentrations of certain serum trace elements in patients with chronic rheumatic heart disease (RHD). Serum analysis of selenium (Se), zinc (Zn), and copper (Cu) trace elements was assayed by atomic absorption spectrophotometry. RHD patients had significantly lower serum concentrations of Se and Zn than control subjects (p<0.05 and p<0.001, respectively). However, the serum Cu concentration was significantly higher in RHD patients than in controls (1.93±0.59 μg/L vs 1.06±0.29 μg/L; p<0.001). Similarly, the Cu/Zn ratio in RHD patients was higher than in control subjects (4.70±0.92 vs 1.68±0.45; p<0.001). Additionally, no significant correlation was found among these trace element concentrations and the functional capacity classes (p>0.05). RHD patients had decreased serum Se and Zn element concentrations and increased serum Cu element concentration. We suggest that Se and Zn deficiency might be contributory factors in the development of rheumatic heart disease, and a high Cu concentration and a high Cu/Zn ratio might reflect an ongoing inflammatory process in this disease.     

Changes and significance of serum angiopoietin-2 levels in patients with coronary heart disease

Coronary heart disease (CHD) is characterized by inflammatory process and endothelial dysfunction. To investigate angiopoietin-2 (Ang-2) profiles, we evaluated serum Ang-2 levels in different types of CHD in 166 subjects. Ang-2 was measured by enzyme-linked immunosorbent assay. Serum Ang-2 levels were significantly elevated in patients with CHD and gradually increased with advance of CHD. Ang-2 was positively correlated with Gensini scores and hs-CRP. Ang-2 might have potential implication in detecting and monitoring the progression of CHD.     

Production of reactive oxygen species by monocyte-derived macrophages from blood of healthy donors and patients with ischemic heart disease

Production of reactive oxygen species (ROS) by macrophages derived from blood monocytes of healthy donors (MP_N) and patients with ischemic heart disease (IHD) (MP_IHD) before, during, and after their incubation with low-density lipoprotein (LDL) isolated from blood plasma of healthy donors (LDL_N) and patients with a high cholesterol level (LDL_H) was investigated by the method of luminol-dependent (spontaneous) and stimulated chemiluminescence (CL) using opsonized zymosan (OZ) or phorbol-12-myristate-13-acetate (PMA) as the CL stimulators. It was shown that proper, luminol-dependent, and zymosan-or PMA-stimulated chemiluminescence of MP_IHD was 1.4-, 1.8-, 2.7-, and 1.6-fold higher than the same types of chemiluminescence of MP_N, respectively, (p<0.05–0.01). Although the effect of OZ on MP_N and MP_IHD was more potent than that of PMA (by 4.3- and 3.2-fold, respectively), but it appeared in 2.5–3.0 times slower than that of PMA. LDL_N and LDL_H incubated with MP_N for the first 15 and 60 min caused the 1.4- and 2.5-increase of the luminol-dependent CL of MP_N; the same treatment of MP_IHD did not influence ROS production by these cells. Repeated increase in the OZ-stimulated CL of MP_N was also observed after preincubation for 15–180 min with LDL_N and LDL_H followed by LDL removal, subsequent MP_N washing and addition of Hanks solution and OZ; the repeated increase in OZ-stimulated CL of MP_N was only observed after incubation with LDL_H than with LDL_N. No increase of CL was observed in experiments with MP_IHD. Thus, more intensive chemiluminescence of macrophages obtained from blood of patients with IHD suggests their in vivo stimulation. LDL_N and LDL_H may cause both primary and secondary (after preincubation) stimulating effect on CL of MP_N but not of MP_IHD. Thus, the analysis of macrophage chemiluminescence is a sensitive test for evaluation the degree of macrophage stimulation; it may be effectively used for monitoring of effectiveness of medical treatment of patients.     

Direct measurements of metabolism, activity and feeding behaviour of pike, Esox Zucius L., in the wild, by the use of heart rate telemetry

Heart rate telemetry records of up to 5 days duration were obtained from pike living in Lochs Kinord and Davan, Scotland. Applying metabolic rate correlations it was found that mean metabolic rate (R) was 1.5 times standard metabolic rate (R_a), The fish rarely worked near their metabolic limits. Activity metabolism (R_a) was much higher than estimates based on mean swimming speed and comprised up to 10% of R. Most activity metabolism was the result of localized bursts of activity. Less than 10% of activity showed evidence of oxygen debt. Specific Dynamic Action or feeding metabolism (R_f) comprised 15–25% of R. Food intake estimated from heart rate was 1.5% wet body weight day⁻¹, consumed in the form of small items captured during the day and digested during the afternoon and night.     

Comparison of epitope specificity of anti-heat shock protein 60/65 IgG type antibodies in the sera of healthy subjects,patients with coronary heart disease and inflammatory bowel disease

Previously, we reported on the presence of antibodies to linear epitopes of human and mycobacterial 60 kD heat shock proteins (HSP) in the sera of healthy blood donors. Since many recent findings indicate that the levels of these antibodies may be altered in coronary heart disease (CHD) and also inflammatory bowel diseases (IBD), it seemed worthwhile to compare the epitope specificity of the anti-HSP60 and anti-HSP65 antibodies in the sera of patients with these diseases to those in healthy subjects. The multipin enzyme-linked immunosorbent assay method was applied with a large overlapping set of synthetic 10-mer peptides covering selected regions of human HSP60 and Mycobacterium bovis HSP65. Sera of 12 healthy persons (HP), 14 CHD, and 14 IBD patients with the same concentration of total anti-HSP60 and HSP65 IgG antibodies were tested. We have identified CHD-specific epitopes in the equatorial domain of the HSP60 protein but in neither region of the HSP65 molecule, indicating that the formation of anti-HSP60 antibodies is not or only partially due to the cross-reaction between human HSP60 and bacterial HSP65. IBD-specific epitopes were found in many regions of the HSP60 and in even more regions of the HSP65 molecule including an IBD-specific T cell epitope in region X as well. These findings indicate that the epitope specificity of the anti-human and anti-mycobacterial HSP60 antibodies associated with various diseases is different.     

The prognostic value of the serum ferritin in a southern Brazilian cohort of patients with Gaucher disease

The clinical utility of serum ferritin as a biomarker of disease severity and prognosis in Gaucher disease (GD) is still debated. Here, we aimed to evaluate ferritin and its relation to clinicolaboratory parameters of GD patients seen at the Reference Center for Gaucher Disease of Rio Grande do Sul, Brazil, so as to gather evidence on the utility of ferritin as a biomarker of this condition. A retrospective chart review was performed collecting pre-and posttreatment data from GD patients. Eighteen patients with ferritin levels available before and after treatment were included in the study. Nine of these participants were males, and seventeen had type I GD. All patients were given either enzyme replacement (n = 16) or substrate reduction therapy (n = 2), and ferritin was found to decrease from 756 [318-1441] ng/mL at baseline to 521 [227-626] ng/mL (p=0.025) after 28.8 month soft treatment. Serum ferritin levels did not correlate with measures of disease severity, but showed an association with age at onset of treatment (ρ= 0.880; n = 18; p < 0.001). In conclusion, although serum ferritin did not correlate with disease severity, after a median 28.8 months of treatment, clinical outcomes had clearly improved, and ferritin levels had decreased.     

普罗布考联合家庭心脏康复训练对冠心病患者射血分数、血脂的影响分析

目的 探讨普罗布考联合家庭康复训练对冠心病患者射血分数及血脂水平的影响.方法 选择2018年6月 ～2020年6月我院收治的冠心病患者110例,按随机数字表法分成对照组和实验组,对照组给予瑞舒伐他汀治疗,实验组在对照组的基础上联合应用普罗布考及家庭康复训练.对两组患者治疗前及治疗后3个月的心功能指标、血脂水平进行检测比...     

Novel P143L polymorphism of the LCAT gene is associated with dyslipidemia in Chinese patients who have coronary atherosclerotic heart disease

Coronary atherosclerotic heart disease (CAD) is a multifactorial disorder resulting from numerous gene-gene and gene-environment interactions. Lecithin:cholesterol acyltransferase (LCAT), a key enzyme in reverse cholesterol transport and the metabolism of high-density lipoprotein (HDL), is thought to be a candidate gene related to dyslipidemia and CAD. Variations in the LCAT gene were investigated in 190 CAD patients and 209 age- and gender-matched controls by denaturing high-performance liquid chromatography, and confirmed by sequencing and RFLP assay. In CAD patients, a novel single-nucleotide polymorphism (P143L) in exon 4 of the LCAT gene was discovered in nine males and two females (frequency of 5.79%), which was found in none of 209 controls. The genotype and allele distribution of P143L is significantly (P<0.04 ) higher in the low HDL-C subgroup than in the normal HDL-C subgroup in both male patients and all CAD patients. P143L was also found to be significantly (P<0.01) associated with the low HDL-C phenotype in both male patients and all CAD patients, with odds-ratios of 7.003 (95% CI 2.243-21.859) and 5.754 (95% CI 1.893-13.785), respectively. Thus, the P143L polymorphism may play a role in causing decreased HDL-C levels, leading to increased risk of dyslipidemia and CAD in Chinese.     

A modified multiplex ligation-dependent probe amplification method for the detection of 22q11.2 copy number variations in patients with congenital heart disease

Background

Copy number variations (CNVs) of chromosomal region 22q11.2 are associated with a subset of patients with congenital heart disease (CHD). Accurate and efficient detection of CNV is important for genetic analysis of CHD. The aim of the study was to introduce a novel approach named CNVplex®, a high-throughput analysis technique designed for efficient detection of chromosomal CNVs, and to explore the prevalence of sub-chromosomal imbalances in 22q11.2 loci in patients with CHD from a single institute.

Results

We developed a novel technique, CNVplex®, for high-throughput detection of sub-chromosomal copy number aberrations. Modified from the multiplex ligation-dependent probe amplification (MLPA) method, it introduced a lengthening ligation system and four universal primer sets, which simplified the synthesis of probes and significantly improved the flexibility of the experiment. We used 110 samples, which were extensively characterized with chromosomal microarray analysis and MLPA, to validate the performance of the newly developed method. Furthermore, CNVplex® was used to screen for sub-chromosomal imbalances in 22q11.2 loci in 818 CHD patients consecutively enrolled from Shanghai Children’s Medical Center. In the methodology development phase, CNVplex® detected all copy number aberrations that were previously identified with both chromosomal microarray analysis and MLPA, demonstrating 100% sensitivity and specificity. In the validation phase, 22q11.2 deletion and 22q11.2 duplication were detected in 39 and 1 of 818 patients with CHD by CNVplex®, respectively. Our data demonstrated that the frequency of 22q11.2 deletion varied among sub-groups of CHD patients. Notably, 22q11.2 deletion was more commonly observed in cases with conotruncal defect (CTD) than in cases with non-CTD (P < 0.001). With higher resolution and more probes against selected chromosomal loci, CNVplex® also identified several individuals with small CNVs and alterations in other chromosomes.

Conclusions

CNVplex® is sensitive and specific in its detection of CNVs, and it is an alternative to MLPA for batch screening of pathogenetic CNVs in known genomic loci.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1590-5) contains supplementary material, which is available to authorized users.     

Study on the variation in X-ray sensitivity of human diploid skin fibroblasts. Normal radiosensitivity of cells derived from patients with Huntington's disease

9 cell strains derived from patients with Huntington's disease and 9 from age- and sex-matched controls were investigated for X-ray sensitivity. No differences in radiosensitivity were observed for the two groups. The two groups taken together reveal a dependence of radiosensitivity on intrinsic cloning efficiency which in turn correlates with donor age. A difference in radiosensitivity between males and females is also indicated although at the borderline of significance. As a parameter for radiosensitivity the dose needed to obtain 0.1% survival appears superior to the Do.