Direct mass spectrometric analysis of body fluids from acutely poisoned patients

A computer-controlled mass spectrometric (MS) analysis of urine, blood and gastric aspirate from 21 drug overdose patients and urine from 15 heroin/morphine addicts, using a direct sample extract introduction into the MS via a membrane separator, is described. The Llewellyn 3 stage membrane separator inlet system allows rapid mass spectrometric analysis, eliminating time-consuming gas chromatographic separation or derivatization of the sample. A computer is used to identify the drugs or drug metabolites according to their fractionation pattern. Normal body fluid components do not interfere significantly. The sensitivity limit of the system is 0.5 mg/100 ml. The total analysis time including the sample work-up is 10 min.     

Methylated DNA as a possible screening marker for neoplastic disease in several body fluids

Early detection appears to be one of the most important approaches to reducing mortality caused by neoplasia. Changes in DNA methylation have been recognized as one of the most common molecular alterations in human tumors. Due to the ubiquity of DNA methylation changes and the possibility to detect methylated DNA in several body fluids, this specifically altered DNA may serve as a potential new screening marker for neoplastic disease.     

Alpha-fetoprotein in pericardial,peritoneal, and pleural fluids: A body fluid matrix evaluation

ObjectivesAlpha-fetoprotein (AFP) measurement in pericardial, peritoneal (ascites), and pleural fluids is sometimes requested by clinicians as supportive evidence in the evaluation of suspected malignancy. As commercially available, Food and Drug Administration (FDA)-cleared AFP assays are not validated for these fluid types, laboratories must complete additional validation studies to comply with regulatory requirements for body fluid testing. The objective of this study was therefore to conduct a matrix evaluation for these body fluid types using the Beckman Access AFP assay on the UniCel DxI 800 immunoassay system.Design and methodsUsing an Institutional Review Board (IRB) approved protocol, previously collected pericardial fluid, peritoneal fluid, pleural fluid, and serum specimens were de-identified and frozen at −20 °C prior to matrix evaluation experiments. Spiked recovery, mixed recovery/linearity, and precision studies were conducted.ResultsIn spiked and mixed recovery studies, the average percent (%) recovery was within predefined acceptable limits (±15%) for all three body fluids. Linearity was observed over the analytical measurement range (AMR) for all three body fluids (slope, intercept, systematic error): pericardial 0.988, −0.1, 6.1%; peritoneal 0.986, 0.0, 4.1%; and pleural 1.016, 0.0, 1.6%. Imprecision was ≤6.0% CV for all three body fluids at both high and low AFP concentrations.ConclusionsMatrix interference with AFP testing was not observed for pericardial, peritoneal, or pleural fluids on the Beckman UniCel DxI 800 system.     

Characteristics of peripheral and peritoneal white blood cells in children with chronic renal failure, dialyzed or not.

OBJECTIVE: To explore further the mechanisms leading to immune deficiency in chronic renal failure and the role of dialysis treatment in these mechanisms. DESIGN: Cross-sectional and longitudinal analysis. PATIENTS: We studied 39 children treated with peritoneal dialysis (PD), 23 children treated with hemodialysis (HD), 33 children not yet dialyzed [chronic renal failure (CRF)], and 27 healthy children. Peritoneal cells were also obtained from PD children for analysis. METHODS: White blood cells (WBCs) were isolated from blood and peritoneal dialysis effluent by centrifugation. The number of CD2+, CD4+, and CD8+ T cells, B cells, and natural killer cells were measured by flow cytometry. RESULTS: The total peripheral blood lymphocyte count was lower in PD children (2.6 x 10(9)/L), HD children (2.1 x 10(9)/L), and CRF children (2.0 x 10(9)/L) compared with healthy children (3.1 x 10(9)/L, p < 0.05). The B lymphocyte count was also lower in PD children (0.34 x 10(9)/L), HD children (0.22 x 10(9)/L), and CRF children (0.33 x 10(9)/L) compared with healthy children (0.52 x 10(9)/L, p < 0.01). Numbers of CD4+ T cells were not different, but numbers of CD8+ T cells were lower in PD children (0.56 x 10(9)/L), HD children (0.63 x 10(9)/L), and CRF children (0.53 x 10(9)/L) compared with healthy children (0.77 x 10(9)/L, p < 0.05). The count of natural killer cells was lower in PD children (0.21 x 10(9)/L), HD children (0.17 x 10(9)/L), and CRF children (0.18 x 10(9)/L) compared with healthy children (0.50 x 10(9)/L, p < 0.0001). The CD4/CD8 ratio of lymphocytes in peritoneal effluent was 0.8 versus 1.9 in peripheral blood (p < 0.001). The CD2/CD19 ratio was not different. The cell subsets remained stable during the first year of PD treatment. The CD2/CD19 ratio in peritoneal effluent was higher in children with a peritonitis incidence > or = 1 per year. CONCLUSIONS: The reduced numbers of B lymphocytes, CD8+ T cells, and natural killer cells found in CRF children, dialyzed or not, may favor the frequent occurrence of infections.     

腹膜透析患者不同腹膜转运类型与炎症反应、营养不良相关关系的研究

目的了解腹膜透析患者腹膜转运类型与炎症反应、营养不良的相关关系.方法对46例腹膜透析患者行腹膜平衡试验(PET),根据PET结果将腹膜转运功能分为高转运(H)、高于平均(HA)、低于平均(LA)和低转运(L)四型.检查血TNF-α、IL-6以及白蛋白水平,并用主观整体评估(SGA)法评估患者的营养状态.结果 TNF-α水平高转运与低转运相比,差异有显著性(P<0.05);而白蛋白高转运与低转运相比,差异也有显著性(P<0.05);IL-6则无显著性差异.根据SGA标准,H营养不良的发生率较L显著增高 (P<0.05).结论腹膜通透性高的患者其炎症反应和营养不良的发生率高,说明高通透性的腹膜转运类型是造成炎症反应、营养不良的主要原因.     

Diagnosis and management of bone disorders in chronic renal failure and dialyzed patients

Renal osteodystrophy is multifactorial. Decreased calcium absorption from the GI tract, secondary to low calcitriol levels; hyperphosphatemia; skeletal resistance to the action of parathormone; and aluminum deposition on the surface of the bones are its main pathogenetic mechanisms. Its biochemical features include abnormalities in serum calcium, phosphate, alkaline phosphatase, parathormone, calcitriol, and aluminum concentration. Radiographic methods are of little use in the characterization of the type of osteodystrophy present, but they may be of help in assessing mineral loss from the skeleton. Clinical manifestations are from bones (pain, deformities, fractures) or from metastatic calcifications. Bone biopsy is the definitive means of diagnosis. The main histologic types of osteodystrophy include osteitis fibrosa, osteomalacia, mixed form (with features of both osteitis fibrosa and osteomalacia), and aluminum osteodystrophy (presenting as either osteomalacia or aplastic lesion). The management of renal osteodystrophy should address all the pathogenetic mechanisms. Correction of the abnormalities in calcium and phosphate metabolism and prevention of aluminum osteodystrophy are the cardinal rules of management. Specific measures (parathyroidectomy, chelation of aluminum) have clear-cut indications and usually require a bone biopsy.     

Neutral steroid metabolites in patients with uraemia and after renal transplantation: a quantitative and qualitative study in body fluids

Steroid metabolites enriched from urine, haemofiltrate, and CAPD-dialysate (Continuous Ambulatory Peritoneal Dialysis) were identified by gas chromatography-mass spectrometry and quantified by capillary gas chromatography. The study included twenty healthy controls, twenty-six non-dialysed uraemics, thirty-nine patients on regular dialysis treatment, and twenty-two allograft recipients. Compared to the 24 h urinary excretion rates of controls the excretion rates of androsterone and etiocholanolone were in the lower normal range up to significantly decreased in the body fluids of all patients, and those of the corticoid metabolites were also significantly decreased. 11-Oxygenated androstanolones in urine from non-dialysed uraemics correlated significantly decreased. 11-oxygenated androstano-levels and were significantly increased, but normal in haemofiltrate and CAPD-dialysate, while in urine of allograft recipients the values were significantly lower.     

大鼠腹股沟脂肪间充质干细胞的分离、培养、鉴定及活体标记

背景：脂肪间充质干细胞具有来源丰富、取材方便、体外有较强增殖能力并具有多向分化的特点,有望成为骨组织工程、细胞治疗等的种子细胞。目的：培养扩增大鼠脂肪源间充质干细胞,以活体标记并鉴定其分化潜能。方法：无菌条件下取大鼠双侧腹股沟脂肪,Ⅰ胶原酶消化法分离培养脂肪源性干细胞,胰蛋白酶消化法传代扩增。取第 3 代脂肪干细胞进行流式检测 HCAM、CD106、CD29、CD49D 和 CD34,生长曲线测定,吉姆萨染色,菲立磁标记,成脂诱导后油红 O 染色及成骨诱导后茜素红染色钙结节。结果与结论：细胞呈长梭形漩涡样生长,第 3 代细胞流式鉴定 CD29 阳性,CD34、HCAM、CD49d、CD106 低表达,生长曲线测定有对数生长期,平台期,菲立磁标记阳性率达 80%,并且在一些诱导剂下分化为脂肪细胞及成骨细胞。提示,来源于大鼠腹股沟脂肪分离获得的脂肪源干细胞易于培养和传代扩增,并可活体标记且在特殊条件下可分化为成骨细胞和脂肪细胞。     

大鼠腹股沟脂肪间充质干细胞的分离、培养、鉴定及活体标记

背景:脂肪间充质干细胞具有来源丰富、取材方便、体外有较强增殖能力并具有多向分化的特点,有望成为骨组织工程、细胞治疗等的种子细胞。目的:培养扩增大鼠脂肪源间充质干细胞,以活体标记并鉴定其分化潜能。方法:无菌条件下取大鼠双侧腹股沟脂肪,Ⅰ胶原酶消化法分离培养脂肪源性干细胞,胰蛋白酶消化法传代扩增。取第 3 代脂肪干细胞进行流式检测 HCAM、CD106、CD29、CD49D 和 CD34,生长曲线测定,吉姆萨染色,菲立磁标记,成脂诱导后油红 O 染色及成骨诱导后茜素红染色钙结节。结果与结论:细胞呈长梭形漩涡样生长,第 3 代细胞流式鉴定 CD29 阳性,CD34、HCAM、CD49d、CD106 低表达,生长曲线测定有对数生长期,平台期,菲立磁标记阳性率达 80%,并且在一些诱导剂下分化为脂肪细胞及成骨细胞。提示,来源于大鼠腹股沟脂肪分离获得的脂肪源干细胞易于培养和传代扩增,并可活体标记且在特殊条件下可分化为成骨细胞和脂肪细胞。     

Low molecular mass IgM in urine of patients with primary and secondary renal diseases

Urine specimens from 50 patients with various renal diseases and from 10 normal subjects were examined for the presence of low molecular mass (7S) IgM by double diffusion, using a specific anti-7S IgM serum. 7S IgM in urine was also quantitated by radial immunodiffusion in 5% polyacrylamide gel. It was found with high frequency in the urine of patients with secondary renal diseases associated with multi-system immunological disorders, especially in systemic lupus erythematosus. Two of the patients with lupus nephritis and positive results for 7S IgM in urine were initially diagnosed as nephrotic syndrome due to a primary renal disease; the symptoms and signs of systemic lupus erythematosus developed later. However, 7S IgM was not detected in the urine of patients with primary renal diseases (except for two cases of nephrotic syndrome), nor in normal subjects. Thus detection of 7S IgM in urine may be helpful in the differential diagnosis of primary and secondary renal diseases.     

Concentration of adipokines in peritoneal effluent: a new marker of acute peritonitis in peritoneal dialysis patients?

BACKGROUND: An early and reliable diagnostic procedure for acute peritonitis in patients on peritoneal dialysis (PD) without typical clinical symptoms remains an important challenge in modern nephrology. During the first days of peritonitis, establishing the diagnosis based on peritoneal effluent pleocytosis and inflammatory markers [C-reactive protein (CRP) or interleukin-6] is not efficient in all cases. Increased peritoneal membrane permeability is a well-known consequence of peritonitis. Therefore, we evaluated the concentrations of selected circulating adipose tissue-derived proteins in the peritoneal effluent of PD patients with episodes of acute peritonitis. Material and METHODS: Concentrations of adiponectin and leptin, in both plasma and peritoneal effluent, were assessed in 24 adult PD patients with peritonitis episodes confirmed by clinical symptoms and/or microbiological tests, and in 23 PD patients without signs and symptoms of inflammation (control group). RESULTS: In peritoneal effluent collected from patients with acute peritonitis (also without pleocytosis or increased CRP), both adiponectin and leptin concentrations were markedly elevated: adiponectin 744.1 (344.2 - 1144.1) ng/mL vs 4.8 (3.1 - 6.5) ng/mL; leptin 16.3 (9.4 - 23.1) ng/mL vs 5.1 (0.5 - 9.6) ng/mL. Receiver operating characteristic analyses revealed that peritoneal effluent adiponectin concentration >180 ng/mL has 100% sensitivity and 100% specificity, while peritoneal effluent leptin concentration >11.0 ng/mL has 58.3% sensitivity and 95.5% specificity for the diagnosis of acute peritonitis. The increases in adiponectin and leptin concentrations in peritoneal effluent were not consequences of changes in their plasma levels. A positive correlation between peritoneal effluent and plasma concentrations of adiponectin and leptin in patients with peritonitis was found. CONCLUSION: Increased concentration of leptin and especially adiponectin in peritoneal effluent seems to be a valuable and new early marker of high peritoneal membrane permeability due to acute peritonitis.     

探讨腹膜不同转运类型对腹膜透析患者的营养状况、炎症及心血管并发症的影响

目的探讨腹膜不同转运类型对腹膜透析患者的营养状况、炎症状态及心血管并发症的影响。方法对101名慢性肾衰竭腹膜透析患者据腹膜PET试验进行分组,即A组为高转运;B组为高平均转运;C组为低平均转运;D组为低转运,对患者的营养状况、炎症状态及心血管并发症进行调查分析。结果①各组营养不良的发生率比较:A组:营养不良者5例,营养不良的发生率为62.5%;B组:营养不良25例,营养不良的发生率为58.1%;C组:营养不良14例,营养不良的发生率为35%;D组:均为营养良好,营养不良的发生率为0。各种腹膜不同转运类型的患者营养不良的发生情况差异具有显著性(P=0.003),其中低转运组营养状况最好,高转运组营养状况最差。②腹膜转运类型与炎症的关系:与C组比较,A组、B组的超敏C反应蛋白水平更高(分别为P=0.020、P=0.021)。③各组心血管并发症的发病率:A组为:62.50%;B组为:55.81%,C组为:47.5%;D组为:30%。心血管并发症的发生情况在4种腹膜转运类型中差异具有显著性(P=0.038),以高转运组发生率最高,低转运组发生率最低。④Pearsons相关性分析显示:患者的腹膜转运类型与心血管并发症(r=0.268,P=0.007)和炎症状态(r=0.312,P=0.002)呈正相关,与SGA评估的营养状况(r=-0.348,P=0.000),具有负相关性。结论腹膜转运特性与CAPD患者的营养状况、炎症状态及心血管并发症的发生密切相关,腹膜功能呈现高转运状态、同时合并有较差的营养状态时易发生MIA综合征。在临床工作中,应常规评估腹膜转运特性,及时调整治疗处方,有益于改善患者的营养状况、减少心血管疾病的发生。     

腹膜透析和血液透析病人行肾移植回顾性研究

目的 通过对比分析腹膜透析病人和血液透析病人接受肾移植的术前、术后临床和血生化指标,了解对肾移植疗效的影响。方法 回顾分析本院1997年前接受肾移植并定期随访病人共452例,其中由腹透进入肾移植的病人共49例为腹透组,由血液透析进入肾移植的病人用计算机随机匹配98例为血透组,对二组病人肾移植前后生化指标、肝炎病毒感染率、术后感染等并发症、移植肾失功、医疗费用等进行评估分析。结果 移植前腹透组血浆白蛋白水平低于血透组,移植后血浆白蛋白水平二组无差异。在达到同样血红蛋白水平,腹透病人所需促红素较低;乙肝、丙肝感染率二组分别为28.57％、2.04％和35.71％、13.27％,腹透组显著低于血透组。术后肾功能恢复情况,移植肾失功率无显著差异。术后感染率腹透组为6.12％低于血透组的12.87％。结论 腹膜透析病人纠正贫血所需促红素量较低,血浆白蛋白水平较低但不影响移植效果。HBV、HCV感染率、术后并发症发生率低。移植后肾功能恢复和维持,移植肾失功率以及总医疗费用与血透病人无显著差异。     

Ischemia-modified albumin,a predictive marker of major adverse cardiovascular events in continuous ambulatory peritoneal dialysis patients

Objectives

The aim of this study was to evaluate the efficiency of ischemia-modified albumin (IMA) for predicting major adverse cardiovascular events (MACE) in continuous ambulatory peritoneal dialysis (CAPD) patients.

Design and methods

A prospective observational study was conducted with 120 CAPD patients and 37 healthy volunteers. Demographic and clinical data were collected. The primary end point is the occurrence of MACE.

Results

A total of 157 participants with a mean age of 43.64 years finally completed this study. The CAPD patients had a significantly high rate of MACE (P = 0.001) and high levels of IMA than healthy controls (P < 0.001). Compared with CAPD patients with normal levels of IMA, the CAPD patients with high levels of IMA (> 85 kU/L) had lower non-MACE survival rate (P < 0.001), which indicated that the high IMA CAPD patients may suffer a high rate of MACE. In addition, the high IMA CAPD patients also had a low level of serum albumin (P < 0.001) and hemoglobin (P = 0.018). The correlation analysis showed that the serum albumin level was the most effective factor influencing IMA (B = − 0.967, P < 0.001).

Conclusions

CAPD patients with high levels of IMA had a high incidence rate of MACE. IMA was a good predictive marker of MACE and might be important in cardiovascular risk stratification of CAPD patients.     

HE4-test of urine and body fluids for diagnosis of gynecologic cancer

Introduction: Serum epididymis protein 4 (HE4) level is a useful biomarker for the management of ovarian and endometrial cancer patients. Urine HE4-test, with its easier access than serum test, has emerged as a new method with promising application for the diagnosis of ovarian cancer.

Areas covered: This review summarizes data regarding the detection and alteration of HE4 in urine samples collected from ovarian cancer patients and controls. The performance and limitation of the assay and potential direction of future study are also discussed.

Expert commentary: Several studies have demonstrated an appreciable efficiency of urine HE4-test in the discrimination of ovarian cancer patients from general population. However, the data is based on small cohorts, and the performance of urine HE4-test need to be validated in larger groups. An algorithm incorporating other important factors may allow a quantitative assessment of cancer possibility. Future studies on the HE4 renal secretion and HE4 degradation dynamics in urine are also required for the establishment of standard protocols for the application of urine HE4-test in clinical settings.     

Lipids, waist circumference and body mass index in haemodialysis patients

In the general population, dyslipidaemia and abdominal obesity are risk factors for cardiovascular disease, but less is known about their roles in patients on maintenance haemodialysis (HD). This study investigated the association between blood lipids and abdominal obesity, as determined by waist circumference (WAC), and body mass index (BMI) in 72 HD patients (32 women). Blood lipid levels were measured using routine laboratory methods. Abdominal obesity, based on the WAC measurement, was found in 62.5% of HD patients (75.0% of women; 52.5% of men). Triglyceride levels were higher in abdominally obese compared with abdominally non-obese men. According to BMI measurements, 34.7% of HD patients were overweight/obese and 9.7% were underweight. In men, WAC and BMI were positively correlated with triglyceride levels. In women, WAC was negatively correlated with high-density lipoprotein (HDL) cholesterol and apolipoprotein A levels. The results indicated that there was a negative association of abdominal obesity and BMI with HDL cholesterol in HD patients.     

Effects of intraperitoneal hyaluronan on peritoneal fluid and solute transport in peritoneal dialysis patients.

BACKGROUND: Hyaluronan (HA) is a glycosaminoglycan found in connective tissues and tissue spaces, including the peritoneal cavity. In vivo studies in a rat model of peritoneal dialysis (PD) have shown that addition of HA to PD solution during an intraperitoneal dwell can alter peritoneal fluid transport and protect the peritoneal membrane from the effects of inflammation and repeated infusions of dialysis solution. The current study sought to evaluate the safety of intraperitoneal HA and its effect on peritoneal fluid and solute transport when administered during a dialysis dwell in humans. METHODS: 13 PD patients were enrolled in a prospective, randomized crossover study involving three dialysis treatments using the following PD solutions: (1) a commercially available PD solution (Dianeal PD-4, 1.36% glucose; Baxter Healthcare Corporation, Alliston, Ontario, Canada); (2) Dianeal PD-4 containing 0.1 g/L HA, and (3) Dianeal PD-4 containing 0.5 g/L HA. Each 6-hour dialysis exchange was separated from the other exchanges by a 2-week washout period. Radioiodinated human serum albumin (RISA) was administered with the dialysis solution to evaluate intraperitoneal volume, net ultrafiltration (UF), and fluid reabsorption. Peritoneal clearances, dialysate/plasma ratios (D/P), and mass transfer area coefficients (MTACs) were determined for sodium, urea, creatinine, albumin, and glucose. Safety was evaluated by monitoring adverse events and changes in serum chemistries. Ten patients completed all three dialysis exchanges and two additional patients completed at least one treatment exchange. RESULTS: There were no reported adverse events related to HA administration and no significant changes in serum chemistries. There were no significant differences in net UF or peritoneal volume profiles among the three treatments. Mean net UF calculated using residual volumes, estimated by RISA dilution, tended to be slightly higher during treatment with solution containing 0.1 g/L HA and 0.5 g/L HA [74 +/- 86 (SE) and 41 +/- 99 mL, respectively] compared to control treatment (-58 +/- 129 mL). Although not statistically significant, there was a trend toward decreased fluid reabsorption during treatment with HA. Solute clearances, D/P ratios, and MTACs were similar for the three treatments. Serum levels of HA were also unaffected by the two treatment solutions. CONCLUSIONS: These data support the acute safety of HA when administered intraperitoneally with the dialysis solution to PD patients. Due to the small sample size and variability in net UF and fluid reabsorption, statistically significant differences were not demonstrated for these parameters. However, a trend toward decreased fluid reabsorption was observed, suggesting that HA may act by a mechanism similar to that observed in animal studies. Further studies are necessary to evaluate whether the beneficial effects of HA observed in animal studies can be shown in humans.     

多重聚合酶链反应检测革兰阳性／阴性细菌、真菌和mecA基因的临床评价

目的：评价多重聚合酶链反应（PCR）直接检测革兰阳性／阴性细菌、真菌和耐甲氧西林葡萄球菌（MRS）的可靠性和准确性。方法：采用细菌16rRNA基因通用引物、革兰阴性细菌特异引物、全真菌通用引物和mecA基因特异引物，通过多重PCR对271份临床无菌性体液扩增检测革兰阳性／阴性细菌、真菌和MRS，同时对这些标本进行细菌、真菌培养和甲氧西林对葡萄球菌MIC测定。结果：多重PCR诊断无菌性体液体革兰阳性／阴性细菌、真菌和MRS感染的特异性、灵敏度、阳性预测值、阴性预测值和诊断效率分别为98.5％、97.0％、97.0％、99.0％、98.2％、99.1％、95.9％、95.9％、99.1％、98.5％、99.6％、100％、83.3％、100％、99.6％、100％、48.0％、100％、61.8％、71.7％。结论：多重PCR直接检测无菌性体液中革兰阳性／阴性细菌、真菌和mecA基因所致的MRS具有敏感、快速、特异、准确的优点，是一种可靠的实验诊断手段，但不能代替培养法。