Paraoxonase 1 activity and phenotype distribution in premenopausal and postmenopausal women.

INTRODUCTION: Postmenopausal women have higher risk of cardiovascular disease. One of the contributing factors could be reduced activity of anti-atherogenic enzyme paraoxonase 1 (PON1). The aim of this study was to examine differences in the lipid status, paraoxonase and arylesterase PON1 activities and PON1 phenotype in women with regular menstrual cycle and in postmenopausal women. MATERIALS AND METHODS: The study included 51 women in reproductive age (25 in follicular and 26 in luteal phase of the menstrual cycle) and 23 women in postmenopause. Lipid parameters in sera were determined using original reagents and according to manufacturer protocol. PON1 activity in serum was assessed by spectrophotometric method with substrates: paraoxon and phenylacetate. PON1 phenotype was determined by double substrate method. RESULTS: Compared to the women in follicular and luteal phase, postmenopausal women have significantly higher concentration of triglyceride [0.9 (0.7-1.3), 0.7 (0.6-1.0) vs. 1.5 (0.9-1.7) mmol/L; P = 0.002], cholesterol [5.10 (4.78-6.10), 5.05 (4.70-5.40) vs. 6.30 (5.73-7.23) mmol/L; P < 0.001], LDL [3.00 (2.56-3.63), 3.00 (2.70-3.70) vs. 3.90 (3.23-4.50) mmol/L; P < 0.001], and apolipoprotein B [0.88 (0.75-1.00), 0.79 (0.68-1.00) vs. 1.07 (0.90-1.24) mmol/L; P = 0.002]. PON1 basal [104 (66-260), 106 (63-250) vs. 93 (71-165) U/L; P = 0.847] and salt-stimulated paraoxonase activity [210 (131-462), 211 (120-442) vs. 180 (139-296) U/L; P = 0.857] as well as arylesterase activity [74 (63-82), 70 (54-91) vs. 70 (60-81) kU/L; P = 0.906] and PON1 phenotype (P = 0.810) were not different in the study groups. CONCLUSION: There are no differences in PON1 activity and PON1 phenotype between women with regular menstrual cycle and postmenopausal women.

Differences in routine laboratory parameters related to cachexia between patients with hematological diseases and patients with solid tumors or heart failure - is there only one cachexia?

BACKGROUND: Cachexia is a state of involuntary weight loss common to many chronic diseases. Experimental data, showing that cachexia is related to the enhancement of acute phase response reaction, led to the new definition of cachexia that included, aside from the principal criterion of weight loss, other "minor criteria", Amongst them are levels of C-reactive protein (CRP), albumin and hemoglobin. However, there is paucity of data regarding possible differences of these laboratory parameters in patients with various diseases known to be related to cachexia. METHODS: CRP, albumin and hemoglobin were evaluated in 119 patients, divided into two disease groups, hematological (ones with diagnosis of non-Hodgkin lymphoma or Hodgkin disease) and non-hematological (solid tumor patients and patients with chronic heart failure). Patients were further subdivided into two nutritional groups, cachectic and non-cachectic ones according to the principal criterion for cacxehia i.e. loss of body weight. RESULTS: We found that cachectic patients had higher levels of CRP, and lower levels of both hemoglobin and albumin compared to non-cachectic patients, regardless of the disease group they fitted. On the other hand, the group of hematological patients had lower levels of CRP primarily due to the differences found in the non-cachectic group. Higher levels of albumin were also found in the hematological group regardless of the nutritional group they fitted. Limitations of cut-off values, proposed by definition, were found, mostly regarding their relatively low sensitivity and low negative predictive value. CONCLUSIONS: As expected, differences in values of routine laboratory parameters used in definition of cachexia were found between cachectic and non-cachectic patients. Their values differed between hematological and non-hematological patients both in cachectic and non-cachectic group. Cut-off levels currently used in definition of cachexia have limitations and should be further evaluated.

Proteomics of inflammatory and oxidative stress response in cows with subclinical and clinical mastitis.

Cow serum proteome was evaluated by three different complementary approaches in the control group, subclinical and clinical mastitis in order to possibly find differential protein expression useful for a better understanding of the pathophysiology of mastitis as well as for an early diagnosis of the disease. The systemic inflammatory and oxidative stress response in cows with subclinical and clinical mastitis were observed. The collected evidence shows a differential protein expression of serpin A3-1, vitronectin-like protein and complement factor H in subclinical mastitis in comparison with the control. It was also found a differential protein expression of inter-alpha-trypsin inhibitor heavy chain H4, serpin A3-1, C4b-binding protein alpha chain, haptoglobin and apolipoprotein A-I in clinical mastitis compared to the control. Among the inflammatory proteins up-regulated in clinical mastitis, vitronectin is over-expressed in both subclinical and clinical mastitis indicating a strong bacterial infection. This suggests vitronectin as an important mediator in the pathogenesis of the onset of mastitis as well as a valuable marker for diagnosis of the subclinical form of the disease. Obtained data could be useful for the detection of mastitis during the subclinical phase and for a better comprehension of the pathophysiological mechanisms involved in the onset of the disease.

External quality assessment in clinical cell analysis by flow cytometry. Why is it so important?

Participation in external quality assessment is an integral part of laboratory work and mandatory when the results have a clinical application, which is one of the requirements of standard 15189 for accreditation of medical laboratories. Institute of Clinical Chemistry, the first laboratory accredited for clinical cell analysis by flow cytometry in Croatia, participated in UKNEQAS for Leukocyte Immunophenotyping in 3 schemes: "Immune Monitoring", "CD34 Stem Cell Enumeration" and "Leukaemia Immunophenotyping". For sample processing on EPICS XL flow cytometer, lyse/no wash preparation technique with ammonium chloride (NH4Cl) or ImmunoPrep lysing reagent was employed. In "Immune monitoring" programme CD45/sideward light scatter (SSC) proposed gating strategy was adopted for lymphocyte subsets, while modified ISHAGE protocol was used for CD34+ cell enumeration. Absolute count determination was performed on flow cytometer using FlowCount beads solution. In the period from the beginning of 2006 until the middle of 2009 a total number of 100 stabilized whole blood samples were processed. The relative and absolute enumeration results for lymphocyte subsets were within tolerable limits, in 97.1 and 97.1% of cases, and 95 and 90% of CD34+ cell enumeration, respectively. In immune monitoring CD45/SSC proposed gating strategy is the most frequent analysis used (> 85% participants) and ISHAGE protocol for CD34+ cell determination with continuous rise from 76 to 83%. A number of participants who accept beads method for absolute count enumeration on flow cytometer get greater, 69 to 86%, while FlowCount was the second of bead-based techniques used (25 and 35%). Sample treatment in lyse/no wash technique using NH4Cl lysing solution was dominant procedure used by more than 1/3 participants, although its home made solution has replaced slowly by commercial reagents. The unacceptable results, 6 of 244, were obtained for 20 most frequently determined cell antigens in "Leukaemia Immunophenotyping" samples screened for leukaemia/lymphoma. Processing results of all participants showed that the deviation from laboratory guidelines and the use of older methods for cell identification, quantification of cell counting on haematology analyser, or usage an antibody conjugated with fluorochrome lesser fluorescence quantum often lead to an unacceptable result, although is noticeable trend to accept new referrals and protocols to reduce the inter-laboratory differences.