Cerebrospinal fluid and serum IL-8, CCL2, and ICAM-1 concentrations in astrocytic brain tumor patients.

BACKGROUND: The aim of the study was the evaluation of serum and CSF concentrations of CCL2, IL-8, and sICAM-1 in patients with astrocytic tumors as compared to a group of non-tumoral patients. METHODS: Chemokine concentrations were measured using the ELISA method. RESULTS: Regardless of the parameter tested and the patient group (brain tumor or non-tumoral patients), statistical differences (P < 0.05) were found between concentrations obtained in CSF compared to values obtained in serum for all proteins tested. CSF IL-8 concentrations were significantly elevated in CNS tumor patients as compared to non-tumoral individuals (P = 0.000); serum CCL2 and sICAM-1 concentrations were significantly decreased in CNS tumors in comparison with the comparative group (P = 0.002 and P = 0.026, respectively). Among proteins tested in the serum, a higher area under the ROC curve (AUC) revealed CCL2 compared to sICAM-1 in differentiating subjects with CNS brain tumors from non-tumoral subjects. AUC for CSF IL-8 was higher than for its index (CSF IL-8/serum IL-8). CONCLUSIONS: For individual biomarkers (IL-8 and CCL2, sICAM-1), measured in CNS brain tumor patients, the appropriate material, respectively CSF or serum, should be chosen and quantitatively tested. Increased cerebrospinal fluid IL-8 with decreased serum CCL2 create a pattern of biomarkers, which may be helpful in the management of CNS astrocytic brain tumors.

Application of EPR dosimetry in bone for ex vivo measurements of doses in radiotherapy patients.

In the present study, bone samples from three patients treated in radiotherapy facilities in Poland were used for the determination of doses absorbed during radiotherapy. The samples were obtained during surgical treatments of patients performed due to medical indications. For the purpose of retrospective dosimetry, sensitivity of the radiation-induced EPR signal was individually calibrated in the samples by re-irradiation of the samples with known doses. The doses reconstructed in bones extracted within 6 months after irradiation were consistent with those calculated by treatment planning systems. The dose reconstructed in the bone removed 6 y after radiotherapy was ∼14% lower than the calculated one.

Antioxidant capacity in renal transplant patients.

The aim of the study was to analyse the relation between total antioxidant capacity and immunosuppressive therapies, renal function and hematocrit in kidney transplant patients. The study included 46 adult patients during the maintenance period (>1 year) following renal transplantation, treated with different combinations of immunosuppressive agents--most commonly with cyclosporine (n = 23) or tacrolimus (n = 15). The total antioxidant capacity (TAOC) of plasma was measured using Trolox-equivalent antioxidant capacity (TEAC) assay. Patients treated with cyclosporine had significantly greater TAOC compared with those treated with tacrolimus (1.16 +/- 0.46 mmol/L vs. 0.80 +/- 0.37 mmol/L, p = 0.018, respectively). There was a significantly negative correlation between TAOC and plasma creatinine (rs = -0.551, p = 0.033) and a positive correlation between TAOC and creatinine clearance or hematocrit in patients treated with tacrolimus but not with cyclosporine (r = 0.525, p = 0.045 or rs = 0.629, p = 0.012, respectively). Immunosuppressive therapy with cyclosporine was associated with higher TAOC. Anemia can be an independent risk factor for an increase of oxidative stress. Although subject numbers werelimited, TAOC was positively associated with renal function in patients treated with tacrolimus.

The value of tumor marker and cytokine analysis for the assessment of regional lymph node status in cervical cancer patients.

The aim of the study was to evaluate the utility of the measurements of the circulating tumor markers, squamous cell carcinoma antigen (SCCA), CA125, carcinoembryonic antigen (CEA), cytokeratin fragment 19 (CYFRA 21.1), and the cytokines, interleukin-6 and vascular endothelial growth factor (VEGF), to estimate regional lymph node involvement in patients with cervical cancer. The study comprised 182 untreated patients with cervical cancer. The regional lymph node status was assessed either by the postsurgical histopathologic examination or by the computed tomography (CT). Concentrations of SCCA, CEA, and CA125 were determined using the Abbott Instruments system, of CYFRA 21.1 by the Roche kits, and of IL-6 and VEGF by the ELISA of R&D Systems (Minneapolis, MN). For the statistical analyses, Mann-Whitney U test and chi(2) test were applied. Serum levels of SCCA, CEA, CA125, CYFRA 21.1, IL-6, and VEGF were measured in patients with specified pelvic and para-aortic lymph node status. SCCA, CA125, and IL-6 levels were found to be significantly higher in patients with lymph node metastases than in those with no lymph node involvement. Also, the percentage of patients with simultaneously elevated concentrations of SCCA and CA125 or SCCA and IL-6 differed depending on the lymph node status and was significantly higher in the series of patients with lymph node metastases. Simultaneous assessment of serum levels of SCCA and CA125 or SCCA and IL-6 in patients with cervical cancer may be useful for the regional lymph node evaluation, especially in patients with advanced stages, when the lymph nodes are examined only by CT, with no histologic confirmation.

Peritoneal fluid cytokines and the differential diagnosis of benign and malignant ovarian tumors and residual/recurrent disease examination.

This study aimed to assess the potential value of peritoneal fluid cytokine examination for the differential diagnosis of ovarian tumors and for evaluating residual or recurrent disease after treatment. The cytokines that are commonly elevated in ovarian cancer, VEGF, IL-6, bFGF, IL-8 and M-CSF, and a reference ovarian tumor marker, CA 125, were measured in peritoneal fluids of 53 previously untreated patients with epithelial ovarian cancer, 18 ovarian cancer patients after surgical treatment and chemotherapy, and 17 patients with benign epithelial ovarian tumors. Non-parametric statistical analysis of data was performed. Ovarian cancer peritoneal fluids, as compared to peritoneal fluids of patients with benign ovarian tumors, contained significantly higher concentrations of IL-6, VEGF and CA 125, and significantly lower concentrations of bFGF and M-CSF, but only the levels of IL-6 and VEGF were significantly higher in peritoneal fluids of stage I and II ovarian cancer patients than of patients with benign ovarian conditions. IL-6 at the cutoff level of 400 pg/mL discriminated benign and malignant ovarian tumors with 92% sensitivity and 60% specificity, while VEGF at the cutoff of 400 pg/mL had 90% sensitivity and 80% specificity. At the cutoff level of 1200 pg/mL, IL-6 had 84% sensitivity and 87% specificity. A radical decrease in local cytokine and CA 125 levels in patients after treatment was independent of therapy outcome. IL-6 and VEGF measurements in peritoneal fluids might be useful for the differential diagnosis of malignant and benign ovarian conditions, but not for residual or recurrent disease examination.

Clinical significance of serum cytokine measurements in untreated colorectal cancer patients: soluble tumor necrosis factor receptor type I--an independent prognostic factor.

The aim of this study was to exploit the potential clinical use of circulating cytokine measurements in colorectal cancer (CRC) patients. The levels of cytokines and cytokine receptors were assessed by ELISA in the sera of 50 healthy volunteers and 157 patients with previously untreated CRC and then related to clinicopathological features and prognosis. All tumors were verified histologically as colorectal adenocarcinomas and staged according to TNM classification. The levels of circulating interleukin (IL)-6, IL-8, macrophage colony-stimulating factor (M-CSF) and interleukin 1 receptor antagonist (IL-1ra) significantly increased with the clinical stage of CRC, and the levels of IL-6, soluble tumor necrosis factor (sTNF) receptor type I (RI), soluble interleukin 2 receptor alpha and TNFalpha with tumor grade, while IL-6, IL-8, M-CSF, IL-1ra and sTNF RI levels significantly rose with bowel wall invasion. None of the cytokine or soluble cytokine receptor levels were influenced by age, gender and colon versus rectum localization. sTNF RI, IL-8, IL-6 and vascular endothelial growth factor measurements demonstrated the highest diagnostic sensitivity. sTNF RI was found elevated in the greatest percentage of all CRC patients, in the greatest proportion of stage I patients and presented the best diagnostic sensitivity. In addition, the sTNF RI level strongly correlated with tumor grade and invasion and proved to be an independent prognostic factor.

Cytokines and endometriosis.

INTRODUCTION: Abnormal expression of cytokines is considered to be involved in the pathogenesis of endometriosis. MATERIAL AND METHODS: In 63 women with various stages of endometriosis preoperative levels of cytokines, including IL-6, IL-8, BVEGF, b-FGF, of TNF RI and TNF RII were analysed employing ELISA tests (R&D kits). RESULTS: In the sera of endometriosis patients significantly augmented levels of IL-6 and of b-FGF were detected as well as a trend for elevated IL-8 levels. CONCLUSION: Elevated serum levels of cytokines may show poor correspondence to the localized pathological process. Endometriosis would find a stricter reflection in cytokine levels in fluids or tissues of the pelvis minor.

Chemical modifications of alpha1,6-fucosyltransferase define amino acid residues of catalytic importance.

alpha1,6-Fucosyltransferase (alpha6FucT) of human platelets was subjected to the action of phenylglyoxal (PLG), pyridoxal-5'-phosphate/NaBH(4) (PLP), and diethyl pyrocarbonate (DEPC) the reagents that selectively modify the structure of amino acids arginine, lysine and histidine, respectively, as well as to N-ethylmaleimide (NEM), mersalyl, p-chloromercuribenzoate (pCMB), iodoacetate, iodoacetamide, and methyl iodide that react with sulfhydryl group of cysteine. In addition, we treated the enzyme with beta-mercaptoethanol, a reagent that disrupts disulfide bonds. All reagents except NEM significantly inactivated alpha6FucT. Protection against the action of PLG, PLP and sulfhydryl modifying reagents was offered by GDP-fucose, GDP, and the acceptor substrate, a transferrin-derived biantennary glycopeptide with terminal GlcNAc residues. Neither donor nor acceptor substrate offered, however, any protection against inactivation by DEPC or beta-mercaptoethanol. We conclude that arginine, cysteine and probably lysine residues are present in, or closely by, the donor and acceptor substrate binding domains of the enzyme, whereas histidine may be a part of its catalytic domain. However, the primary structure of alpha6FucT does not show cysteine residues in proximity to the postulated GDP-fucose-binding site and acceptor substrate binding site of the enzyme that contains two neighboring arginine residues and one lysine residue (Glycobiol. 10 (2000) 503). To rationalize our results we postulate that platelet alpha6FucT is folded through disulfide bonds that bring together donor/acceptor-binding- and cysteine- and lysine-rich, presumably acceptor substrate binding sites, thus creating a catalytic center of the enzyme.

Neutrophils promote the release of alpha-6-fucosyltransferase from blood platelets through the action of cathepsin G and elastase.

Human blood platelets release alpha-6-fucosyltransferase during coagulation of blood or after stimulation with thrombin or other agonists that cause platelet activation (Antoniewicz et al., FEBS Lett. 244 (1989) 388-390). However, in the absence of neutrophils the thrombin-stimulated platelets release only a small fraction of alpha-6-fucosyltransferase activity (Koscielak et al., Acta Biochim. Polon. 42 (1995) 35-40). We show that the effect of neutrophils is reproduced by cathepsin G or (less efficiently) by elastase, the two enzymes that are released by neutrophils during coagulation of blood. We have also localized alpha-6-fucosyltransferase to membrane and alpha-granule fractions of platelets that had been disrupted by nitrogen cavitation. It is concluded that thrombin-activated neutrophils release cathepsin G and elastase that promote degranulation of platelets and hence the secretion of alpha-6-fucosyltransferase.

A novel generation of coupling reagents. Enantiodifferentiating coupling reagents prepared in situ from 2-chloro-4,6-dimethoxy-1,3,5-triazine (CDMT) and chiral tertiary amines.

Coupling of racemic N-protected amino acids with amino components by means of 2-chloro-4,6-dimethoxy-1,3,5-triazine (CDMT) in the presence of chiral tertiary amines such as strychnine, brucine, and sparteine proceeds enantioselectively, affording appropriate amides or dipeptides in 69-85% yield. The configuration of the preferred enantiomer and enantiomeric enrichment depend on the structures of the amine and carboxylic acid. Calculated Kagan enantioselectivity parameters (s) are in the range 1.6-195. Chiral triazinylammonium chlorides formed in situ from CDMT and chiral tertiary amines are postulated as reactive intermediates involved in the process of enantioselective activation of N-protected amino acids.

Assessment of the musculoskeletal load of the trapezius and deltoid muscles during hand activity.

The purpose of the study was to analyse the influence of the precision of a task on tension and fatigue of the trapezius and deltoid muscles. Ten young men took part in experiments. Different levels of force and different frequencies of pressing a button defined the precision of the task. Surface electromyography (EMG) was used. Muscle tension and fatigue were reflected by 2 parameters of the EMG signal: the Root Mean Square amplitude related to the maximum value and changes in the Median Power Frequency. The results showed that hand activities influence the descending part of the trapezius muscle and do not influence the deltoid muscle, and that the precision of work can influence the examined muscles of the arm and shoulder even during work in which only the hand is involved in a performed task.

WW domains of Rsp5p define different functions: determination of roles in fluid phase and uracil permease endocytosis in Saccharomyces cerevisiae.

Rsp5p, ubiquitin-protein ligase, an enzyme of the ubiquitination pathway, contains three WW domains that mediate protein-protein interactions. To determine if these domains adapt Rsp5p to a subset of substrates involved in numerous cellular processes, we generated mutations in individual or combinations of the WW domains. The rsp5-w1, rsp5-w2, and rsp5-w3 mutant alleles complement RSP5 deletions at 30 degrees. Thus, individual WW domains are not essential. Each rsp5-w mutation caused temperature-sensitive growth. Among variants with mutations in multiple WW domains, only rsp5-w1w2 complemented the deletion. Thus, the WW3 domain is sufficient for Rsp5p essential functions. To determine whether rsp5-w mutations affect endocytosis, fluid phase and uracil permease (Fur4p) endocytosis was examined. The WW3 domain is important for both processes. WW2 appears not to be important for fluid phase endocytosis whereas it is important for Fur4p endocytosis. In contrast, the WW1 domain affects fluid phase endocytosis, but it does not appear to function in Fur4p endocytosis. Thus, various WW domains play different roles in the endocytosis of these two substrates. Rsp5p is located in the cytoplasm in a punctate pattern that does not change during the cell cycle. Altering WW domains does not change the location of Rsp5p.

Alterations of routine blood tests in adult patients with soft tissue sarcomas: relationships to cytokine serum levels and prognostic significance.

BACKGROUND: It has been reported that malignancy is often accompanied by hematological alterations and that such alterations may correlate with poor prognosis. It has also been demonstrated that several cytokines may be synthesized by many malignant tumors and that elevated serum levels of some cytokines are associated with changes in blood cell counts in cancer patients. However, so far little is known about the prognostic significance and mechanism of hematological changes in soft tissue sarcomas. The aim of the study was to evaluate the routine blood tests of disturbances in patients with malignant soft-tissue tumors prior to treatment and to correlate these results with selected cytokine serum levels, clinicopathological features of the tumors and patient survival. PATIENTS AND METHODS: 145 patients (75 males, 70 females; mean age 49.97 +/- 16.9 yrs) with histologically confirmed soft tissue sarcomas before treatment were enrolled into the study. In all these patients we evaluated routine blood tests (hemoglobin level HGB, white blood cell count WBC, platelet count PLT, white blood cell differential count-neutrocyte count NE, lymphocyte count LY, monocyte count MN, eosinophile count EO) and serum levels of 13 cytokines and soluble cytokine receptors (IL-6, IL-8, IL-10, TNFalpha, G-CSF, M-CSF, bFGF, VEGF, IL-1ra, sIL-2R. sIL-6R. TNF RI, TNF RII)--ELISA method. Peripheral blood samples from 50 healthy volunteers served as control. Statistical analysis was performed using Kolmogorov-Smirnov and Mann-Whitney U-tests, chi2 test (P < 0.05), where appropriate. For survival analysis the Kaplan-Meier method, log-rank test and multivariate Cox analysis were applied. RESULTS: Alterations of at least one of the standard blood tests were found in 43.4% of all cases. The most frequent alterations were: neutrophilia (28.3% of cases), leukocytosis (27.6%), decreased HGB (25.5%), monocytosis (19.3%) and thrombocytosis (14.5%); they correlated strongly with elevated serum levels of several cytokines and soluble cytokine receptors (particularly: sIL-2R, IL-6, IL-8, M-CSF, VEGF, TNF RI, TNF RII) (P < 0.001). Lymphocytopenia (LY < 1.0) found in 10.3% of patients correlated strongly with increased serum levels of IL-6, sIL-2R, TNF RI. In parallel, we found a significant difference in serum levels of 11 of 13 cytokines (IL-1ra. sIL-2R, IL-6, IL-8, IL-10, TNF RI, TNF RII, TNFalpha, M-CSF, bFGF, VEGF) (P < 0.001) in soft tissue sarcoma patients compared to healthy controls. Hematological alterations were significantly more frequent in patients with advanced tumors. In multivariate analysis we found no prognostic significance of any of the routine blood tests in soft tissue sarcoma patients. CONCLUSION: The results of this study demonstrate that hematological alterations, which occur in over 40% of soft tissue sarcoma cases, are found more frequently in patients with advanced tumors. Strong correlations between the occurrence of hematological abnormalities and elevated serum levels of several cytokines and soluble cytokine receptors, suggest that the former may develop as a result of cytokine misbalance frequently detected in soft tissue sarcoma patients. However, the results of routine blood tests alone are no independent prognostic factor for survival of soft-tissue sarcoma patients.

Practical application of ergonomic settings of typical computerised workstations.

The goal of the study was to check, with regard to ergonomics, workstations equipped with visual display terminals in selected enterprises. Over 180 workstations were tested in 3 enterprises. Most workstations were equipped with computers. The ergonomics of both the parameters of the basic components of the workstation (i.e., a chair and a desk, and the position of the computer at the workstation and its screen with respect to windows) and lighting fittings were analysed. Typical mistakes in the layout of a workstation were chairs inappropriate for computer work, as well as broken chair adjustment mechanisms, which qualified chairs for repair or replacement. Wrong positioning of monitors on the desk and with regard to windows and lighting fittings was also noted.

CRP, TNF-alpha, IL-1ra, IL-6, IL-8 and IL-10 in blood serum of colorectal cancer patients.

Blood serum cytokines: TNFalpha, IL-1ra, IL-6, IL-8, IL-10 as well as CRP were investigated in patients with colorectal cancer, prior treatment and 1, 10 and 42 days after surgery. There was an increase of the levels of CRP, IL-6 and IL-10 in most patients 24 hours after surgery. The levels of IL-1ra were elevated in patients in stage C and in several patients in stage B of the disease and there was a decrease of circulating TNFalpha in stage B patients. On day 10 and 42 after surgery, the levels of cytokines followed various patterns.

The growth of mdp1/rsp5 mutants of Saccharomyces cerevisiae is affected by mutations in the ATP-binding domain of the plasma membrane H+ -ATPase.

Mutations in the PMA1 gene, encoding plasma membrane H+ -ATPase, were isolated that are able to suppress the temperature sensitivity (ts) phenotype of mdp1 mutations located in RSP5, the ubiquitin-protein ligase gene. The mdp1 mutants were previously found to change the mitochondrial/cytosolic distribution of Mod5p-I, the tRNA modifying enzyme, and to affect fluid phase endocytosis. The data presented reveal that mdp1 mutants are also pH sensitive, and hypersensitive to hygromycin B and paromomycin. The ts phenotype, hygromycin B and paromomycin sensitivity are suppressed by pmal-t, but the pH sensitivity, the effect of mdp1 on Mod5p-I cytoplasmic/mitochondrial localization and endocytosis are not. Characterization of pmal-t revealed the substitution of amino acid G(653)V in the ATP-binding domain of the H+ -ATPase. Our results indicate that Rsp5 ubiquitin-protein ligase may also influence, in addition to protein distribution, the functioning of plasma membrane H+ -ATPase and the response of cells to stress.

Triazine dyes as inhibitors and affinity ligands of glycosyltransferases.

The triazine dyes: Cibacron Blue 3GA, Reactive Red 120, Reactive Yellow 86, Reactive Green 19, Reactive Blue 4, Reactive Brown 10 inhibited the activity of a purified preparation of alpha1,6fucosyltransferase (GDP-L-fucose: N-acetyl beta-glucosaminide 6-alpha-L-fucosyltransferase, EC 2.4.1.68) from human blood platelets. Cibacron Blue 3GA and Reactive Red 120 were examined for the nature of the inhibition and both were found to be competitive inhibitors of the enzyme, with Ki = 11 microM and 2 microM, respectively. The two dyes inhibited also serum glycosyltransferases: alpha1,2fucosyltransferase (GDP-L-fucose: beta-D-galactosyl-R2-alpha-L-fucosyltransferase, EC 2.4.1.69), beta1,4galactosyltransferase (UDP-galactose: N-acetyl-D-glucosamine 4-beta-D-galactosyltransferase, EC 2.4.1.90) and beta1,3N-acetylglucosaminyltransferase (UDP-GlcNAc: 4-beta-D-galactosyl-D-glucose). Cibacron Blue 3GA was a more effective inhibitor of the glycosyltransferases that use UDP-linked sugar donors than Reactive Red 120 while the latter was a stronger inhibitor of the fucosyltransferases that use GDP-linked donor. All four glycosyltransferases could be affinity purified on Cibacron Blue 3GA-Agarose columns. The order of elution of glycosyltransferases from the columns with solutions of 0.25-1.0 M potassium iodide also depended upon the structure of nucleotide sugar donor, i.e. whether it contained UDP or GDP. Thus, triazine dyes should interact with the sugar donor binding sites of glycosyltransferases. The main advantages of the use of triazine dyes as affinity ligands for isolation of glycosyltransferases are their universal applicability regardless of enzyme specificity, low cost, and insensitivity to high concentration of other proteins present in the solution.

Purification and characterization of GDP-L-Fuc: N-acetyl beta-D-glucosaminide alpha1-->6fucosyltransferase from human blood platelets.

c-6-L-Fucosyltransferase (alpha1,6FucT; EC 2.4.1.68) from human platelets, the enzyme that is released into serum during coagulation of blood, was purified 100,000-fold. The purification required three sequential chromatographic steps: chromatofocusing, affinity column chromatography on GnGn-Gp(asialo-aglacto-transferrin glycopeptide)-CH-Sepharose, and gel filtration of Sephadex G-200. The final preparation contained a protein that migrated as a single discrete band Mr of 58,000 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under non-reducing conditions, and as a single enzymatically active peak Mr of 58,000 in gel filtration. Although the purified enzyme utilized the biantennary GnGn-Gp as substrate, it was twice as active with the triantennary oligosaccharide when the Man alpha1,3 antenna was substituted with GlcNacbeta1,4. On the other hand the tetraantennary oligosaccharide was not a preferred substrate. The Km values for the substrate asialo-agalactotransferrin-glycopeptide, and GDP-L-fucose were 29 and 28 microM, respectively. The optimum pH of the enzyme was 6.0. The activity of alpha1,6FucT was abolished in the presence of beta-mercaptoethanol. Divalent cations such as Mg2+ and Ca2+ activated, but Cu2+, Zn2+ and Ni2+ strongly inhibited the activity.

[Frequency of human papillomavirus-herpes simplex-cytomegalovirus and Chlamydia trachomatis infection in genital tracts of sexual partners]. / Czestosc wystepowania infekcji wirusowych brodawczaka ludzkiego, opryszczki plciowej, cytomegalii oraz chlamydia trachomatis w narzadach plciowych partnerów seksualnych.

OBJECTIVES: Much attention is given nowadays to the role of Human Papillomavirus, Herpes simplex virus, Cytomegalovirus, and Chlamydia trachomatis--infections in cervical carcinogenesis. Cytomegalovirus, Herpes simplex and Chlamydia trachomatis are now thought to be teratogenic to humans. DESIGN: We investigated the prevalence of HPV, HSV, CMV and Chlamydia trachomatis in genital tracts of sexual partners. MATERIALS AND METHODS: 90 sexual partners were qualified for the research. Examination smears were taken with the dacron swab from the vaginal part of the uterine cervix, cervical canal, the lower vagina from women and from fossa navicularis penis in men. In the group of 67 men we have investigated semen as well. HPV, HSV, CMV and Chlamydia trachomatis were identified using PCR (Polymerse Chain Reaction)--method. RESULTS AND CONCLUSIONS: In 48% of investigated sexual partners we proved the presence of Human Papillomavirus, in 2.2% of women and 2.9% of men--Cytomegalovirus and in 11.1% of women and 14.9% of men--Chlamydia trachomatis. In the investigated biological material we did not find any HSV infection.