Prevalence of hypophosphatemia in sepsis and infection: the role of cytokines.

BACKGROUND: Sepsis occurs following the presence of bacteria in the circulation and is associated with fever, hyperthermia, and hypotension. Hypophosphatemia develops in the early stages of sepsis. High levels of inflammatory cytokines also characterize early sepsis. AIM: The aim of the present study was to correlate hypophosphatemia with cytokines and cytokine receptor levels during early sepsis. We aimed to reestablish the results obtained from patients in an in vivo experimental model, in order to understand the mechanism of hypophosphatemia induction in early sepsis. METHODS: Ninety-nine patients were enrolled in this study and their clinical condition was classified as the presence of infection, sepsis, and bacterial growth in blood cultures. Phosphate levels and cytokine levels were recorded. In order to determine whether hypophosphatemia is correlated to the increased inflammatory cytokines, we injected normal mice with recombinant cytokines and studied their effect on phosphate levels. RESULTS: Our results revealed that 80% of the septic patients had hypophosphatemia associated with very high levels of tumor necrosis factor (TNF)alpha and interleukin (IL)-6 and of soluble IL receptor (sIL)-2R and IL-6R, especially in those patients with positive blood cultures. Injection of IL-6, TNFalpha and IL-1beta in mice markedly decreased the phosphate serum levels. CONCLUSIONS: Significant associations were demonstrated between high levels of inflammatory cytokines and their receptors and between serum phosphate levels, especially in patients with positive blood culture. Our results point to a correlation between the high inflammatory cytokines levels and hypophosphatemia during early sepsis. Cytokine levels and hypophosphatemia may be included in sepsis evaluation and prognosis. Anticytokine strategies might, therefore, reverse hypophosphatemia and other parameters of sepsis.

Elevated inflammatory cytokine levels in bone marrow graft rejection.

Graft rejection and graft failure represent major obstacles in allogeneic bone marrow transplantation (BMT). Cytokines possibly play a central role in the inflammatory and allospecific components of allograft rejection. Therefore, we evaluated inflammatory cytokine levels following BMT in 12 consecutive patients with graft rejection (GR). Seven of the patients underwent BMT from siblings (6 matched and 1 mismatched), 4 patients received bone marrow from other family members (3 mismatched and 1 matched), and 1 patient underwent HLA-matched unrelated BMT. Nine of 12 had a sex-mismatched BMT and 5/12 had an ABO-mismatched BMT. Nine of 12 underwent T cell-depleted (Campath anti-CDw52 moAb) BMT. Rejection was defined as marrow hypoplasia with a peripheral white blood cell count < 0.5 x 10(9)/L 21 days after BMT, in conjunction with the absence of donor cells by polymerase chain reaction analysis using a sex-mismatched probe and/or a tumor-specific probe (BCR/ABL). Twenty-five patients who underwent uneventful BMT with no GR served as controls. The levels of tumor necrosis factor (TNF), interleukin-6 (IL-6), and interleukin-1 (IL-1) were evaluated by a high sensitive RIA or an enzyme immunoassay. The levels of TNF and IL-6 were found to be higher in 10/12 and 7/7 evaluated GR patients, respectively, as compared with controls (P < 0.05). The level of IL-1 was high only in 2/12 patients. TNF elevation occurred in all patients immediately after GR. TNF and IL-6 levels were significantly higher for patients with early rejection (< 35 days after BMT) as compared with patients with late rejection (> 35 days after BMT) (P < 0.049 and P < 0.006, respectively). Eight patients engrafted after the second transplant (2 only transient). All 6 patients with stable engraftment are alive (4 with basic disease), while the 4 patients who did not engraft and the 2 patients with only transient engraftment died. In the 6 patients with no engraftment or only transient engraftment, the elevated TNF levels remained high; in the 6 patients who had stable engraftment after retransplant, TNF levels, but not IL-6 levels, decreased. In conclusion, a majority of the patients with GR displayed high levels of inflammatory cytokines (TNF and IL-6). Dysregulation of inflammatory cytokines may be involved in the pathogenesis of GR.

Effect of cycloheximide on ionic channels in neuroblastoma cell membrane.

The effect of cycloheximide (an inhibitor of protein synthesis) on the ionic currents through sodium and potassium channels was investigated in dialysed voltage-clamped N18 A-1 neuroblastoma cells. The cycloheximide concentration needed for half-inhibition of sodium peak conductance was about 0.5 micrograms/ml for 24 h of incubation. Half-inhibition time of the sodium peak conductance in cells incubated with 15.0 micrograms/ml of cycloheximide was about 9 h. Sodium against potassium ion selectivity, the activation and inactivation parameters were shown to be not affected by cycloheximide. Potassium conductance in similarly treated cells exhibited no consistent changes. The main conclusion is that the decay in peak sodium conductance is caused by diminishing the sodium channel density in the membrane (from 25 to 2.2 channels per micron2). The inhibition effect was evidently mediated by block of protein synthesis and was not the result of direct drug-channel interaction. The half-decay time of sodium peak conductance is interpreted as a possible life-time characteristic of sodium channels in the neuroblastoma cell membrane.

Cytokine dysregulation in chronic graft versus host disease.

Cytokines play a key role in the pathogenesis of chronic Graft versus Host Disease (cGVHD) and various studies have shown abberant production of cytokines by immune cells from GVHD patients. Based on these findings and others showing that high TNF levels precede the development of GVHD, we evaluated inflammatory cytokine levels following BMT and during the development of cGVHD. In this study, patients undergoing bone marrow transplantation (BMT) who consequently developed chronic GVHD were analyzed as to their cytokine production during cGVHD and this was correlated with their clinical manifestations. A positive correlation was found between the severity as well as the number of major clinical complications and high levels of inflammatory cytokines (IL-1 beta IL-6 and TNF alpha) compared to control patients or to normal donors. Patients undergoing BMT who did not develop GVHD, did not produce high levels of IL-1 beta IL-6 or TNF. High levels of cytokines may be used as a tool for assessing novel therapeutic modalities and response to GVHD treatment.

Serum interleukin 1 beta levels as a marker in hairy cell leukemia: correlation with disease status and sIL-2R levels.

Hairy cell leukemia (HCL) is a chronic lymphoproliferative disorder of the B cell lineage. In the search for specific markers with prognostic significance we evaluated the clinical value of IL-1 beta and sIL-2R levels in HCL patients. HCL patients (25) were classified according to their clinical status as "active", "non active" disease or partial or complete remission and response treatment. We found a good correlation between IL-1 beta or IL-2R levels and disease activity: improved clinical status or response to different therapies were associated with decreasing IL-1 beta or sIL-2R levels. In contrast, lack of response to therapy or disease progression was reflected in increases in both IL-1 beta and sIL-2R levels. In some patients increases of both cytokines preceded clinical symptoms. In conclusion our results show that IL-1 beta and s-IL-2R levels may be used as reliable markers for monitoring HCL activity, assessing response to treatment and predicting early progression of disease.

Vascular endothelial growth factor plasma levels correlate to the clinical picture in severe ovarian hyperstimulation syndrome.

OBJECTIVE: To assess the potential involvement of vascular endothelial growth factor in the hyperpermeability characterizing the ovarian hyperstimulation syndrome (OHSS). DESIGN: A controlled clinical study that followed the kinetics of vascular endothelial growth factor in the plasma of patients with severe OHSS from the time of admission to the hospital and until clinical resolution. SETTING: Women hospitalized with severe OHSS in a tertiary medical center. PATIENT(S): Seven patients with severe OHSS after ovulation induction for IVF and seven controls who had received a similar ovulation induction regimen and did not develop the OHSS. INTERVENTION(S): Three blood samples were obtained from each OHSS patient: upon hospitalization for severe OHSS, when significant clinical improvement was evident, and on the first follow-up visit after the patients' discharge. Ascitic fluid was obtained from all OHSS patients by therapeutic paracentesis during the active phase of the syndrome. Blood samples were drawn from the control patients 4 to 6 days after ET. All samples were assayed for vascular endothelial growth factor levels, hematocrit, E2 levels, and white blood cell count. MAIN OUTCOME MEASURE(S): Vascular endothelial growth factor levels were assayed by ELISA. Estradiol was determined by RIA. RESULT(S): Compared with the controls, high levels of vascular endothelial growth factor were detected in the plasma of all patients admitted for severe OHSS. Levels dropped significantly along with clinical improvement, reaching minimum values after complete resolution. A statistically significant correlation was found between plasma vascular endothelial growth factor levels and certain biologic characteristics of OHSS and of capillary leakage such as leukocytosis and increased hematocrit. Ascitic fluid obtained from the study patients also contained high vascular endothelial growth factor levels. CONCLUSION(S): These findings suggest the involvement of vascular endothelial growth factor in the pathogenesis of capillary leakage in the OHSS.

Correlation of serum levels of interleukin-1 family members with disease activity and response to treatment in hairy cell leukemia.

Hairy cell leukemia (HCL) is a well-recognized chronic lymphoproliferative disorder of B cell lineage, which may be regulated by growth factors including cytokines and cytokine antagonists. Previous studies have shown a good correlation between circulating soluble interleukin-2 receptor (sIL-2R) levels and disease activity and response to therapy was always associated with a decrease in sIL-2R levels. The interleukin-1 (IL-1) family of agonists and antagonists may also be involved in the regulation of hematopoietic malignancies. In the present study, we evaluated members of the IL-1 family (IL-1beta, IL-1 receptor antagonist (IL-1Ra) and IL-1 soluble receptors Type I and Type II (IL-1sRI and IL-1sRII) in 23 patients with HCL. Patients were classified according to the clinical state of their disease. Most were treated with 2-chloro-2'-deoxyadercosine (2-CDA) and treatment was associated with a significant decrease in the serum levels of sIL-2R, IL-1beta and IL-1sRII in patients achieving a complete or partial response. In contrast to the above, levels of IL-1Ra increased during response to treatment and clinical response to 2-CDA was associated with an increase of 122% in IL-1Ra levels, in parallel with a decrease of 63% in IL-1beta and 47% in IL-1sRII levels. These results suggest that the balance between IL-1beta, IL-2 and their soluble receptors or antagonists may be involved in the pathogenesis and immunoregulation of HCL. Serum levels of these cytokines may therefore be used to monitor therapeutic efficacy of therapy in HCL and to detect any residual disease.

Cytokines in Gaucher's disease.

Gaucher's disease (GD) is characterized by hepatosplenomegaly, bone marrow infiltration, osteonecrosis, which may all be associated with the presence of pathological macrophages that contain undegraded glycosphingolipids. Levels of serum cytokines, which are soluble products of mononuclear phagocytes (MNP), were evaluated in 24 GD patients. Levels of interleukin-1beta (IL-1beta), interleukin-1 receptor antagonist (IL-1Ra), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), and soluble interleukin-2 receptor (sIL-2R) in GD patients were significantly higher than in normal controls. We attempted to correlate cytokine levels with disease severity. Type I GD patients with more severe clinical manifestations had significantly higher levels of IL-1beta, IL-1Ra and IL-6, relative to type I patients with milder disease. Three patients homozygous for the 1448C mutation with neuropathic type III disease, had significantly higher levels of sIL-2R than type I patients or controls. We speculate that cytokine over-expression may relate to the pathophysiology of some of the clinical manifestations of GD. Thus, the elevated IL-1beta, TNF-alpha and IL-6 levels may induce the bone manifestations, the neutrophil chemotaxis and the increased incidence of hyper-gammaglobulinemia present in GD patients.

Prognostic value of CYFRA 21-1, TPS and CEA in different histologic types of non-small cell lung cancer.

The prognostic value of the tumor markers CYFRA 21-1, tissue polypeptide specific antigen (TPS) and carcinoembryonic antigen (CEA) was investigated in three histologic subtypes of non-small cell lung cancer. Pretreatment serum marker levels were measured in 38 patients with adenocarcinoma (AC), in 43 patients with squamous cell carcinoma (SQC) and in 35 patients with large cell carcinoma (LCC). Univariate analysis in AC showed significant lower survival of patients with elevated levels of TPS, CYFRA 21-1 and CEA. In LCC, elevated levels of TPS and CEA were found to predict lower survival, while in SQC--only TPS was a predictor. A multivariate analysis of survival identified CEA (Relative Risk-5.5; p = 0.004), CYFRA 21-1 (RR-3.4; p = 0.008) and TPS (RR-3.0; p = 0.02) as independent prognostic factors in AC. In SQC, only TPS (RR-2.3; p = 0.03) was such a factor whereas in LC--none of the markers studied retained statistical significance. Thereafter, the combinations of the two strongest prognostic factors in the AC group--CEA and CYFRA 21-1 were explored to divide this group into subsets with different prognosis. In cases where both markers were positive, the relative risk of death was 10.5 times higher as compared to cases where both markers were negative (p = 0.002).

Cytokine evaluation in throat infections.

Inflammatory cytokines have been shown to play an important role in the pathogenesis of various inflammatory processes. In throat infections, intracellular inflammatory cytokines have been detected from the sites of inflammation. The present study aimed to evaluate serum cytokine levels of patients with throat infections and correlate them to the inflammatory parameters and type of inflammation. Significantly higher inflammatory cytokine levels (interleukin [IL]-6 > 7 pg/mL, IL-1 > 1 beta pg/mL, tumor necrosis factor alpha > 1 pg/mL) were detected in most of the patients as opposed to healthy controls. Clinical parameters of infection (fever > 38 degrees C, leukocytosis > 11,000 white blood cells per cubic millimeter, polymorphonuclear neutrophils > 75%) were significantly correlated with high levels of inflammatory cytokines: mainly IL-6 and tumor necrosis factor alpha, and to a lesser degree with IL-1 beta. No correlation, however, was found between the type of inflammation and cytokine levels. The present study indicates a role of inflammatory cytokines in the pathogenesis of throat infections and the need for an anti-inflammatory and anticytokine therapeutic approach.

Changes in interleukin-1 beta and soluble interleukin-2 receptor levels in CSF and serum of schizophrenic patients.

Some evidence points towards a possible autoimmune role in the aetiology of schizophrenia. Experimental findings provide contradictory results regarding abnormalities in cytokine production in this disorder. In the present study we tested the production of cytokines in CSF and serum in 16 schizophrenic patients and 10 healthy controls (tumor necrosis factor alpha - TNF alpha; interleukins IL-1 beta, IL-2, IL-6, soluble IL-2 receptor). Cytokine levels were evaluated by radioactively-labeled antibodies (IL-1 beta, IL-2, IL-6), by enzyme-linked immunoassay (TNF) and by a sandwich enzyme immunoassay (soluble IL-2 receptor). No significant differences were found in either CSF fluid or serum levels of TNF and IL-2 or IL-6. Interleukin-1 beta was significantly decreased in patients' CSF and serum as compared to controls. Soluble interleukin-2 receptor levels were decreased in CSF of patients, but highly increased in their serum in comparison with controls. Changes in various cytokine levels in CSF fluid and serum of schizophrenic patients probably reflect interrelated process of growth, degeneration or neuroimmunological abnormalities, which may all play a role in the pathophysiology of schizophrenia. The present study supports evidence for change in immune activation, probably of peripheral origin, in schizophrenic patients.

[Synthesis and secretion of recombinant protein--the product of v-sis oncogene in Saccharomyces cerevisiae cells]. / Sintez i sekretsiia rekombinantnogo belka--produkta onkogena v-sis v kletkakh drozhzhei Saccharomyces cerevisiae.

The recombinant plasmid DNA YEp secl-v-sis was constructed. This plasmid was able to code for the synthesis and secretion into the cultural medium of the protein-product of oncogene v-sis. Transcription, copy number and stability of the plasmid DNA were studied under the conditions of galactose induction. The v-sis protein was determined by gel electrophoresis and immunoblotting methods and assayed for cell-proliferation activity in the mammalian cell culture.

[General and primary thermoresistance in genetically differing variants of murine neuroblastoma]. / Obshchaia i pervichnaia teploustoichivost' u geneticheski razlichaiushchikhsia variantov neiroblastomy myshi.

General and primary thermoresistance of mouse neuroblastoma clonal cell lines derived from N18A subline was studied: the N18A1 clonal cell line was not treated by heat, the NTR1 was obtained by one-step selection for resistance to the long action of the temperature 40 degrees C, the NHSR1 was obtained by multistep selection for resistance to short-time treatment at 44 degrees C. The NHSR1 clonal line was shown to have higher general and primary thermoresistance by comparison with that of N18A1 cells. The NTR1 cell line, capable of unlimited proliferation at 40 degrees C, did not differ in general resistance but displayed a slower primary resistance compared to that in the N18A1 cells. Cells of all the three clones were found to be capable of temporary increasing in primary thermoresistance, i.e. hardening. A possible contribution of the primary resistance into the general one in cells of all the selected clones has been discussed.

[The isolation and phenotypic characteristics of heat-resistant cells from the Chinese hamster CHO-K1 line]. / Poluchenie i fenotipicheskie osobennosti termorezistentnykh kletok kitaiskogo khomiachka linii CHO-K1.

By sequential selection for a resistance to a short-term heating at 45 degrees C, two independent clones, CHSR5 and CHSR6, have been isolated from ovarian cells of the Chinese hamster (CHO-K1). A subline CHO-A40, capable of proliferation at 40 degrees C, has been obtained as mass culture by culturing CHO-K1 cells for 2 months at 40 degrees C. The lines obtained have no cross-resistance to the temperature used in their selection: the CHSR5 and CHSR6 cells are not capable of reproduction at 40 degrees C, while the CHO cells do not survive over a 60 min heating at 45 degrees C. This bears evidence that mechanisms, underlying the cell resistance under different thermal regimens used, may be different. The heat resistant sublines maintain their heat resistance for 6 months of cultivation at 37 degrees C. The acquisition of heat resistance by cells correlates with their high primary resistance, and with their resistance to colchicine and actinomycin D.

[The temperature dependence of protein kinase A and C activity in variant mouse neuroblastoma cells differing genetically in their heat resistance]. / Temperaturnaia zavisimost' aktivnosti proteinkinaz A i C u variantov kletok neiroblastomy myshi, geneticheski razlichaiushchikhsia po teploustoichivosti.

The ability of the mouse neuroblastoma cell line NTR to proliferate at 40 degrees C correlates with the position of the temperature optimum of protein kinases A and C activities in the region of higher temperatures compared to those for cells of the original line N18AI, and with higher thermostability of protein kinase A after its heating at various elevated temperatures. The found changes in protein kinases A and C in the cells of NTR line mean that the selection of variants, capable of growing at elevated temperatures, is accompanied with conformational protein changes.

[Phenotypic characteristics of heat-resistant murine neuroblastoma cells]. / Osobennosti fenotipa teploustoichivykh kletok neiroblastomy myshi.

One-step selection of murine neuroblastoma (N18TG2 cell line) for the resistance to injuring action of higher temperature resulted in obtaining cell line NTR1 stably capable of proliferating at 40 degrees C. The thermoresistance is shown to be coupled with the changes in some phenotypic features of NTR1 cells: the multiplication rate, saturation density of cells, morphological features, inducibility of long neurite-like processes and adhesion. A possible significance of plasma membrane changes in genetically stable thermoresistance of NTR1 neuroblastoma cells is discussed.

[Ionic currents of neuroblastoma clone N18 A-1 cultured cells under potassium fluoride and phosphate intracellular dialysis]. / Ionnye toki kul'tiviruemykh kletok neiroblastomy klona N18 A-1 v usloviiakh vnutrikletochnogo dializa ftoridom i fosfatom kaliia.

Ionic currents of cells of neuroblastoma clone N18 A-1 was studied under conditions when the internal medium was placed for artificial fluoride or phosphate solutions. The specific membrane leakage resistance was measured to be 8.1 +/- 2.6 kOhm.cm2 and 1.3 +/- 0.3 Kohm.cm2, respectively. The presence of usual sodium and tetraethylammonium sensitive potassium channels is demonstrated. Potassium conductance is shown to amount to 0.25--0.025 of sodium conductance. Dialysis of the cells by phosphate solutions induces a slow outward current, which is not inhibited by tetraethylammonium ions.

[Turnover of potential-dependent ion channels in the membrane of neuroblastoma cells studied using the protein synthesis inhibitor cycloheximide]. / Issledovanie vremeni obmena potentsialzavisimykh ionnykh kanalov v membrane kletok neiroblastomy s pomoshch'iu ingibitora belkovogo sinteza tsiklogeksimida.

The influence of cycloheximide on currents through sodium and potassium channels in dialysed neuroblastoma cells of clone N18A-1 has been studied under voltage clamp conditions. When cells are incubated with cycloheximide, the conductance of sodium decreases, whereas that of potassium changes only slightly, if at all. The cycloheximide concentration, at which sodium conductance is reduced by 50% during 24 hours incubation, was about 0.5 microgram/ml. The half-time of inhibition at the concentration of 15 microgram/ml was about 9 hours, with the time of 50% inhibition of sodium being more complex than single exponential. The density of sodium channels in control and after the maximal inhibition was estimated to be about 25 and 2.2 micron-2, respectively. The sodium-potassium selectivity and the gating mechanism parameters were shown to be unchanged following cycloheximide treatment.

[Changes in the surface and activation of circulating leukocytes after autologous transfusion of UV-irradiated blood]. / Izmeneniia poverkhnosti i aktivatsii tsirkuliruiushchikh leikotsitov pri autotransfuziiakh UF-obluchennoi krovi.

Under study was the dynamics of functional properties of leukocytes of healthy people and patients after UV-irradiation of their blood, mixing the UV-irradiated and non-irradiated blood. It was shown that UV-irradiation and mixing the irradiated and non-irradiated blood facilitated the liberation of bactericidal cation proteins of leukocytes, higher expression of membrane receptors, growth factors, activation of reparation of DNA after its injury by gamma-rays. It proves the role of immediate activation of leukocytes of the circulating blood in trigger mechanisms of effects of autotransfusion of UV-irradiated blood.

Prognostic role of serum cytokeratin 19 fragments in advanced non-small-cell lung cancer: association of marker changes after two chemotherapy cycles with different measures of clinical response and survival.

Prognostic implication of serum cytokeratin 19 fragments (CYFRA 21-1) was explored in 60 advanced NSCLC patients, whereas in 45 patients assessable for serological response a >or=35% CYFRA 21-1 decline after two chemotherapy cycles was strongly associated with non-progression (NP), defined as a sum of objective response (OR)+stable disease (P<0.0001) and survival (P=0.0002). Association of OR with survival was not significant. In multivariate survival analysis, >or=35% marker decline and radiological NP status were found as major determinants of prolonged survival with RR: 0.37 (P=0.01) and 0.63 (P=0.01), respectively. In advanced NSCLC patients, NP reflects therapeutic efficacy better than traditional OR. CYFRA 21-1 >or=35% decline seems to be a reliable surrogate marker of treatment efficacy in terms of survival.