Early expression of the aflatoxin gene cluster in Aspergillus nomiae isolated from Brazil nut.

Aspergillus nomiae is the most important contaminant in Brazil nut due to its high incidence in these nuts and its strong production of carcinogenic metabolites: aflatoxins (AF). Aflatoxin biosynthesis pathway in A. nomiae is poorly studied. Thus, in present investigation, aflatoxin production and gene cluster (aflC, aflQ, aflU, and aflX) expression profile were evaluated on two strains of A. nomiae isolated from Brazil nut samples collected in the Amazon region, and cultivated on Brazil nut-based medium. The results showed that the expression of the aflatoxin gene cluster in A. nomiae, started at day 2 and occurred before the production of aflatoxins. Aflatoxin production (AFB1 and AFG1) was detected on day 3 on both strains. From day 4 onwards, all four toxins were detected and their production kept at similar proportions (AFG1>AFB1>AFG2>AFB2). Thus, the anticipated information obtained through early expression profile results of aflC, aflQ, aflU, and aflX gene cluster in A. nomiae may foresee aflatoxin production before its detection in Brazil nuts.

Blood and body fluids exposure of healthcare workers in a university hospital of Palermo, Italy: a fourteen years long surveillance.

BACKGROUND: Healthcare workers are habitually in direct contact with patients, possible carriers of infectious diseases and with potentially infectious biological materials; therefore, the implementation of standard precautions and good working practices represent an intervention strongly recommended by the Centers for Disease Control and Prevention, and required by Italian law, for the prevention of professional cut wounds. The study focused on assessing the exposure frequency and factors related to biological injuries among healthcare workers in a teaching hospital in Palermo, Italy. METHODS: We performed a 14-years retrospective descriptive analysis on blood and body fluids exposures in healthcare workers, documented by questionnaires administered at the time of injury and by data collected during the follow-up period. The questionnaire included questions concerning personal data (age, sex), job position (role, employment contract, ward), biological exposure (type of exposure, devices used and circumstance of blood and body fluids exposure), precautions adopted (personal protecting equipment, safety devices) and vaccination status. RESULT: A total amount of 899 healthcare workers was investigated. The incidence rate per 100 beds was 10.7. Frequency of exposure to blood and body fluids among healthcare workers was 35.3% in nurses, 31.7% in physicians, 17.6% in students. The mean age of injured healthcare workers was 36 years. The most common blood and body fluids exposures were represented by needlestick injury (76.2%), splash and spill (15.0%) and sharp (8.3%). 585 out of 685 percutaneous exposures were caused by needles (syringe, peripheral venous catheter, butterfly needles, etc.) and occurred mainly to nurses (N=224, 38.3%), physicians (N=184, 31.4% of whom resident physicians=122, 20.1% and hospital doctors=62, 10.6%), students (N=96, 16.4%) and auxiliary personnel (N=77, 13.1%). No seroconversion among exposed healthcare workers was recorded in the whole survey period. Twenty-four healthcare workers (2.6%) received post-exposure prophylaxis against Hepatitis B Virus. CONCLUSION: To our knowledge, this is the first long-term survey on blood and body fluids exposure in Southern Italy. Nurses are the most commonly affected group by biological injuries. Resident physicians and students follow the nurses probably due to a lack of training and experience about biological risk. These last two groups, however, seem to have more awareness of blood and body fluids exposures to which they are susceptible during their training cycle; in fact, they mostly use personal protective equipment compared to other healthcare workers. The blood and body fluids exposures are a preventable and a major occupational hazard in healthcare. This focus highlights the need for interventions to enhance the occupational safety of workers and students.

Metal-Associated Biomarker Responses in Crabs from a Marine Protected Area in Southeastern Brazil.

The environmental quality of a Ramsar wetland site located at the Cananéia-Iguape-Peruíbe Protected Area (CIP-PA), in São Paulo, Brazil, was assessed by geochemical analyses and biomarker assessments (GPx, GST, GSH, GST, MT, LPO, DNA damage) performed in swimming crab Callinectes danae Smith, 1869 organs (posterior and anterior gills and hepatopancreas) to estimate sediment contaminant bioavailability. The results indicated that two sampling stations, PT and PM, exhibited the worst environmental conditions, as sediments collected at both points contained metal contamination, while crabs exhibited significant responses for GPx, GST, and LPO (mostly during winter). Sediment contamination tended to be associated to fine sediments (both seasons) and organic matter (winter). During the summer survey, Pb concentrations in sediments of station PT exceeded the Brazilian Sediment Quality Guidelines (SQGs) and the Canadian Interim Marine Sediment Quality Guidelines. Metal concentrations in sediments sampled in winter were higher compared with summer, with Co, Ni, and Pb exceeding SQGs levels at PT, whereas Co, Ni, Hg, Zn, and Pb exceeded SQGs at PM. Biomarker induction during summer appeared to be caused by natural variables (water salinity and temperature, and molting cycle), whereas oxidative stress and tissue damage during winter appeared to be more clearly linked to metal contamination. Anterior gills presented the clearest signs of seasonal variability, being more responsive to sediment contamination. The results suggest that metals originated from the upper Ribeira de Iguape River are transported toward the estuarine system, causing effects on C. danae individuals. Additionally, seasonality is a strong factor concerning CIP-PA toxicity, since the rainfall regime significantly modifies the freshwater flow and, consequently, estuarine water salinity, suspended particle and metal inputs, as well as the location of depositional areas. Thus, efforts to mitigate CIP-PA contamination should be based on the control of upstream pollution sources.

Biomarkers of exposure to metal contamination and lipid peroxidation in the benthic fish Cathorops spixii from two estuaries in South America, Brazil.

Biomarkers as lipid peroxidation, metallothionein and delta-aminolevulinic acid dehydratase were determined in Cathorops spixii to compare the biological responses of this fish from estuaries with distinct anthropogenic influence. Three areas were selected in two estuaries in accordance with the levels of contamination for the polluted (Santos/São Vicente) and with the hydrodynamic characteristics for the non-polluted (Cananéia) estuary. Water characteristics and mercury levels in C. spixii confirmed a high human influence in the polluted system. In general, the biomarkers showed differences between the estuaries, suggesting disturbances in the specific cell mechanisms due to the presence of multiple xenobiotics in the contaminated system. Therefore, these biomarkers are recommended to promote more accurate information about the exposure to pollutants. Additionally, the study of the effect of the multiple xenobiotics on resident species such as the benthic fish C. spixii can favor a better assessment of the environmental quality of these systems.

Use of Cathorops spixii as bioindicator of pollution of trace metals in the Santos Bay, Brazil.

In the present study Cathorops spixii, was evaluated as a bioindicator fish for trace metal pollution. Concentrations of cobalt (Co), iron (Fe), selenium (Se) and zinc (Zn) were determined by Instrumental Neutron Activation Analysis in liver. Mercury (Hg) and methyl-mercury (MeHg) were analyzed by Cold Vapor Atomic Absorption Spectrometry in muscles and livers. High concentrations of Co, Fe, Se and Zn were observed in C. spixii from Santos Bay in comparison to fish collected in a non-polluted site in the same Brazilian coast. These trace metal concentrations were out of the permissible levels for human consumption. Although, Hg and MeHg levels were low, the C. spixii could still be used as an effective bioindicator to observe trace metal behaviors in the environment in function of the bioaccumulation process observed mainly by other analyzed trace metals. Thus, the use of this species is strongly recommended to monitor the effects and behavior of trace metal pollution in aquatic ecosystems in Brazil due to its bioaccumulation function.

Radiological characterisation of disposed phosphogypsum in Brazil: evaluation of the occupational exposure and environmental impact.

Phosphogypsum, a waste by-product derived from the wet process production of phosphoric acid, represents a serious problem facing the phosphate industry in Brazil. This by-product (mainly calcium sulphate dihydrate) precipitates during the reaction of sulphuric acid with phosphate rock and is stored at a rate of about 4x10(6) kg per day on several piles in Cubatão, Brazil. Contents of natural radionuclides from thorium and uranium series were measured in Brazilian phosphogypsum samples from disposal piles, using high-resolution gamma ray spectrometry and instrumental neutron activation analysis (NAA). These phosphogypsum piles present a potential threat to the surrounding environment and to the individual occupationally exposed. The results obtained in this study show that radionuclides, although present in relatively high concentrations in phosphogypsum, do not imply in significant doses for individuals occupationally exposed. The results obtained for the water activity in the monitor wells showed that the run-off of the piles is influenced by the activity present in the piles, giving indication of a possible groundwater contamination. Sediments from rivers in the area of influence of the pile presented higher concentrations of 238U and 232Th when compared with reference values.

Partitioning of radionuclides and trace elements in phosphogypsum and its source materials based on sequential extraction methods.

Phosphogypsum is a waste produced by the phosphate fertilizer industry. Although phosphogypsum is mainly calcium sulphate dihydrate, it contains elevated levels of impurities, which originate from the source phosphate rock used in the phosphoric acid production. Among these impurities, radionuclides from 238U and 232Th decay series are of most concern due to their radiotoxicity. Other elements, such as rare earth elements (REE) and Ba are also enriched in the phosphogypsum. The bioavailability of radionuclides (226Ra, 210Pb and 232Th), rare earth elements and Ba to the surrounding aquatic system was evaluated by the application of sequential leaching of the phosphogypsum samples from the Brazilian phosphoric acid producers. The sequential extraction results show that most of the radium and lead are located in the "iron oxide" (non-CaSO4) fraction, and that only 13-18% of these radionuclides are distributed in the most labile fraction. Th, REE and Ba were found predominantly in the residual phase, which corresponds to a small fraction of the phosphate rock or monazite that did not react and to insoluble compounds such as sulphates, phosphates and silicates. It can be concluded that although all these elements are enriched in the phosphogypsum samples they are not associated with CaSO4 itself and therefore do not represent a threat to the surrounding aquatic environment.

Emergency maxillofacial radiology. Foreign body localization: report of cases.

Successful emergency removal of foreign bodies of the maxillofacial complex depends on the surgeon's ability to perform exact location procedures. In hospitals and/or offices where advanced imaging technologies are not available, the attendant clinician has to use and rely on variations of conventional radiographic techniques in order to complement the diagnosis and locate the foreign body. Three cases are presented where unconventional radiographic techniques were used, leading to successful surgical removal of foreign bodies.

Expression of heat shock protein 72 in renal cell carcinoma: possible role and prognostic implications in cancer patients.

Renal cancer cells from 43 patients and normal renal cells from 10 of them were studied for the expression of highly stress-inducible heat shock protein 72 (HSP72) by means of immunoperoxidase analysis. It was found that HSP72 was expressed in a significantly higher percentage of renal cancer cells than normal renal cells (P = 0.0001), the mean percentage of positive cells being 33.1 +/- 18% and 8 +/- 5%, respectively. Moreover, a percentage of HSP72-positive cells that was less than the cut-off point (18%, mean value of normal cells +2 S.D.) significantly correlated with shorter disease-free survival (P = 0.002). The renal cancer cell populations taken from the 21 patients who relapsed after a median time of 13 months (range 3-73 months) had a significantly lower percentage of HSP72-positive cells (mean value 25.1 +/- 17%) than the cells taken from the patients who remained tumour-free (mean value 40.8 +/- 15%) after a median period of 72 months (range 19-96 months, P = 0.003). It was also demonstrated that HSP72 expression can be significantly increased by 48-h in vitro incubation with rIFN-gamma (P = 0.007). These data suggest that HSP72 may represent a favourable prognostic factor regardless of stage and histological grade and its expression may be increased by treatment with rIFN-gamma. Further studies are needed in order to investigate the relationship between HSP72 and the immunoeffector cells.

Increasing 4'-epidoxorubicin and fixed ifosfamide doses plus granulocyte-macrophage colony-stimulating factor in advanced soft tissue sarcomas: a pilot study.

PURPOSE: To determine the maximum-tolerated dose (MTD) of 4'-epidoxorubicin (EPI) in combination with full dose of ifosfamide (IFO) when granulocyte-macrophage colony-stimulating factor (GM-CSF) was used, to estimate its clinical efficacy, and to evaluate the mobilization of hematopoietic progenitors. PATIENTS AND METHODS: Previously untreated advanced patients were treated with fixed doses of IFO at 1.8 g/m2/d for 5 days and escalating doses of EPI. The starting dose level of EPI was 50 mg/m2 bolus on days 1 and 2; subsequent levels were 60 mg/m2 and 70 mg/ m2 given on days 1 and 2. GM-CSF (5 micrograms/kg/d) was administered from days +6 to +19. Clinical evaluation of response was performed after three consecutive cycles. Mobilization of hematopoietic progenitors was evaluated as day 14 CFU-GM after the first cycle only. RESULTS: Overall, six, 18, and 13 assessable patients were entered onto each EPI dose level, respectively. The first and the second EPI level were considered feasible. Conversely, at the third level, only six of 13 patients [46%] tolerated full EPI doses at the scheduled time. Therefore, the dose-intensity of the three levels was 100%, 99.7%, and 86.1%, respectively. Overall, 20 of 37 patients (54%) obtained an objective response. The response rates for the three EPI dose levels were significantly different [17%, 33%, and 100%, respectively; test for trend, P < .001]. Considering only lung metastases, the overall response rate was 72% (20%, 66%, and 100% for the three EPI levels, respectively). The most relevant mobilization effect was obtained at the third EPI level, when both GM-CSF and IL-3 were used as in vitro-stimulating factors. CONCLUSION: The third EPI level (70 mg/m2 on days 1 and 2) is the MTD of this program, since it was administered, without dose reduction or treatment delay, for three consecutive cycles in less than half of the patients. Nevertheless, this level proved to be interesting with regard to response rate (13 of 13 objective responses) and in mobilization of the hematopoietic progenitors.

Paclitaxel as a radiosensitiser: a proposed schedule of administration based on in vitro data and pharmacokinetic calculations.

Paclitaxel is efficacious against many human cancers. Because it blocks cells at the radiosensitive G2-M interface, paclitaxel has been investigated as a radiosensitiser. The results have been equivocal and somewhat contradictory. It is impossible to obtain proper pharmacokinetic calculations, aimed at obtaining maximum cytotoxicity and/or radiosensitisation, without knowing (i) how long the drug must be in contact with the cells, (ii) how long the effect lasts after the drug is removed from the cellular environment, (iii) whether the drug acts as a radiosensitiser even when, like cis-platinum, it is added after the radiation and (iv) what the minimum quantity of drug in the cellular environment is required for both chemotoxicity and radiosensitisation. The present work addresses the above questions. Two radioresistant cell lines of human origin were used, A375 melanoma and S549 lung carcinoma, in a clonogenic assay where only colonies with 50 or more cells were counted. For the irradiation, 6 MV X-rays were used. Any G2-M block was quantified by cell cycle kinetics analysis. From the results, a simulation of pharmacokinetics was conducted to calculate the schedule of administration of paclitaxel most likely to achieve and maintain significant chemotoxocity and radiosensitisation. The minimum concentration of paclitaxel for measurable cytotoxicity was 3 nM for both cell lines, but the drug was more toxic to the A549 cells. The minimum concentration for measurable radiosensitisation was 3 nM for A375 and approximately 0.1 nM for A549, but whereas above 3 nM the radiosensitivity increased in A375, it decreased above 1 nM for A549. A minimum of 18 h incubation with the drug was necessary for measurable effects and the radiosensitising effects were lost soon after its removal. There was no radiosensitisation if paclitaxel was added after the radiation, and, at the minimum effective concentrations, it caused only a minor and transient G2-M block. The pharmacokinetic calculations predict that 15 mg/m2 paclitaxel given as a 1 h infusion 5 days/week for 3 weeks during the radiotherapy should achieve both cytotoxicity and radiosensitisation. The mechanism of radiosensitisation by paclitaxel at the concentrations suggested by our results does not appear to be via a G2-M block and is probably concentration dependent. The results imply that low-dose, daily infusions of paclitaxel for as long as possible during a course of radiotherapy are more likely to result in radiosensitisation and prolonged cytotoxicity than high-dose infusions given once a week.

Interleukin-6 and soluble intercellular adhesion molecule-1 in renal cancer patients and cultured renal cancer cells.

Serum interleukin-6 (IL-6) and soluble intercellular adhesion molecule-1 (sICAM-1) levels were measured by enzyme-linked immunosorbent assay in 62 renal cancer patients: 30 were tumor-free after radical nephrectomy and 32 presented with metastatic disease. Serum IL-6 was undetectable in all but one of the tumor-free patients, whereas 41% (13 of 32) of the metastatic patients presented serum IL-6 levels. Furthermore, there was a significant correlation between serum IL-6 levels and a shorter overall survival (p = 0.009). Moreover, serum sICAM-1 levels were significantly higher (p = 0.05) in the metastatic patients with detectable serum IL-6 than in those without IL-6, suggesting a possible link between IL-6 and sICAM-1. The probability of a shorter overall survival was greater in the metastatic patients with both serum IL-6 and elevated sICAM-1 levels (>635 ng/ml), than in those with elevated sICAM-1 but without IL-6 (p = 0.01). The production of IL-6 by 16 freshly dissociated renal cancer cells cultured in vitro was also observed. It appeared that IL-6 levels did not correlate with the expression and release of ICAM-1 by cultured cells, although the highest values of ICAM-1 release were found in cultures synthesizing the highest values of IL-6. In conclusion, in vivo presence of serum IL-6 and elevated sICAM-1 levels is related to an unfavorable prognosis; it can be speculated that the cells capable of releasing high levels of sICAM-1 and IL-6 may negatively influence the antitumor response.

Determination of various nutrients and toxic elements in different Brazilian regional diets by neutron activation analysis.

The concentration of 19 elements (As, Br, Ca, Cd, Cl, Co, Cr, Cu, Fe, Hg, K, Mg, Mn, Na, Rb, Sb, Sc, Se and Zn) was evaluated in some diets taken from different regions of Brazil by Instrumental and Radiochemical Neutron Activation Analysis. Several populations with different socio-economic living conditions and inhabiting in different regions of Brazil were studied in order to estimate and to detect the variability of the mineral and toxic element content among Brazilian populational groups. The data obtained showed a significant difference between the contents of these elements in the diets from the regions studied. The general conclusions from the data obtained in this study were: 1) regarding the daily amounts of essential elements (Ca, Cl, Co, Fe, Mn, Na, K, Fe, Se and Zn), the Santa Catarina 2 diet showed the closest values when compared to the recommended values of RDA (Recommended Dietary Allowance) and/or WHO (World Health Organization). The Santa Catarina 1(low income groups) showed the lowest when compared to the same values. 2) The intake of toxic elements (As, Br, Cd, Hg, Sb) among the diets does not seem to be a major problem when compared to PTWI (Provisional Tolerable Weekly Intake, WHO), except for Hg intakes in regions near gold mining activities, like Manaus and Mato Grosso, where the values found were near the upper limit set by WHO.

Adjuvant immunotherapy treatment of renal carcinoma patients with autologous tumor cells and bacillus Calmette-Guèrin: five-year results of a prospective randomized study.

BACKGROUND: Active specific immunotherapy (ASI) is a strategy that attempts to boost the host's immune response specifically against its own tumor. The purpose of this study was to investigate the effect of adjuvant ASI in patients with renal carcinoma. METHODS: Of 120 consecutive patients, 60 were randomized to a control group and 60 to receive ASI comprised of 3 intradermal injections of 10(7) autologous irradiated tumor cells mixed with 10(7) Bacillus Calmette-Guèrin (in the first 2 vaccinations) or alone. At randomization and 1, 6, and 12 months after completing immunotherapy, the treated patients were evaluated for the development of delayed type cutaneous hypersensitivity (DTCH) response to autologous tumor and autologous normal renal cells. RESULTS: The baseline DTCH responses were negative in all patients. One month after completing ASI, 38 of 54 immunized patients showed a significant (P<0.01) DTCH response to autologous tumor but not to autologous normal renal cells. The response was persistent at 6 months in 25 of 44 patients and at 12 months in 16 of 28 patients. DTCH response remained negative in the nonimmunized control patients. There was no systemic toxicity but local ulcerations that healed in 2 months were observed. After a median follow-up of 61 months, the probability of 5-year disease free survival (DFS) was 63% for treated patients and 72% for control patients. The corresponding probability of 5-year overall survival (OS) was 69% and 78%, respectively. These differences were not statistically significant. CONCLUSIONS: This is the first prospective randomized study of ASI in a large population of patients with renal carcinoma after radical nephrectomy. Our data clearly indicate that ASI can increase the reactivity to autologous tumor, as measured by the DTCH test, but it appears unable to affect DFS and OS of patients.

Expression and release of intercellular adhesion molecule-1 in renal-cancer patients.

We examined ICAM-1 expression in 37 freshly dissociated renal-cancer cell populations. Immunoperoxidase analysis revealed that 31 of the 37 renal tumors expressed ICAM-1 to various degrees; ICAM-1 expression was significantly lower in tumor cells obtained from patients remaining tumor-free after a median follow-up of 60 months (mean value 24.4% +/- 21) than in tumor cells obtained from the relapsed patients (mean value 40.8% +/- 22), and the low expression of this molecule on the cell surface seemed to correlate with favorable clinical behavior. In 41 patients, the mean level of sICAM-I was 551 +/- 260 ng/ml, significantly higher than normal. However, sICAM-1 levels were significantly lower in the 20 tumor-free (mean 467 +/- 158 ng/ml) than in the 21 metastatic patients (mean 631 +/- 318 ng/ml). Eleven renal-cancer cell populations were cultured in order to examine the expression and release of ICAM-1. All of these cells were positive for ICAM-1 expression, which was elevated in 6 cases (> 50%) and low in the remaining 5 cases (18-35%). However, only the 5 cell populations expressing low levels of ICAM-1 released this molecule, showing an inverse correlation with cellular expression. Five of the cell populations were treated for 48 hr with rIFN-gamma, in these cells, both ICAM-1 expression and sICAM-1 levels increased, although sICAM-1 levels in the supernatants of the cell populations with constitutive high ICAM-1 expression remained very low.

Soluble intercellular adhesion molecule-1 and serum cytokines in melanoma patients treated with liposomes containing muramyl tripeptide.

AIMS AND BACKGROUND: A soluble form of intercellular adhesion molecule-1 (sICAM-1) has been recently identified in patients with malignant melanoma. It has been demonstrated that inflammatory cytokines can modulate the cellular expression of ICAM-1 and the shedding of this molecule by cells. To our knowledge, few data exist on serum sICAM-1 levels in cancer patients treated with immunomodulators. Liposomes containing muramyl tripeptide (MLV MTP-PE) can activate monocytes from cancer patients in vitro and in vivo, making them cytotoxic such as tumor necrosis factor-alpha (TNF-alpha) and Interleukin-6 (IL-6). The purpose of the present study was to evaluate the levels of sICAM-1 and their possible correlation with serum inflammatory cytokine levels in melanoma patients treated with MLV MTP-PE. METHODS: The sera from 9 patients with metastatic melanoma treated with MLV MTP-PE, 4 mg i.v. twice a week for 12 weeks, were tested in ELISA system to detect sICAM-1, TNF-alpha, IL-6, Interleukin-1 beta (IL-1 beta) and Interferon-gamma (IFN-gamma) before, and 2 and 24 h after the 1st, 12th and 24th infusion of MLV MTP-PE. RESULTS: Baseline levels of sICAM-1 were elevated in all patients (median 540 ng/ml: range 400-1030 ng/ml). Twenty-four h after the 1st infusion of MLV MTP-PE, we observed 6 increases in sICAM-1 levels, 1 decrease and 2 stable values (median 720 ng/ml: range 410-1820; P = 0.060). Twenty-four h after the 12th infusion, sICAM-1 increased in 3 patients and did not change in 4 (median 790 ng/ml: range 495-1650 ng/ml; P = 0.069). At the 24th infusion, sICAM-1 increased in 4 of 6 evaluable patients and remained stable in 2 (median 802 ng/ml: range 510-1450 ng/ml; P = 0.045). To better analyze the variations in sICAM-1, the patients were arbitrarily divided into two groups according to their clinical behavior: 4 presented stabilization (all lesions, n = 2; some lesions, n = 2) (Group A); 5 presented progressive disease (Group B). In Group A, sICAM-1 levels remained stable or showed a modest increase during treatment (except in 1 patient, who exhibited a substantial variation after the 12th infusion). In contrast, in Group B very high levels of sICAM-1 were observed at the beginning of the study therapy in 1 patient and after the 1st infusion in 3 patients; these values remained high until the 24th infusion. In most of the patients, TNF-alpha and IL-6 increased after the 1st infusion, but not thereafter. IFN-gamma was never detected; IL-1 beta was detectable in a few cases, but only before the infusions. CONCLUSIONS: baseline levels of sICAM-1 were elevated in all patients and further increased during treatment only in patients with more aggressive disease. No correlation was found between sICAM-1 and inflammatory cytokines. It would therefore seem that in patients with advanced disease, higher levels and a progressive increase in sICAM-1 may be unfavorable prognostic factors.

Induction and maintenance of monocyte cytotoxicity during treatment with liposomes containing muramyl tripeptide despite tachyphylaxis to the cytokine response.

Monocyte-mediated cytotoxicity (determined in a 72-h111In release assay) and the circulating levels of tumor necrosis factor alpha (TNF-alpha), interleukin (IL) 1beta, IL-6, IFN-gamma, C-reactive protein, and beta2-microglobulin were determined in 14 melanoma patients treated with multilamellar vesicle liposomes containing muramyl tripeptide phosphatidylethanolamine, 4 mg twice a week for 12 weeks. Monocyte-mediated cytotoxicity increased 24 h after the first infusion in 9 of 14 patients and had reached maximum levels (mean, 44% +/- 8) in all patients by the sixth week; similar values were observed at the 12th week. Once increased in vivo, peripheral blood monocyte cytotoxicity was not susceptible to any further increase after a subsequent in vitro incubation of the monocytes with liposomes. However, the peripheral blood monocytes which were not cytotoxic in vivo were activated by in vitro incubation with liposomes and not by medium. TNF-alpha and IL-6 peaked 2 h after the first infusion and returned to baseline values at 24 h; they were not significantly increased by subsequent treatments. The induction of fever in patients, observed 2 h after the first infusion, correlated with TNF-alpha and IL-6 levels. Similarly, C-reactive protein levels also increased at 24 h, but only after the first dose. No increase in beta2-microglobulin and IL-1beta levels was observed, and IFN-gamma was never detected in serum. Two patients experienced stable disease lasting 7 and 12 months, and 12 patients progressed. These results show that multilamellar vesicle muramyl tripeptide phosphatidylethanolamine administration activates monocyte cytotoxicity and cytokine production (TNF-alpha, IL-6). Chronic treatment with multilamellar vesicle muramyl tripeptide phosphatidylethanolamine results in tachyphylaxis in terms of cytokine secretion but not cytotoxicity. There was no difference between the maximum cytotoxicity levels obtained in vivo and those obtained in vitro using the same agent. A better understanding of immunoregulation is required for a rational application of this and related immunotherapies.

In vitro synergic effect of interferon gamma combined with liposomes containing muramyl tripeptide on human monocyte cytotoxicity against fresh allogeneic and autologous tumor cells.

AIMS: The purpose of the present study was to investigate whether human recombinant interferon-gamma (hrIFN-gamma) can act synergically with various activators in increasing the cytotoxicity of cancer patient monocytes against fresh autologous and allogeneic tumor cells. METHODS: Fresh target cells were obtained by means on the mechanical and enzymatic dissociation of human renal carcinomas. A 375 and SW 626 cell lines were used as positive controls. Monocytes from renal cancer patients and normal volunteers were activated in vitro with lipopolysaccharide, muramyl tripeptide (MTP-PE) or liposomes containing MTP-PE (MTP-PE liposomes), with or without a pre-incubation with hrIFN-gamma and were tested for cytotoxicity by means of a 72-hr 111indium-release assay. All of the patients were tumor free at the time of the study. RESULTS: Cancer patient peripheral blood monocytes were activated in vitro by different immunomodulators and became cytotoxic to freshly dissociated autologous or allogeneic tumor cells. A synergic effect producing maximal cytotoxicity was obtained with an appropriately scheduled combination of hrIFN-gamma (10 U/ml) and MTP-PE liposomes (50 nm/ml), free lipopolysaccharide (10 micrograms/ml) or MTP-PE (100 micrograms/ml). The synergic cytotoxicity was observed against fresh allogeneic and autologous tumor cells, as well as against cultured cells. CONCLUSIONS: All of these data support the possibility of a combined treatment using hrIFN-gamma and MTP-PE liposomes in human studies, particularly when it is borne in mind that liposomes can prevent the direct toxicity of many immunomodulators and that the low levels of hrIFN-gamma required for the synergic activation are not toxic in vivo.

Neutron activation analysis of biological samples at the radiochemistry division of IPEN-CNEN/SP.

Neutron activation analysis is a very useful method for determination of a great number of elements in biological samples. At the Radiochemistry Division of the IPEN-CNEN/SP, this method is being extensively applied to study several materials, such as extracts from medicinal plants, human hair, snake venoms, human lungs, food-stuffs, and corn samples. Both instrumental neutron activation analysis (INAA) and radiochemical neutron activation analysis (RNAA) are used to analyze real samples, as well as biological standard reference materials to evaluate the accuracy and precision of the results.