Acute life-threatening extrinsic allergic alveolitis in a paint controller.

BACKGROUND: Occupational diisocyanate-induced extrinsic allergic alveolitis (EAA) is a rare and probably underestimated diagnosis. Two acute occupational EAA cases have been described in this context, but neither of them concerned hexamethylene diisocyanate (HDI) exposure. AIMS: To investigate the cause of a life-threatening EAA arising at work in a healthy 30-year-old female paint quality controller. METHODS: Occupational medical assessment, workplace evaluation, airborne and biological monitoring and immunodermatological tests. RESULTS: Diagnosis of EAA relied on congruent clinical and radiological information, confirmed occupational HDI exposure and positive IgG antibodies and patch tests. The patient worked in a small laboratory for 7 years, only occasionally using HDI-containing hardeners. While working with HDI for 6 h, she developed breathlessness, rapidly progressing to severe respiratory failure. Workplace HDI airborne exposure values ranged from undetectable levels to 4.25 p.p.b. Biological monitoring of urinary hexamethylene diamine in co-workers ranged from <1.0 to 15.4 µg/g creatinine. Patch tests 8 months later showed delayed skin reaction to HDI at 48 h. Subsequent skin biopsy showed spongiotic dermatitis with infiltration of CD4(+) and CD8(+) T cells. CONCLUSIONS: We believe this is the first reported case of acute life-threatening EAA following exposure to HDI. Low concentrations of airborne HDI and relatively high urinary hexamethylene diamine suggest significant skin absorption of HDI could have significantly contributed to the development of this acute occupational EAA.

Distinctive role of integrin-mediated adhesion in TNF-induced PKB/Akt and NF-kappaB activation and endothelial cell survival.

Tumor necrosis factor (TNF) is a pro-inflammatory cytokine exerting pleiotropic effects on endothelial cells. Depending on the vascular context it can induce endothelial cell activation and survival or death. The microenvironmental cues determining whether endothelial cells will survive or die, however, have remained elusive. Here we report that integrin ligation acts permissive for TNF-induced protein kinase B (PKB/Akt) but not nuclear factor (NF)-kappaB activation. Concomitant activation of PKB/Akt and NF-kappaB is essential for the survival of endothelial cells exposed to TNF. Active PKB/Akt strengthens integrin-dependent endothelial cell adhesion, whereas disruption of actin stress fibers abolishes the protective effect of PKB/Akt. Integrin-mediated adhesion also represses TNF-induced JNK activation, but JNK activity is not required for cell death. The alphaVbeta3/alphaVbeta5 integrin inhibitor EMD121974 sensitizes endothelial cells to TNF-dependent cytotoxicity and active PKB/Akt attenuates this effect. Interferon gamma synergistically enhanced TNF-induced endothelial cell death in all conditions tested. Taken together, these observations reveal a novel permissive role for integrins in TNF-induced PKB/Akt activation and prevention of TNF-induced death distinct of NF-kappaB, and implicate the actin cytoskeleton in PKB/Akt-mediated cell survival. The sensitizing effect of EMD121974 on TNF cytotoxicity may open new perspectives to the therapeutic use of TNF as anticancer agent.

Zoledronate inhibits endothelial cell adhesion, migration and survival through the suppression of multiple, prenylation-dependent signaling pathways.

BACKGROUND: Recent evidence indicates that zoledronate, a nitrogen-containing bisphosphonate used to treat conditions of increased bone resorption, may have anti-angiogenic activity. The endothelial cells signaling events modulated by zoledronate remain largely elusive. OBJECTIVES: The aim of this work was to identify signaling events suppressed by zoledronate in endothelial cells and responsible for some of its biological effects. METHODS: Human umbilical vein endothelial cells (HUVEC) were exposed to zoledronate, isoprenoid analogs (i.e. farnesol and geranylgeraniol) and various inhibitors of signaling, and the effect on adhesion, survival, migration, actin cytoskeleton and signaling events characterized. RESULTS: Zoledronate reduced Ras prenylation, Ras and RhoA translocation to the membrane, and sustained ERK1/2 phosphorylation and tumor necrosis factor (TNF) induced JNK phosphorylation. Isoprenoid analogs attenuated zoledronate effects on HUVEC adhesion, actin stress fibers and focal adhesions, migration and survival. Isoprenoid analogs also restored Ras prenylation, RhoA translocation to the membrane, sustained FAK and ERK1/2 phosphorylation and prevented suppression of protein kinase B (PKB) and JNK phosphorylation in HUVEC exposed to TNF in the presence of zoledronate. Pharmacological inhibition of Rock, a RhoA target mediating actin fiber formation, phosphatidylinositol 3-kinase, an activator of PKB, MEK1/2, an activator of ERK1/2, and JNK, recapitulated individual zoledronate effects, consistent with the involvement of these molecules and pathways and their inhibition in the zoledronate effects. CONCLUSIONS: This work has demonstrated that zoledronate inhibits HUVEC adhesion, survival, migration and actin stress fiber formation by interfering with protein prenylation and has identified ERK1/2, JNK, Rock, FAK and PKB as kinases affected by zoledronate in a prenylation-dependent manner.

Urokinase-type plasminogen activator inhibits alpha 4 beta 1 integrin-mediated T lymphocyte adhesion to fibronectin independently of its catalytic activity.

The urokinase-type plasminogen activator (u-PA)/plasmin system plays an important role in promoting cell migration and invasion, an effect which is largely ascribed to the proteolytic activity of these enzymes. We investigated whether u-PA modulates integrin-dependent T lymphocyte migration and adhesion on fibronectin independently of its plasminogen activator function. Here we report that u-PA reduced the spontaneous and phorbol 12-myristate 13-acetate-induced migration of peripheral blood T lymphocytes on fibronectin by 20-50%, decreased the T lymphocyte and alpha4beta1(+)/alpha5beta1(+) K562 cell adhesion on fibronectin by 30-40%, and completely suppressed integrin alpha4beta1-dependent T lymphocyte and alpha4beta1(+)/alpha5beta1(+) K562 cell adhesion to the LDV-containing 40-kDa fibronectin fragment. The u-PA receptor was not essential for this effect. In contrast, adhesion of alpha4beta1(-)/alpha5beta1(+) K562 cells to an RGD-containing fibronectin fragment was unaffected. A recombinant protein comprising the N-terminal fragment of u-PA, but lacking its proteolytic domain, had the same inhibitory effect. Decreased adhesion was neither associated with a diminished cell surface expression of alpha4beta1 nor with a suppression of alpha4beta1 ligand-binding function. Our results demonstrate that u-PA inhibits alpha4beta1- but not alpha5beta1-mediated lymphocyte/leukocyte adhesion to fibronectin independently of its proteolytic activity. This finding provides additional evidence that matrix proteinases may participate in cell adhesion and migration control independently of their matrix-degrading activity.

Pulse treatment of human vascular endothelial cells with high doses of tumor necrosis factor and interferon-gamma results in simultaneous synergistic and reversible effects on proliferation and morphology.

Regional administration of high doses of tumor necrosis factor (TNF), interferon gamma (IFNgamma) and melphalan to patients with advanced cancers of the limbs, results in rapid and specific tumor necrosis, while the normal adjacent tissues remain unaffected. The tumor vasculature is selectively destroyed by this treatment, and neovascular endothelial cells appear to be an early and specific target of TNF and IFNgamma. To further understand some of the cellular events underlying these in vivo effects, we have investigated the response of human macro- and microvascular endothelial cells in vitro, after exposure to high doses of TNF and IFNgamma (up to 40 x 10(3) U/ml each). TNF and IFNgamma synergistically inhibited endothelial-cell proliferation by up to 80% after 72 hr of treatment. Achievement of synergy required the simultaneous presence of both cytokines. A cytokine pulse as short as 30 min was sufficient to induce maximal growth inhibition measured after 48 hr. Both cytokines also induced progressive and dose-dependent elongation of the endothelial-cell morphology. The effects on endothelial-cell proliferation and morphology were reversible upon removal of the cytokines. Moreover, replating of treated cells onto a fresh substrate immediately resulted in re-acquisition of their normal shape. In contrast to the effect on cell proliferation, there was little or no effect on the rate of endothelial-cell apoptosis. The presented data extend reports on the effects of TNF and IFNgamma on human endothelial cells in vitro, and suggest that the in vivo disruption of the tumor vasculature caused by high doses of TNF and IFNgamma is not due to a direct cytotoxic effect on endothelial cells but occurs through an indirect mechanism.

Evidence for the involvement of endothelial cell integrin alphaVbeta3 in the disruption of the tumor vasculature induced by TNF and IFN-gamma.

Administration of tumor necrosis factor (TNF) and gamma interferon (IFN-gamma) to melanoma patients causes selective disruption of the tumor vasculature but the mechanism of this disruption is unknown. Here we report that exposure of human endothelial cells to TNF and IFN-gamma results in a reduced activation of integrin alphaVbeta3, an adhesion receptor that plays a key role in tumor angiogenesis, leading to a decreased alphaVbeta3-dependent endothelial cell adhesion and survival. Detachment and apoptosis of angiogenic endothelial cells was demonstrated in vivo in melanoma metastases of patients treated with TNF and IFN-gamma. These results implicate integrin alphaVbeta3 in the anti-vascular activity of TNF and IFN-gamma and demonstrate a new mechanism by which cytokines control cell adhesion.

Analysis of prevention program effectiveness with clustered data using generalized estimating equations.

Experimental studies of prevention programs often randomize clusters of individuals rather than individuals to treatment conditions. When the correlation among individuals within clusters is not accounted for in statistical analysis, the standard errors are biased, potentially resulting in misleading conclusions about the significance of treatment effects. This study demonstrates the generalized estimating equations (GEE) method, focusing specifically on the GEE-independent method, to control for within-cluster correlation in regression models with either continuous or binary outcomes. The GEE-independent method yields consistent and robust variance estimates. Data from project DARE, a youth substance abuse prevention program, are used for illustration.

Cluster sampling techniques in quantal response teratology and developmental toxicity studies.

This paper presents a model-free approach for evaluating teratology and developmental toxicity data involving clustered binary responses. In teratology studies, a major statistical problem arises from the effect of intralitter correlation, or the potential for littermates to respond similarly. Some statistical methods impose strict distributional assumptions to account for extra-binomial variation, while others rely on nonparametric resampling and empirical variance estimation techniques. Quasi-likelihood methods and generalized estimating equations (GEE), which model the marginal mean/variance relationship, also avoid strict distributional assumptions. The proposed approach, often used to analyze complex sample survey data, is based on a first-order Taylor series approximation and a between-cluster variance estimation procedure, yielding consistent variance estimates for binomial-based proportions and regression coefficients from dose-response models. The cluster sample technique, presented here in the context of a logistic dose-response model, incorporates many of the advantages of quasi-likelihood methods, are valid for any underlying nested correlation structure, and are adaptable to a variety of analytical settings. The results of a simulation study show the cluster sample technique to be a viable competitor to GEE methods currently receiving attention.

Long-term evaluation of drug abuse resistance education.

Project DARE (Drug Abuse Resistance Education) is the most prevalent school-based drug-use prevention program in the United States, but there is little evidence of its effectiveness. Results from a longitudinal evaluation of the program in 36 schools in Illinois provide only limited support for DARE's impact on student's drug use immediately following the intervention, and no support for either continued or emerging impact on drug use 1 or 2 years after receiving DARE instruction. In addition, DARE had only limited positive effects on psychological variables (i.e., self-esteem) and no effect on social variables (e.g., peer resistance skills). Possible substantive and methodological explanations for the relative lack of DARE's effectiveness observed in this study are discussed.

Developmental neurotoxicity evaluation of orally administered isopropanol in rats.

Isopropanol was administered by gavage to timed-mated rats from Gestation Day (GD) 6 through Postnatal Day (PND) 21. Doses administered were 0, 200, 700, or 1200 mg/kg/day in a volume of 5 ml/kg. The dams were allowed to deliver and body weights and food consumption were recorded during gestation and lactation. Pups were counted, examined, sexed, and weighed on PND 0, 4, 7, 13, 17, 21, 36, 49, and 68. Litters were culled to eight pups (4:4 or 5:3 sex ratio) on PND 4 and litters without acceptable numbers of male and female pups were eliminated from the study. Pups were weaned on PND 22, and two pups from each litter and their dams were killed. Six of these pups from each dose group were perfused in situ for histopathological examination of the central and peripheral nervous system. Brains of the remaining pups were divided into four regions and weighed. Maternal liver and kidney weights were recorded. Weaned pups were assessed for day of testes descent or vaginal opening and for motor activity on PNDs 13, 17, 21, 47, and 58; auditory startle on PNDs 22 and 60; and active avoidance on PNDs 60-64. These pups were euthanized and examined on PND 68. One high-dose dam died on PND 15, but there were no other clinical observations or effects on maternal weight, food consumption, or gestation length. Pup survival, weight, sex ratio, and sexual maturation were unaffected. There were no biologically significant findings in the behavioral tests, no changes in organ weights, and no pathological findings that could be attributed to isopropanol exposure. In conclusion, there was no evidence of developmental neurotoxicity associated with isopropanol exposure as high as 1200 mg/kg/day.

Ratio estimates, the delta method, and quantal response tests for increased carcinogenicity.

This paper demonstrates the use of the delta method for estimating the variance of ratio statistics derived from animal carcinogenicity experiments. The Cochran-Armitage test (Cochran, 1954, Biometrika 10, 417-451; and Armitage, 1955, Biometrics 11, 375-386) is routinely applied to carcinogenicity data as a test for linear trend in lifetime tumor incidence rates. The computing formula for this test derives from the assumption that the denominators of the quantal response rates are fixed. However, when time-at-risk weights are introduced to correct for treatment-related differences in survival, the denominators of the quantal response rates are subject to random variation. The delta method and weighted least squares techniques are applied here to approximate the variance of such ratio statistics and test for a linear dose-response relationship among treatments. This technique is compared to that of Bailer and Portier (1988, Biometrics 44, 417-431), who introduced a survival-adjusted quantal response test for trend in lifetime tumor incidence rates. Their test modifies the usual Cochran-Armitage computing formula by weighting the denominators of the response rates to reflect less-than-whole-animal contributions to risk. Within the framework of a weighted least squares linear regression model that underlies the Cochran-Armitage test, the time-at-risk weights of Bailer and Portier are incorporated using the delta method. Although the delta method approach is slightly more computationally intensive, small-sample simulations indicate that it has superior operating characteristics over the Poly-3 trend test of Bailer and Portier when background tumor incidence rates are low (under 3%) and survival patterns differ markedly across treatments.(ABSTRACT TRUNCATED AT 250 WORDS)

Significance of cytokine production and adhesion molecules in malarial immunopathology.

The pathological expression in malaria infection depends largely on immunopathologic responses induced by the parasite. In the past few years, we have attempted to analyze mechanisms by which inappropriate immune response to some malarial antigens can generate major complications of malaria and particularly neurovascular lesions. To this end, we have undertaken a study aimed at a more precise definition of immunopathological parameters of malaria infection, and more particularly those involved in cerebral malaria (CM). CM, the most severe complication of falciparum infection in man, represents a major problem of public health at the world level.