The Scientific Basis of Uncertainty Factors Used in Setting Occupational Exposure Limits.

The uncertainty factor concept is integrated into health risk assessments for all aspects of public health practice, including by most organizations that derive occupational exposure limits. The use of uncertainty factors is predicated on the assumption that a sufficient reduction in exposure from those at the boundary for the onset of adverse effects will yield a safe exposure level for at least the great majority of the exposed population, including vulnerable subgroups. There are differences in the application of the uncertainty factor approach among groups that conduct occupational assessments; however, there are common areas of uncertainty which are considered by all or nearly all occupational exposure limit-setting organizations. Five key uncertainties that are often examined include interspecies variability in response when extrapolating from animal studies to humans, response variability in humans, uncertainty in estimating a no-effect level from a dose where effects were observed, extrapolation from shorter duration studies to a full life-time exposure, and other insufficiencies in the overall health effects database indicating that the most sensitive adverse effect may not have been evaluated. In addition, a modifying factor is used by some organizations to account for other remaining uncertainties-typically related to exposure scenarios or accounting for the interplay among the five areas noted above. Consideration of uncertainties in occupational exposure limit derivation is a systematic process whereby the factors applied are not arbitrary, although they are mathematically imprecise. As the scientific basis for uncertainty factor application has improved, default uncertainty factors are now used only in the absence of chemical-specific data, and the trend is to replace them with chemical-specific adjustment factors whenever possible. The increased application of scientific data in the development of uncertainty factors for individual chemicals also has the benefit of increasing the transparency of occupational exposure limit derivation. Improved characterization of the scientific basis for uncertainty factors has led to increasing rigor and transparency in their application as part of the overall occupational exposure limit derivation process.

Overexpression of p101 activates PI3Kgamma signaling in T cells and contributes to cell survival.

p101, the regulatory subunit of phosphatidylinositol-3-kinase-gamma (PI3Kgamma), was recently reported as a common site of retroviral insertion in T-cell lymphomas induced in mice by MoFe2-MuLV, a unique recombinant gammaretrovirus. The common interruption of p101 by retroviral integration suggests that the locus encodes an oncogene whose altered expression is related to the induction of T-cell malignancy. To examine a possible role in the malignant process, p101 was overexpressed in human T-cell lines Molt-4 and Jurkat. Transient overexpression of p101 induced apoptosis in recipient cells; however, stable expression could be established in cells that expressed moderate levels of p101. Constitutive p101 overexpression in those cells conferred significant protection against ultraviolet-induced apoptosis. Protection against apoptotic induction was attributed to p101-mediated activation of the Akt pathway. Constitutive overexpression of p101 enhanced the activity of p110gamma and further sensitized it to activation upon stimulation of G protein-coupled receptor. These findings are the first to implicate altered expression of p101 in malignancy, specifically in T-cell lymphoma. The findings further provide insight into the regulation of p110gamma, indicating that the stoichiometry of p110gamma and p101 are important in regulating PI3Kgamma signaling.

Background levels of key biomarkers of chemical exposure within the UK general population--pilot study.

The use of biomarkers is now an accepted measure of chemical uptake (possibly exposure) in risk assessment. However, information on background exposures and biomarker concentrations of many environmental chemicals in the general UK population is limited. This study aims to determine reference ranges for eleven biomarkers of chemical exposure, measurable in urine, within the general adult UK population. The study will involve 400 volunteers throughout the UK and is currently underway. Described here is a pilot study, carried out during August and September 2005 to test the study methodology. The initial results of the postal survey and urinary concentrations for cadmium (UCd) and mercury (UHg) are reported. A total of 78 individuals were recruited by post from the UK Electoral Register, to take part in the pilot study. Participants were asked to complete a questionnaire and provide a urine sample. The overall response rate was 16%, of which 60.3% were female and 39.7% male. Those living in suburban areas accounted for 60% of respondents, current smokers 12.8% and vegetarians 1.3%. Levels of UCd were higher in females compared to males and smoking status influenced levels; smokers displayed higher levels of UCd than individuals who had previously smoked or who had never smoked. The mean, median and range of UHg was 1.12, 0.55 (

Investigation of the possible involvement of ketonic derivatives of polycyclic hydrocarbons in hydrocarbon carcinogenesis.

5,6-Dihydro-5-oxo-7,12-dimethylbenz(a)anthracene and 5,6-dihydro-6-oxo-7,12-dimethylbenz(a)anthracene are described. These compounds, which are isomeric with 7,12-dimethylbenz(a)anthracene-5,6-oxide (the K-region epoxide), were inactive in tests for tumor-initiating activity in mouse skin and tumor production in the s.c. tissue of the mouse.

Relevance of the rat lung tumor response to particle overload for human risk assessment-Update and interpretation of new data since ILSI 2000.

The relevance of particle-overload related lung tumors in rats for human risk assessment following chronic inhalation exposures to poorly soluble particulates (PSP) has been a controversial issue for more than three decades. In 1998, an ILSI (International Life Sciences) Working Group of health scientists was convened to address this issue of applicability of experimental study findings of lung neoplasms in rats for lifetime-exposed production workers to PSPs. A full consensus view was not reached by the Workshop participants, although it was generally acknowledged that the findings of lung tumors in rats following chronic inhalation, particle-overload PSP exposures occurred only in rats and no other tested species; and that there was an absence of lung cancers in PSP-exposed production workers. Since the publication of the ILSI Workshop report in 2000, there have been important new data published on the human relevance issue. A thorough and comprehensive review of the health effects literature on poorly soluble particles/lung overload was undertaken and published by an ECETOC (European Centre for Ecotoxicology and Toxicology of Chemicals) Task Force in 2013. One of the significant conclusions derived from that technical report was that the rat is unique amongst all species in developing lung tumors under chronic inhalation overload exposures to PSPs. Accordingly, the objective of this review is to provide important insights on the fundamental differences in pulmonary responses between experimentally-exposed rats, other experimental species and occupationally-exposed humans. Briefly, five central factors are described by the following issues. Focusing on these five interrelated/convergent factors clearly demonstrate an inappropriateness in concluding that the findings of lung tumors in rats exposed chronically to high concentrations of PSPs are accurate representations of the risks of lung cancer in PSP-exposed production workers. The most plausible conclusion that can be reached is that results from chronic particle-overload inhalation studies with PSPs in rats have no relevance for determining lung cancer risks in production workers exposed for a working lifetime to these poorly soluble particulate-types.

flvi-2, a target of retroviral insertional mutagenesis in feline thymic lymphosarcomas, encodes bmi-1.

LC-FeLV is a myc-containing strain of feline leukemia virus which induces thymic lymphosarcoma in the domestic cat with short latency. A locus in feline DNA, termed flvi-2, is commonly interrupted in naturally occurring and experimentally induced thymic lymphosarcomas containing LC-FeLV; thus, interruption of a gene encoded by flvi-2 may cooperate with the myc oncogene in the induction of T-cell tumors by LC-FeLV. Clones homologous to flvi-2 have been isolated from a normal human thymus cDNA library. Nucleotide sequence analysis of the cDNA clones demonstrates that flvi-2 encodes bmi-1, a gene previously identified as a target for MoMuLV integration and as a myc-collaborator in retrovirally-induced B-cell lymphomas in E mu-myc transgenic mice. In feline thymic lymphomas, retroviral integrations occur downstream of the gene, and result in enhanced expression of a bmi-1 transcript of normal size. These findings demonstrate the interruption of bmi-1 in natural as well as experimentally induced tumors, implicate the activation of bmi-1 in the induction of T-cell as well as B-cell lymphoma, and support the premise that bmi-1 functions as a myc collaborator.

The Bmi-1 oncoprotein interacts with dinG and MPh2: the role of RING finger domains.

Experimentally-induced mutations in the C3HC4 RING finger domain of the Bmi-1 oncoprotein block its ability to induce lymphomas in mice. In this report, the role of the Bmi-1 RING finger in mediating protein-protein interactions is examined using the yeast two-hybrid system. Bmi-1 interacts directly with the RING finger protein dinG/RING1B. Heterodimerization of the two proteins requires the intact RING finger structures of both Bmi-1 and dinG. Although the RING finger domains are necessary for dimerization, they are not sufficient for this process as residues outside the C3HC4 motif are also required. Thus, binding specificity may be partly conferred by residues outside the RING motif. Both Bmi-1 and dinG interact with the Polyhomeotic protein MPh2 through binding domains apart from the RING finger. The data suggest a model whereby Bmi-1, dinG, and MPh2 form a stable heterotrimeric complex in which each protein contributes to the binding of the others.

Evolutionary conservation and chromosomal localization of flvi-1.

A locus in feline DNA, termed flvi-1, has been identified as harboring retroviral integrations commonly found in natural feline lymphomas induced by infection with feline leukemia virus (FeLV). Southern blot analysis of human and murine DNA using restriction fragments representing flvi-1 demonstrates its phylogenetic conservation among mammals, flvi-1 is localized to murine chromosome 2, proximal portion of band E, by in situ hybridization to metaphase chromosomes. This position is adjacent to that of another putative proto-oncogene, sfpi-1, although probes representing flvi-1 and sfpi-1 do not cross-hybridize. The repeated implication of flvi-1 in natural feline leukemogenesis, its evolutionary conservation and its chromosomal position support the hypothesis that flvi-1 may represent a previously unidentified protooncogene.

Mechanisms that contribute to the development of lymphoid malignancies: roles for genetic alterations and cytokine production.

Recent studies have defined genetic alterations commonly associated with transformed lymphocytes. This review suggests roles for these alterations in the development of lymphoid neoplasms. Damage to the genes encoding proteins that function in intracellular signaling, transcription, or regulation of the cell cycle has been identified and linked at varying degrees to the progression of certain lymphoid malignancies. An understanding of the mechanistic consequences following such genetic alterations is essential to an understanding of the development of these lymphoid neoplasms. In contrast, it is also becoming clear that the dysregulated expression of proteins that are not genetically altered can also contribute to the progression of lymphoid malignancies. One such example is the excessive expression of "normal" lymphokines of cytokines which accompanies many lymphoproliferative diseases. The dysregulated expression of cytokines during malignancy can result in the augmentation of growth of transformed lymphocytes, as well as an alteration of the anti-tumor immune response. The latter mechanism is especially important because evasion of the impending immune response is a prerequisite for the progression of lymphoproliferative diseases. Taken together, this review supports the notion that the development of lymphoid malignancies is multifactorial, involving genetic alterations as well as dysregulated cytokine expression.

Cooperating events in lymphomagenesis mediated by feline leukemia virus.

Feline leukemia virus (FeLV)-mediated lymphomagenesis in the domestic cat has been examined as a model of lymphoid malignancy in a naturally outbreeding population. The pathogenesis of two distinct, naturally occurring types of FeLV-induced tumors has been investigated: (1) a thymic lymphoma of T-cell origin, typical of FeLV-induced lymphoma, and (2) an extrathymic, extranodal lymphoma of non-B non-T-cell origin. The genetic features of these tumors are clearly distinguishable, and include determinants encoded both by the virus and the host. Virally encoded determinants of pathogenesis include the long terminal repeat (LTR) and the envelope SU protein. Cellular determinants include the involvement of a set of proto-oncogenes, and other factors characteristic of the specific cell type of origin of the tumor. Functional studies are aimed at evaluating the action and interaction of these genetic determinants in the pathogenesis of lymphoma in an animal model system.

Identification and analysis of a third mouse Polycomb gene, MPc3.

A new mouse Polycomb (Pc) gene, MPc3, has been identified. The MPc3 protein contains the highly conserved chromodomain and carboxy-terminal COOH box of other known Pc proteins from diverse species. Like other Pc proteins, MPc3 physically interacts with the RING finger proteins RING1A and dinG/RING1B. MPc3 maps to the distal arm of mouse chromosome 11 (11E2), a region that contains other known Pc genes in addition to several disease loci that may be linked to abnormal Pc gene function.

Complexity and content of the DNA and RNA in Trypanosoma cruzi.

The content and sequence complexity of the nuclear DNA and messenger RNA for epimastigotes of Trypanosoma cruzi were determined. From analysis of nuclear DNA reassociation studies and microspectrofluorometric measurements of laser induced fluorescence of cellular DNA, T. cruzi is found to be a diploid organism with a nuclear DNA content of 2.5 x 10(8) nucleotide pairs (2.8 x 10(-13) g) and a kinetoplast DNA content of 4.9 x 10(7) nucleotide pairs (5.4 x 10(-14) g). Reassociation kinetics of nuclear DNA of average length 0.4 kb reveals three kinetic components: a moderately repetitive component with a reiteration frequency of 5.1 x 10(3) present in 9% of the fragments, a lowly repetitive component with a reiteration frequency of 32 present in 51% of the fragments, and a single-copy component present in 23% of the fragments. By saturation hybridization of total polysomal RNA to 3H-labeled single-copy DNA, it was determined that 68% of the single-copy DNA was represented in the epimastigote polysomal RNA. This corresponds to ca. 12 000 different mRNA species. Of these, ca. 9000 are present as poly(A)+-RNA, while the remaining 3000 appear not to be polyadenylated. Kinetic analysis of the poly(A)+-RNA population indicates it is composed of at least three classes of RNA's of different abundancy levels: two sequences which occur ca. 3000 per cell, ca. 750 sequences which occur about 20 times per cell, and ca. 15 500 sequences which occur 1-2 times per cell.

Simian AIDS-associated lymphoma in rhesus and cynomolgus monkeys recapitulates the primary pathobiological features of AIDS-associated non-Hodgkin's lymphoma.

Non-Hodgkin's lymphomas occur with increased frequency (3-6%) in HIV-infected individuals. These AIDS-associated lymphomas (AALs) exhibit characteristics that distinguish them from lymphomas in the general population. A proposed model for the pathogenesis of AAL includes the following: (1) Tumorigenesis is multistep; (2) tumors occur in long-term survivors; (3) tumors are of clonal B cell origin; (4) HIV acts early and is an indirect effector; (5) tumor cells are infected with EBV; and (6) specific genetic lesions occur in tumor cells. Many aspects of this process remain to be tested in an animal model system. Since 1984, necropsy examinations have been performed on more than 1000 SIV-infected rhesus and cynomolgus monkeys at the Tulane Regional Primate Research Center. Lymphoid malignancies were detected in a proportion of SIV-infected animals. These SAIDS-associated lymphomas (SALs) have been studied to determine the extent to which their pathological features recapitulate a working model for the pathogenesis of AAL. The results show that lymphomas occur in SIV-infected rhesus macaques at 4% incidence, similar to that of AAL, and that the incidence of SAL in cynomolgus macaques is eightfold higher. Analysis of SAL from both species of macaques demonstrated significant similarity to the hallmark pathobiological features of AAL. These findings indicate that the HIV-infected human and the SIV-infected macaque share a common pathobiology and mechanism of lymphomagenesis.

Rhesus lymphocryptovirus infection during the progression of SAIDS and SAIDS-associated lymphoma in the rhesus macaque.

SAIDS-associated lymphoma (SAL) represents a monoclonal expansion of B-cell origin in which simian immunodeficiency virus (SIV) infection is not detected. However, tumor cells are frequently infected with rhesus lymphocryptovirus (RhLCV), a rhesus homologue of Epstein-Barr virus (EBV). In previous studies, the incidence of RhLCV infection in SAL was determined to be 89% as measured by polymerase chain reaction (PCR) and/or in situ hybridization. The main objective of the present study was to ascertain whether the level of RhLCV infection in the SIV-infected macaque is influenced as a function of SAIDS progression, and/or whether increased levels of RhLCV infection may correlate with the development of SAL. To this end, RhLCV infection was evaluated in three independent groups: (1) in lymphomas from SIV-infected rhesus macaques, (2) in peripheral blood mononuclear cells (PBMC) from a cohort of 69 randomly selected healthy animals, and (3) in PBMC collected from 22 SIV-infected animals at various times during progression to SAIDS or SAL. The relative levels of RhLCV infection were evaluated by PCR/Southern blot analysis, visual comparison to a standard dilution series, and assignment of relative signal intensity to a uniform classification scheme. The data show that SIV-infected monkeys have a generally higher RhLCV load in PBMC than do healthy animals, but that the virus load varies widely among animals during disease progression. Increased RhLCV load does not occur uniformly during the progression of SAIDS, although evidence indicates an increased RhLCV viral load in the development of SAL.

Variation in simian immunodeficiency virus env V1 region in simian AIDS-associated lymphoma.

Genetic variation of SIV env during the course of infection provides a large population pool that is continually shaped by selective forces in vivo and may influence the development of clinical disease. SAIDS-associated lymphoma (SAL) in the SIV-infected macaque is typically a clonal or oligoclonal mass of B cell origin, extranodal in anatomic distribution, in which SIV is restricted largely to infiltrating macrophages. To explore the degree of genetic variation in SIV env represented in SAL, a 480-bp DNA fragment containing the V1 region was PCR amplified from seven cases of SAL and from a nonneoplastic lymph node of an SIV-infected macaque. The nucleotide sequence of the V1 region was determined from at least 10 clones from multiple independent amplification reactions of each tissue. Overall, the degree of V1 variability within lymphomas was found not to be restricted but to resemble the heterogeneity reported in SIV-infected lymphoid and other tissues. V1 variation in the nonneoplastic lymph node was unexpectedly limited, perhaps related to the unusual disease condition associated with SAIDS in that animal. Unlike observations from SIV-infected tissues of animals without neoplastic disease, no increase was detected in the number of O- or N-linked glycosylation sites in the V1 regions isolated from lymphomas as compared with the original inoculum. These findings suggest that, within the microenvironment of the lymphoma, the immune evasion conferred by increased glycosylation may offer little selective advantage.

Comparative hazards of chrysotile asbestos and its substitutes: A European perspective.

Although the use of amphibole asbestos (crocidolite and amosite) has been banned in most European countries because of its known effects on the lung and pleura, chrysotile asbestos remains in use in a number of widely used products, notably asbestos cement and friction linings in vehicle brakes and clutches. A ban on chrysotile throughout the European Union for these remaining applications is currently under consideration, but this requires confidence in the safety of substitute materials. The main substitutes for the residual uses of chrysotile are p-aramid, polyvinyl alcohol (PVA), and cellulose fibers, and it is these materials that are evaluated here. Because it critically affects both exposure concentrations and deposition in the lung, diameter is a key determinant of the intrinsic hazard of a fiber; the propensity of a material to release fibers into the air is also important. It is generally accepted that to be pathogenic to the lung or pleura, fibers must be long, thin, and durable; fiber chemistry may also be significant. These basic principles are used in a pragmatic way to form a judgement on the relative safety of the substitute materials, taking into account what is known about their hazardous properties and also the potential for uncontrolled exposures during a lifetime of use (including disposal). We conclude that chrysotile asbestos is intrinsically more hazardous than p-aramid, PVA, or cellulose fibers and that its continued use in asbestos-cement products and friction materials is not justifiable in the face of available technically adequate substitutes.

Expression of c-myc, c-myb, and c-sis in fibroblasts from affected and unaffected skin of patients with systemic sclerosis.

We examined c-sis, c-myc, and c-myb proto-oncogene expression in fibroblasts cultured from affected and unaffected skin of patients with systemic sclerosis (SSc), and from healthy donor skin. Total cellular RNA from cultured dermal fibroblasts was used in slot blot analysis and scanning densitometry or phosphorimaging to quantify steady-state levels of proto-oncogene mRNAs. PDGF B-chain levels in culture supernatants of fibroblasts were determined by ELISA. Our results demonstrate that steady-state levels of c-myc and c-myb mRNA were elevated 1.5- to 5.6-fold in intralesional fibroblasts from SSc patients as compared to other cells examined. Levels of c-sis mRNA and PDGF-B protein were comparable regardless of source. Elevated c-myc and c-myb expression may be indicative of, and may contribute to, fibroblast activation in SSc.

Control of IL-6 expression and response in fibroblasts from patients with systemic sclerosis.

Systemic sclerosis (SSc) is an autoimmune connective tissue disease of unknown etiology in which aberrant fibroblast function results in fibrosis of the skin and internal organs. A distinguishing feature of dermal fibroblasts cultured from SSc lesions is that they produce constitutively, i.e., without exogenous stimulation, as much as 30-fold more interleukin-6 (IL-6) than do normal fibroblasts. The present study indicates that the mechanism of constitutive IL-6 secretion involves the accumulation of IL-6 mRNA in affected SSc fibroblasts, mediated by the constitutive binding of nuclear factors to the IL-6 promoter. DNA-protein complexes formed using nuclear extracts of constitutively expressing cells are distinct from those using extracts of normal cells, with or without exogenous stimulation of IL-6; thus, the mechanisms which regulate constitutive and inducible IL-6 gene expression are apparently distinct. The data also demonstrate that dermal fibroblasts respond very rapidly to IL-6 by increasing expression of the IL-6 gene, thus suggesting a mechanism for the establishment and/or persistence of constitutive expression. The constitutive secretion of IL-6 may play an important role in the perpetuation of the local immune dysregulation and fibroblast activation in the SSc lesion.

B-cell leukemia in a rhesus macaque (Macaca mulatta) infected with simian immunodeficiency virus.

Conditions associated with abnormal B-cell proliferation have an increased incidence in the HIV-infected population. A longitudinal study conducted at the Tulane Regional Primate Research Center has followed more than 1,000 rhesus macaques infected with simian-immunodeficiency virus (SIV) since 1984. While spontaneous B-cell malignancy in SIV-negative macaques has not been reported, 42 cases of SIV-associated-lymphoma (SAL) have been documented in this cohort. Recently we identified a single case of B-cell leukemia, first suggested by clinical abnormalities and confirmed and further characterized by molecular analysis. The case is important because it models the occurrence of B-cell leukemia in the human AIDS patient and because it extends our understanding of the B-cell lymphoproliferative diseases associated with AIDS.

Hexavalent chromium produces DNA strand breakage but not unscheduled DNA synthesis at sub-cytotoxic concentrations in hepatocytes.

Rat hepatocytes were used to investigate the possible induction of unscheduled DNA synthesis (UDS) and the extent of DNA strand breaks induced by sodium dichromate (a representative chromium(VI) compound) and chromium acetate hydroxide (chromium(III)) in vitro. Cytotoxicity, measured using tetrazolium salt (MTT) reduction assay, was found at a much higher dose of chromium(III), (> 50 microM), compared to that of chromium(VI), (> 2.5 microM), in cultured hepatocytes over 20 h treatment at 37 degrees C. Chromium(VI), but not chromium(III), stimulated minimal UDS in hepatocytes at sub-cytotoxic concentrations. A positive UDS response was only observed at cytotoxic concentration. DNA strand breaks in hepatocytes were induced by chromium(VI) following incubation at 37 degrees C for 1 h at doses of 10, 20 and 40 microM sodium dichromate. The subsequent ligation of such strand breaks in hepatocytes treated with 40 microM chromium(VI) for 1 h at 37 degrees C was demonstrated. The majority of strand breaks was repaired within 30 min following removal of the chromate. In conclusion, chromate-induced DNA strand breakage, possibly involving the formation of oxygen radicals and lack of significant UDS have some analogy to those produced by ionizing radiation.