Agricultural safety efforts by county health departments in Wisconsin

OBJECTIVE: The authors sought to improve the agriculture safety prevention efforts of county health departments in Wisconsin by examining current programs, staffs' perceptions of the farm safety problem, and the need for new resources. METHODS: A survey instrument was completed by a professional staff member of the local health department in each of Wisconsin's 69 counties. RESULTS: Usable responses were obtained from 84% of the counties. Forty-five percent of the responding staff members conducted some agricultural safety and health programs, most often health screenings or group meetings conducted collaboratively with county agricultural Extension agents. There were no major differences in county demographics or other service provision variables between staff members who conducted programs and those who did not. Staff members perceived the largest barriers to better safety as lack of staff time and difficulty getting farmers to attend safety programs. Most failed to place more emphasis on training agricultural workers to permanently correct hazards than on training them to work safely around hazards. However, the staff members ranked safety inspection checklists as the most needed new material and ranked Extension agents and farmers as the most appropriate people to conduct inspections using such checklists. CONCLUSION: County public health professionals want more staff time and new materials to increase the effectiveness of their agricultural safety efforts. Encouraging agricultural workers and family members to identify and correct hazards would be a more effective use of staff time than training people to work safely around hazards.     

Studies on the mechanism of resistance to rapamycin in human cancer cells

Rapamycin is a potent cytostatic agent that arrests cells in the G1 phase of the cell cycle. The relationships between cellular sensitivity to rapamycin, drug accumulation, expression of mammalian target of rapamycin (mTOR), and inhibition of growth factor activation of ribosomal p70S6 kinase (p70(S6k)) and dephosphorylation of pH acid stable protein I (eukaryotic initiation factor 4E binding protein) were examined. We show that some cell lines derived from childhood tumors are highly sensitive to growth inhibition by rapamycin, whereas others have high intrinsic resistance (>1000-fold). Accumulation and retention of [14C]rapamycin were similar in sensitive and resistant cells, with all cells examined demonstrating a stable tight binding component. Western analysis showed levels of mTOR were similar in each cell line (<2-fold variation). The activity of p70(S6k), activated downstream of mTOR, was similar in four cell lines (range, 11.75-41. 8 pmol/2 x 10(6) cells/30 min), but activity was equally inhibited in cells that were highly resistant to rapamycin-induced growth arrest. Rapamycin equally inhibited serum-induced phosphorylation of pH acid stable protein I in Rh1 (intrinsically resistant) and sensitive Rh30 cells. In serum-fasted Rh30 and Rh1 cells, the addition of serum rapidly induced c-MYC (protein) levels. Rapamycin blocked induction in Rh30 cells but not in Rh1 cells. Serum-fasted Rh30/rapa10K cells, selected for high level acquired resistance to rapamycin, showed >/=10-fold increased c-MYC compared with Rh30. These results suggest that the ability of rapamycin to inhibit c-MYC induction correlates with intrinsic sensitivity, whereas failure of rapamycin to inhibit induction or overexpression of c-MYC correlates with intrinsic and acquired resistance, respectively.     

Shelf life of sliced raw potatoes of various cultivar varieties--substitution of bisulfites

Treatment of colorectal liver metastases with the HSVtk/GCV approach and adenoviral vectors is highly toxic. We present a nontoxic alternative using the cell type-specific CEA promoter instead of the widely used hCMV immediate-early promoter to drive tk gene expression in the context of a recombinant adenovirus. Analysis of CEA promoter-dependent tk gene expression showed significant activity of this promoter in several human and rat tumor-derived cell lines but not in rat primary hepatocytes and in mouse liver, whereas the CMV promoter was highly active in all cell types and tissues investigated. CEA promoter-dependent tk gene expression was sufficient to kill 100% of cancer cells in vitro, even if less than 10% were infected by the adenoviral vector, indicating a significant bystander effect. Moreover, treatment of subcutaneous tumors in SCID mice with Ad.CEA-tk led to a several-fold reduction of tumor growth, and tail vein injection of a high dose of Ad.CEA-tk caused no side-effects in the liver. The CMV promoter was more potent than the CEA promoter in mediating GCV sensitivity to cancer cells in vitro and in vivo, but even a 20-fold reduction of the dose of Ad.CMV-tk did not prevent its liver cell toxicity after systemic application to mice and still resulted in the death of all animals within 4 days after the start of GCV treatment. These results indicate that restriction of tk gene expression to tumor cells in the liver prevents systemic toxicity. Moreover, the CEA promoter is a safe and efficient tool for tumor cell-specific expression of suicide genes in the liver.     

Type I IFNs inhibit human dendritic cell IL-12 production and Th1 cell development

We have investigated the role of type I IFNs (IFN-alpha and -beta) in human T cell differentiation using anti-CD3 mAb and allogeneic, in vitro-derived dendritic cells (DC) as APCs. DC were very efficient activators of naive CD4+ T cells, providing necessary costimulation and soluble factors to support Th1 differentiation and expansion. Addition of IFN-alphabeta to DC/T cell cultures resulted in induction of T cell IL-10 production and inhibition of IFN-gamma, TNF-alpha, and LT secretion. Diminished T cell IFN-gamma production correlated with IFN-alphabeta-mediated inhibition of the p40 chain of the IL-12 heterodimer secreted by DC. Suppression of p40 IL-12 and IFN-gamma was not due to increased levels of IL-10 in these cultures, and production of IFN-gamma could be restored by exogenous IL-12. These data indicate that type I IFNs inhibit DC p40 IL-12 expression, which is required for development of IFN-gamma-producing CD4+ T cells. Furthermore, when T cells were restimulated without IFN-beta, these cells induced less p40 IL-12 from DC, suggesting that the functional properties of T cells may regulate DC function. Thus, IFN-alphabeta inhibits both IL-12-dependent and independent Th1 cytokine production and provides a mechanism for inhibition of IL-12-mediated immunity in viral infections.     

Molecular characterization of cDNAs encoding low-molecular-weight heat shock proteins of soybean

Three cDNA clones (GmHSP23.9, GmHSP22.3, and GmHSP22.5) representing three different members of the low-molecular-weight (LMW) heat shock protein (HSP) gene superfamily were isolated and characterized. A fourth cDNA clone, pFS2033, was partially characterized previously as a full-length genomic clone GmHSP22.0. The deduced amino acid sequences of all four cDNA clones have the conserved carboxyl-terminal LMW HSP domain. Sequence and hydropathy analyses of GmHSP22, GmHSP22.3, and GmHSP22.5, representing HSPs in the 20 to 24 kDa range, indicate they contain amino-terminal signal peptides. The mRNAs from GmHSP22, GmHSP22.3, and GmHSP22.5 were preferentially associated in vivo with endoplasmic reticulum (ER)-bound polysomes. GmHSP22 and GmHSP22.5 encode strikingly similar proteins; they are 78% identical and 90% conserved at the amino acid sequence level, and both possess the C-terminal tetrapeptide KDEL which is similar to the consensus ER retention motif KDEL; the encoded polypeptides can be clearly resolved from each other by two-dimensional gel analysis of their hybrid-arrest translation products. GmHSP22.3 is less closely related to GmHSP22 (48% identical and 70% conserved) and GmHSP22.5 (47% identical and 65% conserved). The fourth cDNA clone, GmHSP23.9, encodes a HSP of ca. 24 kDa with an amino terminus that has characteristics of some mitochondrial transit sequences, and in contrast to GmHSP22, GmHSP22.3, and GmHSP22.5, the corresponding mRNA is preferentially associated in vivo with free polysomes. It is proposed that the LMW HSP gene superfamily be expanded to at least six classes to include a mitochondrial class and an additional endomembrane class of LMW HSPs.     

Identification of an Arabidopsis thaliana mutation (vsm1) that restricts systemic movement of tobamoviruses

Following inoculation, many plant viruses spread locally from cell to cell until they reach the vascular system, through which they then move to other parts of the plant, resulting in systemic infection. To isolate host genes involved in systemic transport of plant viruses, ethyl methanesulfonate-mutagenized Arabidopsis thaliana plants were screened for significant delays in the systemic movement of turnip vein clearing virus (TCVC). One such mutant, designated vsm1 (virus systemic movement), was identified. Unlike the wild-type plants, vsm1 did not develop viral disease and did not allow the systemic spread of the virus. The local viral movement within the inoculated vsm1 leaves, however, was not affected. TVCV systemic movement within the vsm1 plants was likely blocked at the step of viral entry into the host plant vasculature from the infected leaf tissue. vsm1 plants also restricted the systemic movement of another tobamovirus but not of an unrelated carmovirus.     

Correlation of nicotinic binding with neurochemical markers in Alzheimer's disease

The loss of neocortical synapses that occurs in Alzheimer's disease (AD) has been shown to correlate with cognitive decline. In addition, marked losses in the cholinergic system in AD, specifically choline acetyltransferase (ChAT) activity and high affinity presynaptic neuronal nicotinic cholinergic receptors (nAChRs), have also been described. We hypothesized that in AD, the loss of [3H]-ligand binding to nAChRs, which are largely presynaptic, would correlate with changes in two other presynaptic markers: synaptophysin (Syn), a measure of synaptic density, and ChAT activity. The midfrontal (MF) cortex of 36 autopsy confirmed (NIA and CERAD criteria) AD patients (mean death age +/- SD 80.1 +/- 8.4 years) who met NINDS-ADRDA criteria for a clinical diagnosis of probable or possible AD, and 11 nondemented controls (mean death age +/- SD 77.9 +/- 8.0) were examined. Synapse counts were quantified by a dotimmunobinding assay for Syn. ChAT activity was assessed by standard biochemical assays. Nicotinic cholinergic receptor binding was assayed using the high affinity nicotinic agonist [3H]-(+/-)-epibatidine ([3H]-EPI). The mean +/- SD Syn in AD (83.4 +/- 31.9 arbitrary units (AU)/mg protein) was significantly lower than controls (126.1 +/- 19.9, p = 0.0003; t-test). The mean ChAT activity in AD (139.0 +/- 75.6 nmol ACh/hr/100 mg protein) was significantly lower than controls (219.6 +/- 70.8, p = 0.004). The mean [3H]-EPI total binding in AD (6.2 +/- 2.8 fmol/mg protein) was significantly lower than controls (14.8 +/- 3.2; p < 0.0001). Syn correlated with [3H]-EPI binding in AD (r = 0.48, p = 0.006; Pearson) but ChAT did not (r = -0.20, p = 0.34). We conclude that loss of high affinity nAChR binding correlates with loss of synapses in AD. The lack of correlation between [3H]-EPI binding and ChAT activity suggests that the targeted receptor populations may not be located exclusively on cholinergic neurons.