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A bacterium that was able to utilize Emkarate 1550 (E1550), a synthetic lubricant ester, as the sole source of carbon was isolated. The isolate was tentatively identified as Micrococcus roseus. The components of the E1550 ester, octanoate, decanoate, and 1,1,1-tris(hydroxymethyl)propane (TMP), were detected in the culture medium of cells growing on the ester. The TMP tertiary alcohol accumulated during growth and was not utilized by this isolate. The detection of the components of the ester in the supernatant of cultures indicated that one of the first steps in its degradation was cleavage of the ester bonds. Esterase activity was significantly enhanced in cells grown on E1550 compared with esterase activity measured in cells grown on acetate.

Expression of granulocyte-macrophage colony-stimulating factor (GM-CSF) by metastatic Lewis lung carcinoma cells (LLC-LN7) was previously shown to contribute to the maintenance of phenotypic characteristics associated with an increased capacity to metastasize. In the present study, pre-incubation of LLC-LN7 cells with neutralizing anti-GM-CSF antibodies diminished the capacity of the tumor cells to form experimental metastases after i.v. inoculation, while pre-incubation with recombinant GM-CSF (rGM-CSF) increased formation of metastases. In the presence of rGM-CSF, the LLC-LN7 cells exhibited an increased capacity to migrate, invade through a reconstituted basement membrane, and adhere to lung tissue. Studies to identify the signal transduction pathway through which GM-CSF enhanced the in vitro metastatic properties of the LLC-LN7 tumor cells implicated protein kinase A (PKA). Signaling through PKA was suggested by the demonstration that the stimulation of tumor-cell motility by GM-CSF was blocked in the presence of the adenylate cyclase inhibitor nicotinic acid, or the PKA inhibitors A3 or KT5720. In addition, the role of PKA as a signaling mechanism for GM-CSF was assessed by using REV-LN7 cells, which are LLC-LN7 cells that have been stably transfected with an expression vector encoding a mutant PKA RI alpha subunit and which, in turn, express a cAMP-resistant PKA. Adherence and invasion by the PKA-defective REV-LN7 cells were not stimulated by rGM-CSF, contrasting with the stimulation observed for wild-type LLC-LN7 cells. These data suggest that rGM-CSF can further enhance the in vitro metastatic characteristics of LLC-LN7 tumor cells and that this is dependent on signal transduction through PKA.

Production of granulocyte-macrophage colony-stimulating factor (GM-CSF) by metastatic Lewis lung carcinoma tumors (LLC-LN7) has been shown to increase their migration and invasion in vitro, and metastasis in vivo. The present studies showed that treatment of the LLC-LN7 cells by 2 days culture with 1 alpha, 25-dihydroxyvitamin D3 [1,25(OH)2D3] inhibited their production of GM-CSF. The 1,25(OH)2D3-treated cells also became less migratory and invasive. This inhibitory effect on tumor motility could not be readily reversed upon removal of 1,25(OH)2D3. The mechanism by which 1,25(OH)2D3 reduced tumor motility was attributed to the inhibition of tumor GM-CSF production since the capacity to migrate and invade could be restored by supplementing the medium with recombinant GM-CSF. In vivo studies showed that treatment of LLC-LN7-bearing mice with 1,25(OH)2D3 had no effect on growth of s.c. primary tumors, but reduced the formation of tumor metastases. These results show that 1,25(OH)2D3 can interrupt the autocrine motility-stimulation cascade by reducing tumor production of GM-CSF, thus reducing the expression of properties that are characteristic of metastatic tumor cells.

Metastatic Lewis lung carcinoma (LLC) tumors stimulate myelopoiesis and, consequently, induce bone marrow cells to become immune suppressive to T cell blastogenesis and macrophage activation for tumor necrosis factor alpha (TNF-alpha) secretion. The suppressor cells phenotypically resembled granulocytic-monocytic progenitor cells. In order to diminish the presence of these immune suppressor cells, LLC-bearing mice were treated with low doses of gamma interferon (IFN-gamma) (100 units/mouse) plus TNF-alpha (10 units/mouse). Treatment of LLC-bearing mice with these low doses of IFN-gamma plus TNF-alpha diminished the suppressive activity of their bone marrow cells, as measured by the effect on normal macrophage activation to secrete TNF-alpha. In in vivo adoptive transfer studies, bone marrow from placebo-treated LLC-bearers stimulated tumor establishment and metastasis, while the bone marrow of IFN-gamma-plus TNF-alpha-treated tumor-bearers diminished LLC establishment and metastasis. The effect of the low dose treatments with IFN-gamma and/or TNF-alpha on the recurrence of excised s.c. tumors was also assessed. Treatment of mice following tumor excision with either IFN-gamma, TNF-alpha, or the combination of IFN-gamma plus TNF-alpha reduced recurrence. However, in the animals with recurring tumors only the combined IFN-gamma plus TNF-alpha treatment effectively diminished the development of lung metastases. These results demonstrate that low dose IFN-gamma plus TNF-alpha treatment diminishes the presence of suppressor and tumor growth-promoting activities of bone marrow and reduces tumor recurrence and metastasis.

Objective: To provide economic, epidemiologic, and clinical data on initial and subsequent hospitalizations for firearm injuries., Design: Nonconcurrent prospective study; data obtained by medical records review., Setting: Public university teaching hospital, designated a level I trauma center., Subjects: Two hundred fifty persons first hospitalized for firearm injuries at UC Davis Medical Center, Sacramento, between January 1, 1984, and June 30, 1985, and followed by medical records review to June 30, 1989., Results: The aggregate hospital charge for 250 firearm injuries, exclusive of professional fees, was $3,745,496, of which 80% was borne directly or indirectly by public funds. The charge for initial hospitalizations was $3,297,506. Mean and median initial charges per case were $13,190 and $5,996 respectively; range, $787-$494,152. The five patients with charges over $100,000 accounted for 33% of all charges; 36% of all patient days were attributable to the eight patients with hospitalizations lasting more than 30 days. Thirty-one patients were rehospitalized a total of 71 times; charges for rehospitalization totaled $447,990. Three fourths of all charges resulted from handgun injuries., Conclusions: The costs for hospital treatment of firearm injuries are substantial. Avoiding prolonged hospitalization may be helpful in controlling these costs, but will be difficult to achieve. Primary prevention of firearm injuries may be the most effective cost-control measure.

Using both polymerase-chain-reaction analysis and the soft-agar colony-forming unit assay, granulocyte-macrophage colony-stimulating factor (GM-CSF) was shown to be expressed by cloned metastatic Lewis-lung-carcinoma (LLC-LN7) cells but not by non-metastatic LLC-C8 cells. Furthermore, the metastatic LLC-LN7 cells were shown to respond both to autologous GM-CSF and to exogenous recombinant GM-CSF (rGM-CSF). In the presence of neutralizing anti-GM-CSF antibodies, the metastatic LLC cells became less able to migrate or to adhere and invade through a reconstituted basement membrane. Moreover, the addition of rGM-CSF further enhanced the capacity of the metastatic LLC cells to adhere to the reconstituted basement membrane. This stimulation of metastatic properties of the LLC cells by either autologous or exogenous GM-CSF was associated with enhanced endogenous protein phosphorylation. Two proteins of approximately Mr 45,000 and Mr 64,000 were the dominant target proteins to be phosphorylated by the presence of GM-CSF. These results suggest that autologous GM-CSF may function as an autocrine stimulator of the metastatic properties of metastatic LLC cells.

The modulatory effects of serotonin on T-cell activity were investigated. T-cell blastogenesis of normal spleen cells was slightly stimulated by the addition of low doses (1 and 10 ng/ml) of the inducer of serotonin release, fenfluramine. In contrast to the stimulatory effects of low doses of fenfluramine, high doses of fenfluramine (1 and 10 ug/ml) or of exogenously added serotonin (> or = 0.1 ug/ml) inhibited T-cell activation. Both the stimulation by low dose fenfluramine and the inhibition by high dose fenfluramine were accentuated by pretreating mice with tryptophan to heighten intracellular stores of serotonin and then inducing serotonin release. Pretreatment of mice with the serotonin inhibitor p-chlorophenylalanine (PCPA) abolished the fenfluramine inhibition of T-cell activation indicating that the fenfluramine inhibitory effect was mediated via endogenous spleen cell-derived serotonin. However, the PCPA treatment diminished T-cell activation. These results suggest that endogenous serotonin causes a biphasic dose-response effect on T-cell activity with serotonin being required for optimal T-cell function, low doses being immune stimulatory and higher doses being suppressive.

Normal bone marrow cells become immunosuppressive when cultured with supernatants of metastatic Lewis lung carcinoma (LLC-LN7) cells. The suppressor-inducing activities in the LLC-LN7 supernatants are interleukin-3 and granulocyte/macrophage-colony-stimulating factor. In the present study, the mechanisms by which these induced suppressor cells (LLCsup-BM) mediate their immunosuppression were investigated. The suppression by LLCsup-BM of splenic concanavalin CA blastogenesis was not dependent on cell contact since immunosuppression occurred regardless of whether the LLCsup-BM were separated from the responder spleen cells by a permeable membrane or if the LLCsup-BM were cocultured with the spleen cells. Culture supernatants of LLCsup-BM also inhibited T cell blastogenesis, being more suppressive than were supernatants of control bone marrow cells, which had been precultured with medium. The suppression by the soluble inhibitors elaborated from the LLCsup-BM was not restricted to the inhibition of T cell function as the supernatants also inhibited the natural killer activity of normal spleen cells. Studies to determine the identity of the suppressive activity produced by the LLCsup-BM showed increased levels of transforming growth factor beta (TGF beta) in their supernatants. Immunosuppressive bone marrow and spleen cells obtained from mice bearing metastatic LLC-LN7 tumors also secreted more TGF beta than did the cells obtained from normal mice. When anti-TGF beta antibodies were added to the LLCsup-BM supernatants, the suppressive activity was diminished. These results suggest that the LLCsup-BM mediate at least part of their immunosuppression through production of TGF beta.

Complete pool fencing and effective bystander resuscitation are both believed to reduce the risk of childhood drowning. The relationship between support for, and prevalence of, a complete pool barrier and current cardiopulmonary resuscitation certification was investigated among an equal probability sample of 795 owners of residential swimming pools in Sacramento County, California. Only 50% (95% confidence interval [CI] 44%, 56%) of respondents who favored a cardiopulmonary resuscitation certification requirement for pool owners represented a household with any members so certified. Only 35% (95% CI 26%, 44%) of respondents who endorsed a complete barrier requirement for all pools had a fence surrounding their own pool. Support for a cardiopulmonary resuscitation requirement was associated with a modestly higher prevalence of current cardiopulmonary resuscitation certification (46% vs 33%, difference = 13%, 95% CI for difference 2%, 24%). Endorsement of a pool barrier requirement was associated with a substantially higher prevalence of complete pool fencing (35% vs 7%, difference = 28%, 95% CI for difference 19%, 37%). The proportion of the pool-owning population endorsing these risk reduction behaviors is much larger than the proportion actually adopting them. The results suggest that an effective pool drowning-prevention program will rely primarily on legislative approaches, with health education serving as a useful adjunct.

Progressive growth of metastatic Lewis lung carcinoma (LLC) tumors results in a concurrent stimulation of myelopoiesis and the appearance of immune-suppressive bone marrow cells. The present study has shown that normal bone marrow cells could be induced to become immune-suppressive by 3 days of culture with supernatants of cloned metastatic LLC-LN7 variant cells. The capacity of the LLC-LN7 supernatants to stimulate the appearance of suppressor cells was directly proportional to the concentration of supernatant used in the bone marrow culture. When adoptively transferred with a LLC-LN7 tumor inoculum, the supernatant-induced suppressor bone marrow cells increased the rate of appearance of palpable tumors and the frequency of tumor establishment. The LLC-LN7 supernatants containing suppressor-cell-inducing activity also had colony-stimulating factor (CSF) activity. The CSF activity produced by the LLC-LN7 cells could be diminished with neutralizing antibodies to either granulocyte/monocyte(GM-) CSF or to interleukin-3 (IL-3). Likewise, the suppressor-inducing activity in the LLC-LN7 supernatants was diminished by pretreatment with anti-GM-CSF or anti-IL-3. The combination of anti-GM-CSF and anti-IL-3 completely neutralized all suppressor-inducing activity produced by the LLC-LN7 cells. These results suggest that the secretion of IL-3 and GM-CSF by LLC-LN7 tumor cells is a mechanism by which the tumors stimulate myelopoiesis and induce normal bone marrow cells to become immune-suppressive. Bone marrow cells that are induced to become immune-suppressive by culture with LLC-LN7 supernatants can, in turn, facilitate the establishment of tumor in vivo.

We present an epidemiologic characterization of deaths from motor vehicle immersions, based on 77 deaths in 63 motor vehicle immersions in Sacramento County, CA, during 1974-85. All persons were autopsied; all but one of the deaths were due to drowning. Average annual mortality rates per million person years were 12 for males, four for females, 30 for Hispanics, six for White non-Hispanics, Blacks and Asians. Seventy-one percent of drivers and 60 percent of passengers had a blood alcohol concentration greater than or equal to 22 mmol/L. Most cases (57 percent) had an Injury Severity Score of 1 (minor injury) or 0 (no injury). Alcohol use was associated with higher Injury Severity Scores. Road curvature of 20 degrees or greater was far more common at crash sites than at matched control sites one mile away (OR = 6.57, 95% CI = 2.93, 14.71). Guard rail placement along highly curved sections of roadway may be an effective preventive measure.

The myelopoietic stimulation which occurs in mice bearing metastatic Lewis lung carcinoma (LLC-C3) tumors is accompanied by immune suppression and the appearance of myelopoiesis-associated immune suppressor cells in the bone marrow and spleen. Low doses of recombinant murine interferon-gamma (IFN-gamma) plus recombinant human tumor necrosis factor-alpha (TNF-alpha) were used to limit myelopoiesis and, in turn, reduce the presence of myelopoiesis-associated immune suppressor cells in LLC-C3 tumor bearers. Neither IFN-gamma nor TNF-alpha alone had any effect in vitro on the growth of myeloid progenitor cells into colonies or on the suppressive activity of bone-marrow cells from LLC-C3-bearing mice. However, the combination of low doses of IFN-gamma and TNF-alpha synergistically inhibited both the growth of myeloid progenitor cells into colonies and the suppressive activity of bone-marrow cells from tumor-bearers. Similar results were obtained in vivo. When used alone, neither IFN-gamma nor TNF-alpha had any effect on myelopoiesis or on suppressor-cell activity. When combined, IFN-gamma plus TNF-alpha synergistically suppressed myelopoiesis and the presence of immune suppressive cells both in the bone marrow and in the spleen of tumor bearers. T-lymphocyte blastogenic and NK cytotoxic activities of the tumor-bearers were restored only after treatment with both IFN-gamma and TNF-alpha.

In previous studies, we demonstrated that immune-suppressive bone marrow cells appeared during a period of myelopoietic stimulation in mice bearing Lewis lung carcinoma (LLC) tumors and could be induced from normal bone marrow cells during 3 days of culture with supernatants of LLC variant cells. We have now shown that the capacity of LLC variants to induce immune suppressor cells was associated with the capacity to produce colony-stimulating factor (CSF) activities. The LLC variants that secreted more CSF activities also produced more bone marrow immune suppressor cell-inducing activity. The induction of immune-suppressive bone marrow cells was dependent on bone marrow cell proliferation because irradiation of bone marrow cells prior to culture with the CSF-containing supernatants blocked induction of suppressor cells. Culture supernatants of WEHI-3 cells and of pokeweed mitogen-stimulated spleen cells, rich sources of interleukin 3 (IL-3) and granulocyte-monocyte-CSF (GM-CSF), as well as recombinant mouse GM-CSF and recombinant mouse IL-3, also stimulated bone marrow suppressor cell activity. The combination of GM-CSF with IL-3 resulted in a synergistic rather than simply an additive induction of bone marrow suppressor cells. Conditioned medium of CSF-1-producing L cells did not induce bone marrow immune suppressor cells. These results suggest that during heightened periods of myelopoiesis, induction of bone marrow-derived immune suppressor cells may be stimulated by CSFs, such as by the singular or combined effects of GM-CSF and IL-3.

Cloned Lewis lung carcinoma (LLC) variants were used in an in vitro migration model for dissemination, to determine if prostaglandin E2 (PGE2) produced by nonmetastatic LLC cells could directly stimulate dissemination of metastatic LLC cells and to identify an intracellular mechanism for such an effect. The migration of metastatic LLC clones was stimulated not only by exogenous PGE2 but also by nonmetastatic LLC cells, by their production of a migration-stimulatory factor which was sensitive to indomethacin and anti-PGE2 antibodies. Nonmetastatic LLC clones were unresponsive to migration stimulation by PGE2. The results of in vivo metastasis studies were consistent with those of in vitro migration studies. In vivo lung metastasis was increased by PGE2, as well as by nonmetastatic cells when they were either admixed with the metastatic LLC inoculum, irradiated and injected adjacent to the metastatic LLC tumor, or localized in chambers and implanted s.c. into mice given injections of metastatic LLC cells. Indomethacin blocked metastasis stimulation by nonmetastatic cells. The in vitro PGE2 stimulation of metastatic LLC cells appeared to be linked to a cyclic AMP (cAMP) response, since migration could also be stimulated by dibutyryl-cyclic AMP and blockage of a cAMP response with nicotinic acid ablated the PGE2 stimulation of migration. In vivo metastasis could be stimulated by elevation of cAMP with aminophylline. The differential responsiveness of metastatic versus nonmetastatic LLC cells to PGE2 could not be due to PGE2-adenylate cyclase coupling, since PGE2 increased the cAMP levels in cultures of both metastatic and nonmetastatic LLC cells. There was, however, a difference in the cyclic AMP-dependent protein kinase (PKA) response to PGE2, with PKA activity of metastatic LLC being stimulated by PGE2 and by the adenylate cyclase-stimulator forskolin, whereas PKA of nonmetastatic LLC was not stimulated by these cAMP elevators, suggesting a dysfunction in the cAMP-PKA coupling.

Training in cardiopulmonary resuscitation for pool owners and mandatory placement of a barrier around pools, two commonly suggested strategies for prevention of drowning, will depend, in part, on the support of pool owners to be successfully implemented. To measure this support, an equal probability sample of 795 Sacramento County households with pools was surveyed. An 80% response rate was achieved. A large majority (86%) favored voluntary cardiopulmonary resuscitation training, and a plurality (40%) favored required cardiopulmonary resuscitation certification for pool owners. However, 61% opposed a universal barrier requirement, and 49% objected to a barrier requirement for new pools only. Respondents with small children at home were more likely (P = .0001) to support a required barrier. The previous occurrence of a significant immersion event had surprisingly little effect. The results suggest some specific directions for programs to prevent swimming pool drownings.

We conducted a two-part study of unintentional firearm deaths in California. First, we analyzed death certificate data for the 688 unintentional firearm deaths of California residents occurring during 1977-1983. Mortality rates were 7.5 for males, 0.9 for females, 4.8 for whites, and 5.3 for blacks, all per million persons per year. Males ages 15-24 had the highest rate (17 per million persons per year). We then investigated the 131 childhood deaths in greater detail, using coroners' or medical examiners' reports. Most of these shootings occurred at a residence. Handguns were involved more frequently than predicted by their reported availability in homes in the region. Almost two thirds of child deaths resulted from head wounds, reinforcing the importance of primary prevention. At least 40% of child deaths in this study appeared in part to be attributable to defects in firearm performance or current firearm design practices, suggesting that improvements should be sought and mandated.

Between 1977 and 1983, eighty-eight California children 0 to 14 years of age were unintentionally shot and killed either by other children or by themselves. The majority of cases occurred while children were playing with guns they had found. Easy accessibility to guns, the resemblance of guns to toys, and gun malfunctions were all contributing factors. Handguns, particularly those of .22 caliber, were frequently involved.

Previous epidemiological studies of drowning in the United States have dealt primarily with children. The epidemiology and prevention of drowning in adulthood may be very different. To test this general hypothesis, we analyzed the 293 drownings occurring among Sacramento County residents 20 years of age and above during 1974 to 1985. Drowning rates were highest for men 20-29 years of age (11.5 per 100,000 population) and blacks (7.5 per 100,000 population). Swimmers, boaters, and motor vehicle occupants were most frequently represented. Alcohol use was involved in 48% of cases overall and 77% of those involving motor vehicle occupants. A history of seizure disorder was another contributing factor. Important differences do exist in drowning epidemiology between children and adults. Our results suggest that preventing drowning will be more problematic among adults than among children. The study was also used to test the sensitivity of two commonly used methods of case ascertainment for cases in Sacramento County. A manual review of coroner's records had a sensitivity of 96%. A computerized review of death certificate data from the state's vital statistics data base had a sensitivity of 79%. The sources and implications of these differences are discussed.

The adrenocorticotrophic activities of two synthetic polypeptides, Synacthen (Ciba) and DW 75 (Sandoz), were compared in two groups of patients having a variety of arthritic disorders. Compared with Synacthen, injected either intramuscularly or intravenously, intravenous DW 75 was found to have a longer duration of adrenocorticotrophic activity (measured by plasma 11-hydroxycorticosteroid levels) in both groups of patients, while intramuscularly administered DW 75 did not appear to be absorbed well.