Reduced endoplasmic reticulum (ER)-to-Golgi protein trafficking contributes to ER stress in lipotoxic mouse beta cells by promoting protein overload.

AIMS/HYPOTHESIS: Saturated fatty acids augment endoplasmic reticulum (ER) stress in pancreatic beta cells and this is implicated in the loss of beta cell mass that accompanies type 2 diabetes. However, the mechanisms underlying the induction of ER stress are unclear. Our aim was to establish whether saturated fatty acids cause defects in ER-to-Golgi protein trafficking, which may thereby contribute to ER stress via protein overload. METHODS: Cells of the mouse insulinoma cell line MIN6 were transfected with temperature-sensitive vesicular stomatitis virus G protein (VSVG) tagged with green fluorescent protein to quantify the rate of ER-to-Golgi protein trafficking. I14 antibody, which detects only correctly folded VSVG, was employed to probe the folding environment of the ER. ER stress markers were monitored by western blotting. RESULTS: Pretreatment with palmitate, but not oleate, significantly reduced the rate of ER-to-Golgi protein trafficking assessed using VSVG. This was not secondary to ER stress, since thapsigargin, which compromises chaperone function by depletion of ER calcium, markedly inhibited VSVG folding and promoted strong ER stress but only slightly reduced protein trafficking. Blockade of ER-to-Golgi protein trafficking with brefeldin A (BFA) was sufficient to trigger ER stress, but neither BFA nor palmitate compromised VSVG folding. CONCLUSIONS/INTERPRETATION: Reductions in ER-to-Golgi protein trafficking potentially contribute to ER stress during lipoapoptosis. In this case ER stress would be triggered by protein overload, rather than a disruption of the protein-folding capacity of the ER.

Endoplasmic reticulum stress contributes to beta cell apoptosis in type 2 diabetes.

AIMS/HYPOTHESIS: Increased lipid supply causes beta cell death, which may contribute to reduced beta cell mass in type 2 diabetes. We investigated whether endoplasmic reticulum (ER) stress is necessary for lipid-induced apoptosis in beta cells and also whether ER stress is present in islets of an animal model of diabetes and of humans with type 2 diabetes. METHODS: Expression of genes involved in ER stress was evaluated in insulin-secreting MIN6 cells exposed to elevated lipids, in islets isolated from db/db mice and in pancreas sections of humans with type 2 diabetes. Overproduction of the ER chaperone heat shock 70 kDa protein 5 (HSPA5, previously known as immunoglobulin heavy chain binding protein [BIP]) was performed to assess whether attenuation of ER stress affected lipid-induced apoptosis. RESULTS: We demonstrated that the pro-apoptotic fatty acid palmitate triggers a comprehensive ER stress response in MIN6 cells, which was virtually absent using non-apoptotic fatty acid oleate. Time-dependent increases in mRNA levels for activating transcription factor 4 (Atf4), DNA-damage inducible transcript 3 (Ddit3, previously known as C/EBP homologous protein [Chop]) and DnaJ homologue (HSP40) C3 (Dnajc3, previously known as p58) correlated with increased apoptosis in palmitate- but not in oleate-treated MIN6 cells. Attenuation of ER stress by overproduction of HSPA5 in MIN6 cells significantly protected against lipid-induced apoptosis. In islets of db/db mice, a variety of marker genes of ER stress were also upregulated. Increased processing (activation) of X-box binding protein 1 (Xbp1) mRNA was also observed, confirming the existence of ER stress. Finally, we observed increased islet protein production of HSPA5, DDIT3, DNAJC3 and BCL2-associated X protein in human pancreas sections of type 2 diabetes subjects. CONCLUSIONS/INTERPRETATION: Our results provide evidence that ER stress occurs in type 2 diabetes and is required for aspects of the underlying beta cell failure.

Plasma ascorbate in a population of children: influence of age, gender, vitamin C intake, BMI and smoke exposure

The objective of this study is to determine the influence of several personal and lifestyle factors on the levels of circulating vitamin C in a population of children. To accomplish this objective, blood samples were collected from 511 healthy children residing in the Greater San Juan area. The population was stratified into 4 percentile groups (approaching quartiles) according to plasma ascorbate levels from lowest to highest concentrations. Comparisons were made between percentile groups on the basis of age, gender, body mass index (BMI), dietary intake of vitamin C (corrected and uncorrected for energy intake) and exposure to environmental tobacco smoke (ETS). Smoke exposure was determined using urinary cotinine, which is a highly sensitive bioindicator for ETS. Dietary vitamin C was determined via one 24hr recall questionnaire. When all 4 percentile groups were used as a basis of comparison, no differences were noted for any of the factors between groups, however when comparing percentile group 1 (lowest blood ascorbate) to an aggregate value of percentile, groups 2-4, it was found that vitamin C intake (corrected for energy intake) paralleled blood values with a statistically significant association. Among personal and environmental factors only exposure to ETS showed a significant difference in blood levels between groups 2-4 and group 1. No differences between percentile groups were noted for age gender or BMI. These results emphasize that ETS is strongly associated with lowered blood ascorbate levels with the obvious implication of reduced antioxidant protection and increased risk of adverse health consequences.

Blood and nutritional profiles of an individual consuming a high protein-low fat diet

This is the case of a normal weight, physically active 24-year old Puerto Rican woman consuming a highly unusual diet. Through careful selection of foods, the diet contains a high percent protein, a low percent fat, adequate fiber and zero cholesterol. Popular commercial diets high in protein all contain high fat, high cholesterol and low fiber. Blood samples were taken and dietary recalls were collected for 6 consecutive days to evaluate hematological and nutritional parameters. A blood lipid profile showed low circulating levels of cholesterol and triglycerides and a beneficial HDL/LDL ratio. However, nutritional analysis revealed insufficient ingestion of vitamin D and an unhealthy balance of servings from the food pyramid. Long-term consequences of this diet could put the subject at risk for kidney and bone diseases. Immediate discontinuation of the diet is the preferred recommendation to the subject. This case report illustrates the danger of adapting a self-prescribed eating plan without the consultation of a dietitian or other qualified health professional.

Pneumocystis carinii pneumonia alters expression and distribution of lung collectins SP-A and SP-D.

Surfactant proteins SP-A and SP-D, members of the collectin family, have been shown to play a significant role in lung host defense. Both proteins selectively bind Pneumocystis carinii (PC) organisms and modulate the interaction of this pathogen with alveolar macrophages. We hypothesized that the expression and distribution of lung collectins SP-A and SP-D is altered by PC lung infection. PC organisms (2 x 10(5)) were inoculated intratracheally into C.B-17 scid/scid mice that do not require steroids for immunosuppression. Four weeks after inoculation, bronchoalveolar lavage (BAL) fluid was fractionated into three fractions-cell pellet, large aggregate (LA), and small aggregate (SA) surfactant-and each fraction was analyzed for the expression of surfactant components. In uninfected mice, the majority of SP-A (62% +/- 10%) was found in association with lipids in the LA fraction, while 55% +/- 14% of SP-D was distributed in the SA fraction. In contrast, both hydrophobic proteins SP-B and SP-C were associated exclusively with LA. PC infection resulted in major changes in the expression of all surfactant components. Total protein content of LA was unchanged by PC infection (115% +/- 18% of control), whereas SA protein content markedly increased (240% +/- 18% of control level, P <.001). In contrast, the phospholipid content of LA was significantly decreased (53% +/- 5% of control level, P <.001), whereas the SA phospholipid content of infected mice was increased (172% +/- 16% of control level, P <.001). By Western blotting, PC pneumonia (PCP) induced a 3-fold increase in the total alveolar SP-D protein that was reflected mainly in increases in SA SP-D (454% +/- 135% of control, P <.05). The total alveolar SP-A protein content was also increased in PCP because of a large increase in SP-A in SA (720% +/- 115% of control, P <.05); SP-A levels in LA were unchanged. The increases in lung collectin expression were selective, because PCP resulted in the down-regulation of both SP-B and SP-C in LA (5% +/- 2% and 13% +/- 2% of control, respectively, P <.001). We conclude that PCP induces marked elevations in alveolar collectin levels because of increased expression and accumulation of SP-A and SP-D protein in SA surfactant.

Plasma ascorbic acid concentrations in a population of rhesus monkeys (Macaca mulatta).

Although monkeys frequently are used as animal models for ascorbic acid studies whose results are extrapolated to humans, little information is available on the normal levels of this vitamin in large populations of animals classified by sex, age, or physiologic state such as pregnancy or lactation. The purpose of this report is to provide these values and compare them to the same parameters in humans, pointing out similar and dissimilar trends. Plasma samples were obtained from a troop of 167 rhesus monkeys (Macaca mulatta) and analyzed for ascorbic acid by using the 2,4-dinitrophenyl hydrazine method. Results obtained for ascorbic acid concentrations in plasma showed no differences between sexes. A significant (P< 0.0001) lowering effect was observed in aging versus young animals. Pregnant and nonpregnant females had similar ascorbate values, and lactating monkeys had slightly elevated levels. We conclude that rhesus monkeys and humans exhibit some of the same characteristics of ascorbic acid metabolism, such as an age-related decrease in ascorbate and the maintenance of these levels during lactation. However, a difference between species was noted with gender. Women maintain higher ascorbate concentrations than do men, whereas no differences in concentrations of this vitamin were observed between female and male monkeys.

Determinants of environmental tobacco smoke in a population of Puerto Rican children.

This study was designed to determine among various personal, socioeconomic, and environmental factors those which had the greatest influence on exposure to environmental tobacco smoke (ETS) in a population of children residing in a tropical environment and to compare these results with those obtained in the literature of tobacco exposed children in temperate climates. Urine specimens were collected from 606 healthy Puerto Rican children (2-12 years) living in an industrial area and analyzed for cotinine, a quantitative biomarker for exposure to ETS. Parents completed a questionnaire covering smoking habits and socioeconomic information. Seventy per cent of the children were reported to be exposed to ETS, 50% resulting from exposure to smoke from either or both parents. Major determinants to ETS exposure were found to be presence of smoker, number of smokers, identity of smoker, number of cigarettes smoked in the household and child age with the youngest children suffering twice the exposure of older children. Non-determinants were exposure to smoke other than from the parent, sex of the child, season of the year and several socioeconomic factors including civil and employment status of the mother, mother's age and educational background and whether food stamps were being received. Results of a multiple regression analysis showed that our predictors accounted for 40% of cotinine appearing in the urine. Reasons for this relatively low value may be due in part to precision of our analytic method and lower levels of ambient smoke in our population vs. others that reported higher R(2) values. Predictions from questionnaire information for high ETS exposure were not always the same as those indicated by urinary cotinine emphasizing that the bioindicator, which indicates the actual inhalation of ETS, is a better predictor of exposure than responses from a questionnaire.

Treatment of tropical sprue: the work of Dr. Bailey K. Ashford examined in retrospect.

Tropical sprue is a malabsorption syndrome that responds to treatment with folic acid and a broad-spectrum antibiotic. Eighty years ago, prior to the identification of the vitamins and the discovery of penicillin, clinical trials often consisted of best guess treatments based upon current knowledge and available technology. Dietary interventions were emerging as effective treatments for alleviation of diseases such as pellagra and beri beri. Representative of his generation of clinicians, Bailey K. Ashford, MD, one of the pivotal figures in academic medicine in Puerto Rico, carried out dietary studies in his patients with tropical sprue. This historical retrospective presents an examination of the diets used by Ashford in terms of nutrient content and comparison to current recommended dietary allowances. Results show the diets to be inadequate for sustained nutrient value and especially low in folic acid. In summary, Ashford recognized the basic causes of tropical sprue but was unable to effectively treat the syndrome due to lack of adequate resources.

P. carinii induces selective alterations in component expression and biophysical activity of lung surfactant.

Studies of Pneumocystis carinii pneumonia (PCP) suggest an important role for the surfactant system in the pathogenesis of the hypoxemic respiratory insufficiency associated with this infection. We hypothesized that PCP induces selective alterations in alveolar surfactant component expression and resultant biophysical properties. PCP was induced by intratracheal inoculation of 2 x 10(5) P. carinii organisms into C.B-17 scid/scid mice. Six weeks after inoculation, large (LA)- and small (SA)-aggregate surfactant fractions were prepared from bronchoalveolar lavage fluids and analyzed for expression of surfactant components and for biophysical activity. Total phospholipid content was significantly reduced in LA surfactant fractions from mice infected with PCP (53 +/- 15% of uninfected mice; P < 0.05). Quantitation of hydrophobic surfactant protein (SP) content demonstrated significant reductions of alveolar SP-B and SP-C protein levels in mice with PCP compared with those in uninfected mice (46 +/- 7 and 19 +/- 6%, respectively; P < 0.05 for both). The reductions in phospholipid, SP-B, and SP-C in LA fractions measured during PCP were associated with an increase in the minimum surface tension of LAs as measured by pulsating bubble surfactometer (13.1 +/- 1.1 vs. 5.4 +/- 1.8 mN/m; P < 0.05). In contrast to decreases in the hydrophobic SPs, SP-D content in the SA fraction was markedly increased (343 +/- 30% of control value; P < 0. 05) and SP-A levels in LA surfactant were maintained (93 +/- 26% of control value) during P. carinii infection. In all cases, the changes in SP content were reflected by commensurate changes in the levels of mRNA. We conclude that PCP induces selective alterations in surfactant component expression, including profound decreases in hydrophobic protein contents and resultant increases in surface tension. These changes, demonstrated in an immunologically relevant animal model, suggest that alterations in surfactant could contribute to the hypoxemic respiratory insufficiency observed in PCP.

Granulocyte-macrophage colony-stimulating factor in the innate immune response to Pneumocystis carinii pneumonia in mice.

Innate immunity plays an important role in pulmonary host defense against Pneumocystis carinii, an important pathogen in individuals with impaired cell-mediated immunity. We investigated the role of GM-CSF in host defense in a model of P. carinii pneumonia induced by intratracheal inoculation of CD4-depleted mice. Lung GM-CSF levels increased progressively during the infection and were significantly greater than those in uninfected controls 3, 4, and 5 wk after inoculation. When GM-CSF gene-targeted mice (GM-/-) depleted of CD4+ cells were inoculated with P. carinii, the intensities of infection and inflammation were increased significantly compared with those in CD4-depleted wild-type mice. In contrast, transgenic expression of GM-CSF directed solely in the lungs of GM-/- mice (using the surfactant protein C promoter) dramatically decreased the intensity of infection and inflammation 4 wk after inoculation. The concentrations of surfactant proteins A and D were greater in both uninfected and infected GM-/- mice compared with those in wild-type controls, suggesting that this component of the innate response was preserved in the GM-/- mice. However, alveolar macrophages (AM) from GM-/- mice demonstrated impaired phagocytosis of purified murine P. carinii organisms in vitro compared with AM from wild-type mice. Similarly, AM production of TNF-alpha in response to P. carinii in vitro was totally absent in AM from GM-/- mice, while GM-CSF-replete mice produced abundant TNF in this setting. Thus, GM-CSF plays a critical role in the inflammatory response to P. carinii in the setting of impaired cell-mediated immunity through effects on AM activation.

Inhibition of lung surfactant protein B expression during Pneumocystis carinii pneumonia in mice.

The pathogenesis of Pneumocystis carinii pneumonia (PCP) suggests an important role for dysfunction of the pulmonary surfactant system in the hypoxemic respiratory insufficiency associated with this infection. Surfactant protein B (SP-B) is a hydrophobic protein shown to be essential for normal surfactant function in vivo. Therefore, we hypothesized that the inhibition of SP-B expression occurs during PCP, and we tested this hypothesis in two immunodeficient animal models. PCP was induced in C.B-17 scid/scid mice by intratracheal inoculation of P. carinii organisms. Infected lung homogenates, obtained at time points up to 6 weeks after inoculation, were analyzed for SP-B and mRNA content. When a comparison was made with uninfected scid controls, the densitometric quantitation of Western blots of lung homogenates demonstrated significant reductions in 8 kd SP-B in mice infected with P. carinii 4 weeks after inoculation (16% of the control value). Northern blot analysis showed a concomitant decrease in SP-B mRNA to 24% of the control level. The decrease in SP-B and mRNA levels in lung homogenates of infected mice was reflected in lower SP-B levels in the surfactant. An enzyme-linked immunosorbent assay for the SP-B level in surfactant prepared from bronchoalveolar lavage samples of infected scid mice demonstrated a significant reduction in alveolar SP-B content (45% of the control value). In contrast to the results with SP-B, neither the SP-A protein content nor the mRNA level was significantly altered by PCP infection. To confirm these observations, SP-B expression was studied in an additional animal model of PCP. The SP-B content of lung homogenates from BALB/c mice depleted of CD4+ T cells and infected with P. carinii was also reduced (51% of the control value). We conclude that P. carinii induces selective inhibition of the expression of SP-B in two mouse models of PCP and that this down-regulation is mediated at the level of mRNA expression. Therefore, an acquired deficiency of SP-B is likely to be an important contributor to the pathogenesis of hypoxemic respiratory failure that is observed in patients with PCP.

Urokinase-type plasminogen activator in inflammatory cell recruitment and host defense against Pneumocystis carinii in mice.

Effective host defense against Pneumocystis carinii depends upon the integrated actions of inflammatory cells and mediators in the lungs. Using immunocompetent and immunosuppressed mice, our laboratory has defined inflammatory changes in the lungs in response to P. carinii. However, the essential molecules and mechanisms required for cellular recruitment and for host defense against P. carinii are undefined. We hypothesized that urokinase-type plasminogen activator (uPA), a protease intimately involved in inflammatory cell migration and activation, is required for clearance of P. carinii. To test this hypothesis in vivo, we compared the intensity of P. carinii infection and inflammation in the lungs of mice lacking the uPA gene (uPA knockout mice) and in the lungs of wild-type mice. After intratracheal inoculation with P. carinii organisms, uPA knockout mice developed uniformly heavy P. carinii pneumonia while wild-type mice cleared the P. carinii inoculum. Bronchoalveolar lavage fluid from uPA knockout mice contained significantly smaller numbers of cells than did lavage fluid from wild-type mice. We conclude that deletion of the uPA gene prevents the clearance of P. carinii and reduces inflammatory cell recruitment. Therefore, uPA is an important participant in the network of inflammatory events required for the clearance of P. carinii, confirming an important role for this molecule in pulmonary host defense against opportunistic pathogens.

Interaction of rat Pneumocystis carinii and rat alveolar epithelial cells in vitro.

During Pneumocystis carinii pneumonia, P. carinii trophic forms adhere tightly to type I alveolar epithelial cells (AECs). However, the manner in which the interaction between P. carinii organisms and AECs results in clinical pneumonia has not been explored. To investigate this interaction in vitro, we established a culture system using rat P. carinii and primary cultures of rat AECs. We hypothesized that binding of P. carinii to AECs would alter the metabolic, structural, and barrier functions of confluent AECs. Using fluorescently labeled P. carinii, we demonstrated that P. carinii bound to AECs in a dose-dependent manner. During P. carinii-AEC interaction, both the AECs and the P. carinii organisms remained metabolically active. Immunofluorescent staining demonstrated that AEC expression of the junctional proteins E-cadherin and occludin and the structural protein cytokeratin 8 were unaffected by P. carinii binding. To evaluate the effect of P. carinii on AEC barrier function, transepithelial resistance across AEC monolayers was measured during interaction with organisms. Culture with P. carinii did not result in loss of AEC barrier function but in fact increased AEC transepithelial resistance in a dose- and time-dependent manner. We conclude that the direct interaction of P. carinii with AECs does not disrupt AEC metabolic, structural, or barrier function. Therefore, we speculate that additional inflammatory cells and/or their signals are required to induce the epithelial derangements characteristic of P. carinii pneumonia.

Susceptibility to Pneumocystis carinii in mice is dependent on simultaneous deletion of IFN-gamma and type 1 and 2 TNF receptor genes.

Pneumocystis carinii pneumonia is an important cause of morbidity and mortality in immunosuppressed patients, particularly HIV-infected individuals. An improved understanding of pulmonary host response, including the cytokines required for defense, could suggest novel immunotherapeutic approaches to this infection. The cytokines IFN-gamma and TNF have contributory roles in host defense against P. carinii, but their combined and interactive importance is unclear. To test the roles of these cytokines in defense against P. carinii directly, organisms were inoculated intratracheally into wild-type mice and into three groups of gene-deleted mice: those lacking genes for IFN-gamma (IFN-gamma(-/-)), for TNF receptors 1 and 2 (TNFR(-/-)), and for both IFN-gamma and TNFR (TNFR-IFN-gamma(-/-)). Four weeks after P. carinii inoculation, lungs of the wild-type, IFN-gamma(-/-), and TNFR(-/-) mice demonstrated clearance of P. carinii and only mild inflammation. However, TNFR-IFN-gamma(-/-) mice demonstrated severe P. carinii infection and lung inflammation. Our findings demonstrate conclusively that deletion of either IFN-gamma or TNF activity alone does not block clearance of P. carinii. However, simultaneous deletion of IFN-gamma and TNF receptor genes results in susceptibility to P. carinii. Rather than focusing exclusively on individual cytokines, our data suggest that immunotherapy targeted at maximizing both the IFN-gamma and TNF responses to P. carinii may be required to augment host defense against this important opportunistic pathogen.

Exposure of Puerto Rican children to environmental tobacco smoke.

BACKGROUND: Many studies of environmental tobacco smoke (ETS) have been conducted in northern, industrialized countries. As yet, however, no studies have been carried out on ETS exposure with nonsmokers living in tropical environments. METHODS: Urine specimens were collected from 175 healthy Puerto Rican children (2-11 years) living in an industrial area and were analyzed for cotinine, a quantitative biomarker for exposure to ETS. Their parents completed a questionnaire covering smoking habits. RESULTS: Seventy percent of children were exposed to ETS. Quantitatively, exposure to smoke in households consuming more than 1 pack per day (ppd) caused a doubling of cotinine excretion compared with households consuming less than 1 ppd. Smoke from mothers made the greatest contribution to cotinine, followed by smoke from fathers, with smoke from other persons having no effect. Degree of exposure was inversely related to age of the child. CONCLUSIONS: Young children (2-4 years) were detected to have significantly greater exposure to ETS than older children (5-11 years) and in the younger group the effect seemed to be from the mother's smoking much more than the father's, with other persons contributing negligible amounts. This suggests an obvious strategy for prevention of exposure to ETS in young children.

Dietary analysis of meals served in the breakfast and lunch programs of Puerto Rican schools.

BACKGROUND: Nutritional analysis of meals in the Federally-sponsored Breakfast and Lunch Programs in Stateside Schools has recently been completed. However due to ethnic and cultural differences, the findings may not be directly applicable to similar nutrition programs in Puerto Rico. It is our aim to carry out an analysis of meals served in Federal programs in Puerto Rico and to compare results to the stateside study. METHODS: Twenty eight different breakfast meals and 96 different lunch meals being cycled in elementary, middle and secondary schools throughout the entire island of Puerto Rico were analyzed for content using the Minnesota Nutrition Data System 32 and compared with: 1) compliance to meal pattern requirements of federal programs, 2) dietary guidelines for Americans (DG) and 3) recommended dietary allowances (RDA's). RESULTS: Breakfasts and lunches served in Puerto Rican Schools satisfy federal meal pattern requirements however most frequently offered foods different from programs in the mainland, reflecting ethnic and cultural food preferences. In terms of DG's adequate protein was present, cholesterol content was satisfactory but meals had excess percent energy from fat as well as excess energy from saturated fat, high sodium and a lower than recommended level of energy from carbohydrate. In terms of RDA's meals had prescribed levels of vitamin A, vitamin B12, vitamin C, calcium, folacin, magnesium, phosphorus and potassium. Below prescribed levels included vitamin B6, copper, vitamin E, energy, fiber, iron, niacin and zinc. CONCLUSIONS: While differences in food preferences exist between foods available in the Breakfast and Lunch Programs in Puerto Rican and U.S., schools, they have similar strengths and weaknesses when compared to compliance with U.S. Dietary Guidelines and with recommended dietary allowances.

Regulation of interleukin 8 gene expression by oxidant stress.

Interleukin 8 (IL-8) is a recently described cytokine that functions as a potent neutrophil chemoattractant and activator. We sought to examine the link between the generation of reactive oxygen intermediates (ROI) and the regulation of IL-8 gene expression to specifically test the hypothesis that ROI would induce production of IL-8 mRNA and protein. In lipopolysaccharide-stimulated human whole blood, the OH radical scavenger dimethyl sulfoxide (Me2SO) dramatically inhibited (approximately 90%) IL-8 production, but had minimal effects on the production of tumor necrosis factor, interleukin 1 beta (IL-1), and IL-6. To determine whether NADPH-oxidase-generated free radicals were critical in the regulation of IL-8, studies were performed using blood from patients with chronic granulomatous disease. In both normal individuals and patients with chronic granulomatous disease, production of IL-8 could be initiated with lipopolysaccharide, phytohemagglutinin, or aggregated immune complexes, and this production could be inhibited by Me2SO (1% v/v). To examine if oxidant stress represents a ubiquitous mechanism for the induction of IL-8, experiments were performed in cultured cell lines. In the human hepatoma cell line Hep-G2, Me2SO dose-dependently inhibited tumor necrosis factor-stimulated IL-8 production, with a 74 +/- 1% reduction observed at a Me2SO concentration of 1%. Direct exposure to ROI demonstrated that H2O2 stimulated IL-8 production in a dose-dependent manner in Hep-G2 cells, A549 pulmonary type II epithelial cells, and human skin fibroblasts; this induction could be prevented by addition of catalase. The production of IL-8 appeared to be specific to an oxidant stress since exposure of the cells to heat shock or chemical stress did not induce expression of IL-8. These studies demonstrate that oxidant stress is an important regulator of IL-8 gene expression and support the hypothesis that low levels of ROI may serve to initiate IL-8 production which then serves to recruit neutrophils to sites of inflammation.

Glycation of blood proteins during pregnancy and lactation in the rat.

Non-enzymatic glycation of blood proteins is a time and concentration dependent process and has been used clinically to monitor carbohydrate metabolism during human pregnancy. Since gestation in rats is of much shorter duration than in humans (3 weeks vs 9 mos) the question was raised whether similar differences in glycated proteins could be observed. Therefore, levels of glucose, glycated hemoglobin and fructosamine were measured during normal pregnancy and lactation in rats. Glucose levels during late pregnancy were significantly lower than in non pregnant and early pregnant rats. During lactation glucose levels return to normal. Glycated hemoglobin paralleled glucose decrease during late pregnancy and increased during lactation. Fructosamine followed a similar pattern. Therefore glycated hemoglobin and fructosamine appear to be reliable indicators of glucose status during gestation and lactation similar to humans and may have value as predictors of gestational diabetes mellitus once a suitable rat model is developed.

Cigarette smoking-nutritional implications.

Although the effects of cigarette smoking on a variety of diseases, from cancer through emphysema and cardiovascular illness are well documented, direct effects on the levels of macro- and micronutrients in the body are reported less frequently. In fact, imbalances in these nutrients may have a role in many of the pathological conditions attributed to smoking. Tobacco smoke contains numerous compounds emitted as gases and condensed tar particles, many of them being oxidants and prooxidants, capable of producing free radicals thus enhancing lipid peroxidation in biological membranes. Vitamin E, vitamin C, B-carotene and selenium are involved in the overall cellular anti-oxidant defense against deleterious effects of reactive oxygen species. Smoking has been shown to lower the level of vitamin C and B-carotene in plasma. Cadmium, naturally found in tobacco, decreases the bioavailability of selenium and acts antagonistically to zinc, a cofactor for the antioxidant enzyme, superoxide dismutase. Vitamin E, the principle lipid-soluble antioxidant, may be at suboptimal levels in tissues of smokers. In addition, tobacco constituents have been shown to reduce levels of several vitamins of the B-complex. Nutritional status in smokers may be further compromised by an inadequate diet. Data from the Second National Health and Nutrition Examination Survey indicates that smokers are less likely to consume fruits and vegetables, particularly those high in vitamin C and carotenes. Cessation of smoking is the obvious solution to ending cigarette-related problems. In the world as it is, however, the medical community should be responsible for making recommendations to lower the risk in smokers to tobacco related diseases. Nutritionists could have a role in this process. There exists a lively debate as to where levels of nutrients should be set. Additional vitamin C has already been recommended for smokers. Should other antioxidants also be increased? Arguments for the against are considered.

Use of games to improve scores on memory-type examinations.

Memory facilitating techniques such as word games are commonly used teaching tools, but are they effective transmitters of information? To answer this question, lists of scientists were compiled and tested in matching-type exams using three different methods of exam preparation. These included two traditional methods: 1) identities of the scientists supplied (DEFINED); 2) identities of the scientists looked up in library references (LOOK UP) and; one non traditional method: the identities of the scientists were deduced by solving a word puzzle (WORD PUZZLE). Results between a pre- and post-test showed significant increases using all methods, however, WORD PUZZLE had greater increases than did DEFINED or LOOK UP. A questionnaire indicated that WORD PUZZLE was the preferred method of study and students felt equally confident using WORD PUZZLE as a method of preparation as with the two more traditional methods. It is concluded that word games should be encouraged as a memory improving method.