Nils-Gunnar Ilbäck

Exercise in coxsackie B3 myocarditis: Effects on heart lymphocyte subpopulations and the inflammatory reaction

To investigate whether exercise in coxsackie B3 virus infection is detrimental to the myocardium, Balb/c mice were inoculated with the virus and exercised to exhaustion on a motor-driven treadmill up to 48 hours after the inoculation. This infection evokes myocarditis. The inflammatory and necrotic lesions in the ventricular myocardium 7 days after the inoculation covered 4.32% of the tissue section area in the nonexercised group. Exercise at 0 hours did not affect this myocardial damage (4.77%), whereas exercise at 48 hours after the inoculation increased the lesion to 7.85% (p less than 0.05). Lethality was not influenced by exercise. The response pattern of myocardial lymphocyte subpopulations was studied with an immune histochemical staining technique. The number of T cytotoxic, T suppressor cells increased threefold (p less than 0.01), and the T cytotoxic, suppressor/T helper cell ratio increased twofold (p less than 0.01) with exercise at 48 hours but was unchanged with exercise at 0 hours. The number of class II expressing cells decreased with exercise at 48 hours (p less than 0.05) and was negatively correlated (p less than 0.01) with the size of the inflammatory reaction. The development of myocardial inflammatory and necrotic lesions seems to be dependent on the presence and cooperation of class II expressing cells and T killer cells. Furthermore, failure to restrict physical activity in the acute phase of this infection may well contribute to the progression of the disease.

Methyl mercury exposure via placenta and milk impairs natural killer (NK) cell function in newborn rats

The effect of methyl mercury (MeHg) exposure (3.9 micrograms/g diet) on the development of immune function was studied in the newborn Sprague-Dawley rat after MeHg exposure via placenta and/or milk. No consistent alterations were observed between control and treated offspring (at the age of 15 days) on the following parameters: body weights, lymphoid organ weights or cell number, and the lymphoproliferative response to B-cell mitogen. The lymphoproliferative response to T-cell mitogen was increased in thymocytes (by 30-48%), but decreased in splenocytes (by 30-32%). This decreased activity was only observed in the groups exposed during lactation. White blood cell counts (WBC) were increased in all groups. Natural killer (NK) cell activity was reduced (by 42%, P less than 0.01) in the group that was exposed both via placenta and milk. These results indicate that placental and lactational transfer of MeHg does adversely affect the developing immune system of the rat.

Influence of sodium selenite on 203Hg absorption, distribution, and elimination in male mice exposed to methyl203Hg.

To study the effects of long-term selenium supplementation on absorption, distribution, and elimination of methylmercury (MeHg) in mice, three groups of male mice (Balb/c CA) were exposed for 7 wk to 0, 0.6, and 3 ppm sodium selenite in tap water. They were then given a single oral dose of Me203Hg (2 μmol/kg) by gastric intubation, and elimination of203Hg was followed by whole-body counting for 49 d at the same Se exposure as previously. Twenty-four hours and 49 d after dosage, 6–7 animals/group were sampled for analysis of203Hg distribution in the body. Glutathione peroxidase (GSH-PX) activity in blood and selenium levels in the liver were used as measures of selenium status. Gastrointestinal absorption of Me203Hg was not influenced by the Se status of the animals. Selenium supplementation of MeHg-exposed mice caused an enhanced whole-body elimination of Hg, but selenium-supplemented animals did not have lower Hg levels in the brain and kidney than nonsupplemented animals. The effect of selenium on the accumulation, of Hg in the brain was dose-dependent, a high dose (3 ppm Se) causing a higher initial accumulation of Hg. The intracellular distribution of203Hg in the liver and kidney was not affected by Se. The results indicate that selenium treatment of MeHg-exposed mice may have a positive effection the health of the animals by decreasing the total body burden of MeHg.

Vaccination of Balb/c mice against enteroviral mediated myocarditis

A non-virulent strain of Coxsackie B3 (CB3) virus was used to produce a subunit vaccine. It contains the capsid proteins VP1, 2, 3 and probably 4 and can be made RNA-free. It is based on the ISCOM technology ensuring non-toxic properties and good adjuvant effect. Vaccinated animals at doses above 16 ng were completely protected from mortality when challenged with a myocarditic strain of CB3. Histologically no inflammatory lesions were found in the heart. This was corroborated using immune histological techniques with monoclonal antibodies against lymphocyte subsets. Even at a dose of 0.16 ng a delayed mortality was observed. Neutralizing antibody titres rose to 512, thus ensuring a circulating level well above that considered protective. It is suggested that vaccination might be a possible way of prophylaxis for myocarditis and even dilated cardiomyopathy, the latter presently being the chief cause of heart transplantation. By persistence or triggering of autoimmune phenomena Coxsackie virus is incriminated as the first step in pathogenesis.

Trace element distribution in heart tissue sections studied by nuclear microscopy is changed in Coxsackie virus B3 myocarditis in methyl mercury-exposed mice.

Methyl mercury (MeHg) has been shown to change Coxsackie virus type B3 (CB3) myocarditis in a direction compatible with the development of chronic disease. Murine models of CB3 myocarditis closely mimic the pathogenesis in humans. There are also indications that metals, such as mercury, and trace elements may interact and adversely affect viral replication and development of inflammatory lesions. The effects of low-dose MeHg exposure on myocardial trace element distribution, as determined by means of nuclear microscopy, was studied in CB3 myocarditis. Balb/c mice were fed a MeHg-containing diet (3.9 mg/kg diet) for 12 wk prior to infection. Areas of inflammatory lesions in the myocardium were identified by traditional histologic examination, and serial tissue sections in these selected areas were used for immune histology (macrophages), in situ hybridization of virus genomes, and nuclear microscopy of tissue trace element distribution. Areas with no inflammation or virus were compared with areas of ongoing inflammation and viral replication. In the inflammatory lesions of MeHg-exposed mice as compared to nonexposed mice, the myocardial contents of calcium (Ca), manganese (Mn), and iron (Fe) were significantly increased, whereas the zinc (Zn) content was decreased. The increased Ca and decreased Zn contents in the inflamed heart may partly explain a more severe disease in MeHg-exposed individuals. Although not significant in the present study, with a limited number of mice, the inflammatory and necrotic lesions in the ventricular myocardium on d 7 of the infection was increased by 50% (from 2.2% to 3.3% of the tissue section area) in MeHg-exposed mice and, also, there was a tendency of increased persistence of virus with MeHg exposure. No increased MeHg uptake, either in the inflammatory lesions or in the areas of noninflamed heart tissue in infected mice, could be detected. The present results indicate that a “competition” exists between potentially toxic heavy metals from the environment/diet and important trace elements in the body and that a disturbed trace element balance adversely influences the development of pathophysiologic changes in inflammatory heart disease.

Effects of2,3,7,8-tetrachlorodibenzo-p-dioxin on blood and spleen natural killer (NK) cell activity in the mouse

The immunotoxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) were studied in male A/J mice after a loading dose of 5 micrograms TCDD/kg body wt. followed by 3 weekly maintenance doses of 1.42 micrograms TCDD/kg b.w. administered intraperitoneally. Tissue samples and immune cells were prepared on two occasions, i.e. on days 28 and 120 after the first injection of TCDD. This dose of TCDD evoked classical histological signs of liver damage and lipid accumulation, as well as thymic atrophy. Red (RBC) blood cell counts were significantly lowered in the TCDD group on day 28, but were normal on day 120. White (WBC) blood cell counts were normal in the TCDD group. Natural killer (NK) cell activity increased 3.4-fold (P less than 0.01) and 2.2-fold (P less than 0.01) in the blood and spleen, respectively, after 28 days, and these effects persisted on day 120. The increased NK-cell activity occurred concomitantly with a decreased proliferative response of spleen lymphocytes to the T-cell mitogen concanavalin A after both 28 (65%) and 120 days (58%). The proliferative response of spleen cells to the B-cell mitogen lipopolysaccharide seemed, however, unaffected. We have thus shown for the first time that TCDD induces an increased activity of NK cells that occurs simultaneously in the blood and spleen. This effect may indicate a general compensatory activation of the bodys defences brought about by disturbances in the function of other arms of the immune system.

Interactions Among Infections, Nutrients and Xenobiotics

During recent years there have been several incidents in which symptoms of disease have been linked to consumption of food contaminated by chemical substances (e.g., 2,3,7,8-tetrachlorodibenzo-p-dioxin, TCDD). Furthermore, outbreaks of infections in food-producing animals have attracted major attention regarding the safety of consumers, e.g., Bovine Spongiform Encephalitis (BSE) and influenza in chicken. As shown for several xenobiotics in an increasing number of experimental studies, even low-dose xenobiotic exposure may impair immune function over time, as well as microorganism virulence, resulting in more severe infectious diseases and associated complications. Moreover, during ongoing infection, xenobiotic uptake and distribution are often changed resulting in increased toxic insult to the host. The interactions among infectious agents, nutrients, and xenobiotics have thus become a developing concern and new avenue of research in food toxicology as well as in food-borne diseases. From a health perspective, in the risk assessment of xenobiotics in our food and environment, synergistic effects among microorganisms, nutrients, and xenobiotics will have to be considered. Otherwise, such effects may gradually change the disease panorama in society.

Effects of selenium supplementation on virus-induced inflammatory heart disease

The effects of 10 wk of selenium (Se) supplementation (5 ppm) in drinking water on immune responses and resistance to a myocarditic Coxsackie virus B3 (CB3) infection were studied in female Balb/c mice. Se supplementation reduced CB3-induced mortality: at day 14 postinoculation, survival was 58% in the Se-treated group as compared to 25% in the untreated group. Whole-blood glutathione peroxidase (GSH-Px) activity was elevated by 68% (p < 0.001) and Se content in the liver by 24% (p < 0.001). Red (RBC) and white blood cell (WBC) counts, as well as the number of cells in the spleen and thymus, were unaffected. The cellular counts of T-lymphocytes (CD4+, CD8+) and natural killer (NK+) cells in the blood were not affected. However, the CD4+/CD8+ ratio (5.2) tended to increase after Se supplementation (5.9). The spleen lymphoproliferative response to T and B-cell mitogens were increased by 9 and 43%, respectively (ns), in the Se-supplemented group. The total NK cell activity in blood and spleen showed minor increases, but when the activity in the blood was expressed per cell, the increase amounted to 35% (ns) with Se supplementation. The inflammatory and necrotic lesions in the ventricular myocardium at 7 and 14 d postinoculation were not significantly reduced by Se treatment, probably owing to the increased survival with Se even of mice with the most pronounced heart damage; comparable untreated mice were estimated to have died at day 14. Results indicate that modest doses of Se can improve immune function, which may increase the general resistance to this viral infection.

Altered distribution of 109cadmium in mice during viral infection

The distribution of the toxic heavy metal cadmium (Cd) was studied in Coxsackie virus B3 (CB3)-infected Balb/c mice by whole-body autoradiography and gamma-counting. The distribution of 109Cd was studied 4 days post CB3-inoculation and 10 min after intravenous injection of 0.21 microgram of Cd/kg body weight. Whole-body autoradiography results showed that the distribution of 109Cd is greatly changed during this viral infection. This newly discovered distribution was mainly visible as a greatly increased accumulation in the renal and adrenal cortices. After impulse counting of selected organs it was found that the normal accumulation of 109Cd in the kidneys (184,354 +/- 30,961 c.p.m.) was increased by 47% (P less than 0.05) during CB3 infection (270,503 +/- 54,780 c.p.m.). In contrast to healthy animals, some infected mice showed accumulation of 109Cd in the spleen. These results show for the first time that an invading micro-organism can change the distribution of an environmental pollutant.

A common viral infection can change nickel target organ distribution.

The autoradiographic distribution of the toxic heavy metal nickel (Ni) was studied at 4 and 7 days post-coxsackievirus B3 (CB3) infection in Balb/c mice. The distribution of the iv injected 63Ni was studied 10 min, 4 hr, and 24 hr after administration. Results clearly show that the site of 63Ni accumulation is greatly changed during this viral infection. This newly discovered distribution was mainly visible as a greatly increased accumulation in the pancreas and the wall of the ventricular myocardium. Healthy animals showed almost no 63Ni accumulation in these tissues. These results for the first time show that an invading microorganism can change the distribution of an environmental pollutant.