Induction of Th1 chemokine secretion in dermal fibroblasts by vanadium pentoxide.

Vanadium is a soft, silvery­grey metal with a number of different oxidation states. The most common commercial form of vanadium is vanadium pentoxide (V2O5). All vanadium compounds are considered toxic. An increase in skin rashes has been observed in certain vanadium workers, including the development of atopic dermatitis. However, to the best of our knowledge, no prior in vivo or in vitro studies have evaluated the effect of vanadium exposure in human dermal fibroblasts. The present study evaluated the effect of V2O5 on proliferation and chemokine secretion in dermal fibroblasts. The results revealed that V2O5 had no significant effect on the viability or proliferation of fibroblasts, however it was able to induce the secretion of T­helper (Th)1 chemokines from dermal fibroblasts, synergistically increasing the effect of important Th1 cytokines, including interferon­Î³ and tumor necrosis factor­α. Through these processes, V2O5 may lead to the induction and perpetuation of an inflammatory reaction in dermal tissue. The induction and perpetuation of inflammation in the dermis and the variety of involved candidate genes may be at the base of V2O5­induced effects following occupational and environmental exposures. Further studies are necessary to evaluate dermal integrity and manifestations in subjects who are occupationally exposed, or living in polluted areas.

Sex-based differences in lymphocyte proliferation in the spleen after vanadium inhalation.

Vanadium (V) is a transition metal often adhered to particulate matter and released into the atmosphere as vanadium pentoxide (V2O5) by the burning of fossil fuels. This air pollutant causes adverse effects in the immune system. Lymphocytosis and splenomegaly have been reported with increased white pulp in mice after V inhalation. The effect of V on the immune system as related to sex has been poorly investigated. This study sought to determine if V inhalation (a) produced lymphoproliferation that could explain the changes previously observed in the spleen and in peripheral blood lymphocyte counts and (b) whether any observed effects differed due to gender. Immunohistochemical analyses of Ki-67, a specific proliferation marker, was made in the spleens of CD-1 male and female mice exposed for 1 h, twice a week, over a 12-week period to V2O5 (at 1.4 mg V2O5/m(3)) by whole-body inhalation; similar analyses were performed on spleens of control mice exposed to vehicle (filtered air). The results showed that in male mice there was a significant increase in percentage of Ki-67 immunopositive lymphocytes starting from the second week and until the end of the exposure. The Ki-67 signal was cytoplasmic and nuclear in the exposed males, while in controls the signal was only nuclear. In female mice, V inhalation singificantly increased the percentage of proliferating lymphocytes only after 1 week of exposure. Ki-67 signal was observed only in the nucleus of lymphocytes from the control and exposed females. The results here help to explain the splenomegaly and lymphocytosis observed previously in male mice and support the lymphoproliferative effect induced by V. Lastly, the finding that there was a sex difference in the effect of vanadium on lymphocyte proliferation suggests a role for sex hormones in potential protection against V immunotoxicity; however, further studies are needed to support this hypothesis.

An enzyme-linked immunosorbent assay (ELISA) for the determination of mucin levels in bronchoalveolar lavage fluid.

INTRODUCTION: A method to measure the mucin concentration in bronchoalveolar lavage (BAL) fluid was developed to aid efforts to identify pharmacologically the mechanisms that modulate pathophysiological mucin secretion. Mucins are the major macromolecular components of mucus. In the airways, mucus is the first line of defense against inhaled microorganisms (infection) and particulates (irritation). METHODS: An enzyme-linked immunosorbent assay (ELISA) was developed, comparing two monoclonal anti-mucin antibodies (A10G5 and 45M1) raised to human mucin, to quantify the mucin in BAL fluid from animal models of pulmonary inflammation. To validate the ELISA method, rats were exposed to ovalbumin (OVA, in sensitized rats), lipopolysaccharide (LPS), vanadium pentoxide (V(2)O(5)), or saline. One hundred microliters of BAL fluid was analyzed for mucin concentration. Pooled BAL fluid from untreated rats was used as an internal "plate standard", as a standard mucin that cross-reacts with A10G5 was unavailable. RESULTS: We found both antibodies reacted with rat, human, and guinea-pig mucin; where the 45M1 antibody also reacted with the mucin in porcine BAL, while A10G5 did not. We determined the mucin concentration in each BAL fluid sample relative to the standard, defined as a mucin concentration of 100 plate units. BAL fluid from LPS (218+/-25 plate units, n=5), OVA (386+/-31, n=3), V(2)O(5) (1208+/-450, n=6) challenged rats displayed significantly elevated mucin concentration over their saline controls (126+/-22, n=12). Subsequently, the 45M1 antibody displayed immunoreactivity with a commercially available crude preparation of porcine stomach mucin, allowing us to calculate the concentration of mucin directly compared to the known concentration of the porcine stomach mucin standard. Both the 45M1 and A10G5 based ELISA assays detected higher mucin content in the saline challenged rat than the saline challenged guinea pig BAL. DISCUSSION: The recent availability of the 45M1 antibody and the use of the crude purification of porcine stomach mucin as a reference standard should allow for direct comparison of mucin concentration in BAL (and other fluids).

Induction of the lung myofibroblast PDGF receptor system by urban ambient particles from Mexico City.

Platelet-derived growth factor (PDGF) and its receptor system regulate mesenchymal cell proliferation. We recently reported that emission-source fly-ash particles and asbestos fibers induce the PDGF alpha-receptor through a macrophage-dependent pathway, and upregulation of this receptor greatly enhances the mitogenic response of lung myofibroblasts to PDGF (Lindroos and colleagues, Am. J. Respir. Cell Mol. Biol. 1997;16:283-292). In the present study we investigated the effect of particulate matter <= 10 micrometers in size (PM10) from the southern, central, and northern regions of Mexico City on PDGF receptor induction and compared these urban, ambient particles with Mt. St. Helen's volcanic ash particles as a negative control. All Mexico City PM10 samples, but not volcanic ash, stimulated rat alveolar macrophages to secrete a soluble, upregulatory factor(s) for the PDGF alpha-receptor on early passage rat lung myofibroblasts. The macrophage-derived upregulatory activity was blocked by the interleukin (IL)-1 receptor antagonist. The ability of PM10 to stimulate IL-1beta release was blocked in part by a recombinant endotoxin neutralizing protein (rENP). Lipopolysaccharide/endotoxin (LPS) and vanadium, both constituents that were present within these PM10 samples, also stimulated macrophages to secrete factor(s) that upregulated PDGF-Ralpha on lung myofibroblasts. Direct exposure of myofibroblasts to PM10 also elicited upregulation of the PDGF alpha-receptor, and this effect was blocked by rENP and mimicked by LPS, but not vanadium. These findings suggest that PM10 particles induce expression of the PDGF receptor system through macrophage-dependent and -independent mechanisms involving endotoxin and metals.