In Vivo Activation of the Intracrine Vitamin D Pathway in Innate Immune Cells and Mammary Tissue during a Bacterial Infection

Numerous in vitro studies have shown that toll-like receptor signaling induces 25-hydroxyvitamin D3 1α-hydroxylase (1α-OHase; CYP27B1) expression in macrophages from various species. 1α-OHase is the primary enzyme that converts 25-hydroxyvitamin D3 to 1,25-dihydroxyvitamin D3 (1,25(OH)2D3). Subsequently, synthesis of 1,25(OH)2D3 by 1α-OHase in macrophages has been shown to modulate innate immune responses of macrophages. Despite the numerous in vitro studies that have shown 1α-OHase expression is induced in macrophages, however, evidence that 1α-OHase expression is induced by pathogens in vivo is limited. The objective of this study was to evaluate 1α-OHase gene expression in macrophages and mammary tissue during an in vivo bacterial infection with Streptococcus uberis. In tissue and secreted cells from the infected mammary glands, 1α-OHase gene expression was significantly increased compared to expression in tissue and cells from the healthy mammary tissue. Separation of the cells by FACS9 revealed that 1α-OHase was predominantly expressed in the CD14+ cells isolated from the infected mammary tissue. The 24-hydroxylase gene, a gene that is highly upregulated by 1,25(OH)2D3, was significantly more expressed in tissue and cells from the infected mammary tissue than from the healthy uninfected mammary tissue thus indicating significant local 1,25(OH)2D3 production at the infection site. In conclusion, this study provides the first in vivo evidence that 1α-OHase expression is upregulated in macrophages in response to bacterial infection and that 1α-OHase at the site of infection provides 1,25(OH)2D3 for local regulation of vitamin D responsive genes

SMOKE AND VIRAL-INFECTION CAUSE CILIA LOSS DETECTABLE BY BRONCHOALVEOLAR LAVAGE CYTOLOGY AND DYNEIN ELISA

The purpose of this study was to quantitate cilia loss following airway epithelial cell injury. Two models of airway injury were used: (1) Ex vivo acute cigarette smoke exposure model: Bovine lungs, obtained directly after slaughter, were ventilated with air or cigarette smoke for 5 min followed immediately by bronchoalveolar lavage (BAL). The bronchi were examined histologically and bronchial and alveolar fractions of BAL fluid were examined for cell counts, cell differentials, and cilia dynein concentrations using a specific 13S dynein ELISA. Smoke exposure resulted in a marked loss of ciliated cells from the bronchial luminal surface (2,364 +/- 351 versus 11,090 +/- 542 ciliated cells/mm2; p = 0.0001), a comparable increase in ciliated cells in the bronchial BAL fraction (0.90 x 10(6) cells/mm3 versus 0.15 x 10(6) cells/mm3; p = 0.0003) and a significant increase in bronchial fluid dynein concentrations (24.5 +/- 6.0 micrograms/ml versus 8.9 +/- 2.2 micrograms/ml; p = 0.03) compared with that in air-exposed lungs. The dynein concentrations strongly correlated with the absolute number of ciliated cells recovered in the bronchial lavage (r = 0.80; p < 0.0001). (2) In vivo viral infection model: Healthy cattle underwent bronchoscopy 3 days before and 7 days after inoculation with bovine respiratory syncytial virus (BRSV). BAL fluid was examined as in the first model. Following BRSV inoculation, airway exfoliation of ciliated cells and squamous metaplasia were observed histologically, bronchial ciliated cell counts doubled (0.011 +/- 0.003 x 10(6) cells/mm3 versus 0.026 +/- 0.006 x 10(6) cells/mm3; p = 0.002) and bronchial dynein concentrations increased threefold (2.2 +/- 1.0 micrograms/ml versus 7.2 +/- 1.9 micrograms/ml; p = 0.02).(ABSTRACT TRUNCATED AT 250 WORDS