Evaluation of the effects of short-chain fatty acids and extracellular pH on bovine neutrophil function in vitro.
OBJECTIVE
To investigate the effects of short-chain fatty acids (SCFAs) and pH on neutrophil oxidative burst, phagocytosis, and morphology after exposure to acetate, propionate, butyrate, or succinate at pH 5.5 and 6.7.


METHODS
Neutrophils isolated from bovine blood samples and Porphyromonas levii, Prevotella spp, and Bacteroides fragilis isolated from lesions of cattle with acute interdigital phlegmon (foot rot).


METHODS
Bacteria were cultured in strictly anaerobic conditions. Bacterial SCFA production was measured with high-performance liquid chromatography. Neutrophils were isolated, stimulated with phorbol 12-myristate 13-acetate (PMA) or opsonized zymosan (OZ), and incubated with dihydroethidium or dichlorofluorescein diacetate to measure production of O(2)and H(2)O(2), respectively. Phagocytosis was assessed after exposure to serum-opsonized bacteria. Cellular morphology was assessed with differential staining.


RESULTS
All bacteria produced at least 3 of the 4 SCFAs. Production of both O(2) and H(2)O(2) was markedly curtailed in PMA-stimulated neutrophils exposed to SCFA at pH 5.5, compared with production at pH 6.7. Succinate caused a significant dose-dependent decrease in O(2) production at pH 6.7 in OZ-stimulated neutrophils. Monoprotic SCFAs elicited a significant increase in H(2)O(2) production in OZ-stimulated neutrophils at pH 6.7 but a significant decrease at pH 5.5. Monoprotic SCFAs significantly increased phagocytosis at pH 6.7 but decreased phagocytic activity at pH 5.5. Cellular necrosis was observed in cells exposed to SCFAs at pH 5.5.


CONCLUSIONS
Establishment and persistence of anaerobic bacteria in cattle with foot rot infection may result in part from neutrophil dysfunction secondary to the effects of bacterially secreted SCFA in acidotic microenvironments.
Mills
Steven W
SW
Department of Biological Sciences, Faculty of Science, University of Calgary, Calgary, AB T2N 1N4, Canada.
Montgomery
Shaylyn H
SH
Morck
Douglas W
DW
D000704:Analysis of Variance; D000818:Animals; D001441:Bacteroides fragilis / Q000737:chemistry*; D002417:Cattle; D002418:Cattle Diseases / Q000276:immunology* / Q000382:microbiology*; D002481:Cellulitis / Q000276:immunology / Q000382:microbiology / Q000662:veterinary*; D002851:Chromatography, High Pressure Liquid / Q000662:veterinary; D005232:Fatty Acids, Volatile / Q000302:isolation & purification / Q000633:toxicity*; D006861:Hydrogen Peroxide / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D009504:Neutrophils / Q000166:cytology / Q000187:drug effects* / Q000276:immunology; D010587:Phagocytosis / Q000187:drug effects; D017861:Porphyromonas / Q000737:chemistry*; D018720:Prevotella / Q000737:chemistry*; D016897:Respiratory Burst / Q000187:drug effects
D005232:Fatty Acids, Volatile; D006861:Hydrogen Peroxide