Immunolocalization of pro-inflammatory cytokines in normal and inflamed hoof of buffalo calves.

Laminitis is characterized by inflammation of the sensitive laminae of the hoof, loss of lamellar basement membrane and infiltration of mononuclear inflammatory cells in the laminar tissue. Buffaloes commonly suffer from subclinical mastitis resulting insidious economic losses to the farmers in terms of decreased milk yield and weight gain and increased cost of treatment. Effective tools to treat lameness in buffaloes are still lacking due to incomplete understanding of its complex patho-physiology. Moreover, there is paucity of data on the in situ localization of macrophages, pro-inflammatory cytokines, TLR-4 and TLR9 in inflamed hoof of buffalo calf. The present study identifies immunopositive endothelial cells, macrophages, IL8, IL-1?, TNF-? and TLR9 cells in the normal and inflamed hoof of buffalo calves. Labeling of PIMs with the anti-human macrophage antibody (MCA874G) demonstrated frequent occurrence of these cells in inflamed hoof. Further, there was increased immunopositive expression of IL8, IL-1?, TNF-? and TLR9 cells in the inflamed hoof of buffalo calf. TLR4 mRNA expression showed almost six fold significant increase (p<0.05) in the inflamed hoof compared to normal hoof. We present first data on the immunolocalization of macrophages, various pro-inflammatory cytokines, TLR4 and TLR9 in the normal and inflamed hoof of buffalo calves. The data taken together suggest that there is increase in the immunopositivity for various pro-inflammatory cytokines along with TLR4 and TLR9 in the inflamed buffalo calf hoof indicating that activation of TLRs may contribute significantly to the pathogenesis of hoof inflammation in buffalo calves.