Kelly S. Warmington

Effect of C-C chemokine receptor 2 (CCR2) knockout on type-2 (Schistosomal antigen-elicited) pulmonary granuloma formation. Analysis of cellular recruitment and cytokine responses

Monocyte chemotactic protein (MCP)-1 is postulated to play a role in cellular recruitment during inflammatory reactions. C-C chemokine receptor 2 (CCR2) is considered the major G-protein coupled receptor for MCP-1/JE. We reported that mice with knockout of the CCR2 gene display partially impaired type-1 granuloma formation. The present study similarly examined the effect of CCR2 deficiency on synchronously developing type-2 (Th2) cytokine-mediated lung granulomas elicited by embolization of beads coated with Ags of Schistosoma mansoni eggs. Systemically, blood monocytes were reduced by about half throughout the 8-day study period. At the local level, granuloma size and macrophage content were impaired during the early growth phase (days 1 to 2). By day 4, granuloma sizes were similar to controls. In granulomatous lungs, CCR2 knockout increased mRNA for CCR2 agonists, MCP-1, MCP-3, and MCP-5, but reduced IL-4 and IFNgamma mRNA. The latter was possibly related to decreased CD4+ T cell recruitment. Regionally, draining lymph nodes showed panlymphoid hyperplasia with impaired production of IFNgamma, IL-2, and IL-4, but not IL-5, IL-10, or IL-13. Analysis of procollagen gene expression indicated transient impairment of procollagen III transcripts on day 4 of granuloma formation. These findings indicate that agonists of CCR2 contribute to multiple facets of type-2 hypersensitivity granulomatous inflammation.

Role of Macrophage Inflammatory Protein (Mip)-Lα in Granulomatous Inflammation

Macrophage inflammatory protein (MIP)-1 is a low MW, LPS-inducible monocyte and neutrophil chemotactic cytokine which may be important in inflammation and pulmonary granuloma formation. The present study examined the kinetic generation, and in vivo relevance of murine MIP- lα utilizing synchronous pulmonary Schistosoma mansoni egg granulomas. Antigenic MIP-lα was measured in 24 hr supernatants from whole granulomas cultured (700/ml) with or without soluble egg antigen (SEA) utilizing an ELISA developed in our laboratory. Intact primary granulomas isolated at various times of development from normal mice showed low backgroud levels of MIP- lα production (<1 ng/ml), however, when challenged with antigen demonstrated significant production of MIP-lα beginning at day 8 (2.6 ng/ml) and peaking at day 16 (16.8 ng/ml). Intact pulmonary granulomas isolated from acutely infected mice demonstrated high backgroud levels of MIP-lα production (peaking at day 2; 7.8 ng/ml). Likewise, background levels from chronic infection granulomas (day 2; 5.6 ng/ml) were similar to acute granulomas. Antigen stimulation increased expression of MIP- lα at all time points with granulomas from acutely infected (peaking at day 2; 16 ± 4.6 ng/ml) but not chronically infected (peaking at day 8; 3.45 ± 1.3 ng/ml) mice. Treatment of mice with polyclonal rabbit anti-mouse MIP-lα (6,125 ± 310 um2) abrogated 8 day primary pulmonary granuloma formation when compared to normal serum control group (12,704 ± 1154 um2). Anti-MIP-lα sera also decreased granuloma formation in lungs of acutely (4-day; 27,897 ± 2400 um2), but not chronically (4-day; 15257 + 1058 um2) infected mice compared to normal serum treated groups (36,010 ± 2507 and 17319 ± 1016 um2, respectively).