Abstract
Abstract: :
Purpose: The purpose of this study was to localize and quantitate pro-inflammatory [substance P (SP)] and anti-inflammatory [vasoactive intestinal peptide (VIP)] neuropeptides in susceptible (C57BL/6, B6) vs. resistant (BALB/c) mice after P. aeruginosa. infection. Methods: Corneas and cervical lymph nodes (CLN) from both groups were removed before and at 1 and 7 days post-infection (p.i.) with 1.0x106 CFU P. aeruginosa, ATCC 19660. Frozen corneal and CLN sections were immunostained with antibody to SP and VIP. Corneas and CLN also were removed similarly for neuropeptide protein quantitation by competitive ELISA. Appropriate staining and ELISA controls were included for similar analysis. Results: Immunostaining for SP-containing nerve fibers showed higher intensity in uninfected B6 compared to BALB/c cornea. Staining intensity for VIP was similar in both mouse strains. SP staining in uninfected CLN was similar in both groups, but VIP staining was greater in BALB/c mice. These results were confirmed by ELISA analysis. Corneal immunostaining for SP was of increased intensity subepithelially and in the superficial stroma at 1 day p.i. and in the stroma of B6 mice at 7 days p.i. VIP immunostaining was similar in the cornea of both mouse strains at day 1 p.i., but was of greater intensity in the BALB/c cornea at day 7 p.i. SP and VIP staining of CLN was similar at day 1 p.i. At day 7 p.i. immunostaining for SP was more intense in B6 mice, while VIP staining was more intense in BALB/c mice. ELISA analysis for SP confirmed the corneal immunostaining data and showed significantly (p<0.05) increased protein levels of SP in B6 cornea at both time points. There was no difference in VIP levels at day 1 p.i., but increased protein levels were seen in BALB/c cornea at day 7 p.i. (p<0.05). ELISA analysis of CLN showed similar levels of SP and VIP in B6 and BALB/c mice at 1 day p.i., but at 7 days p.i. SP levels were higher in B6 and VIP levels were higher in BALB/c CLN (p<0.05 for both). Conclusion: Overall, in cornea and CLN B6 mice appear more SP (pro-inflammatory) regulated, while BALB/c mice appear more VIP (anti-inflammatory) regulated. Support: RO1EY02986; P30EY04068.
Keywords: Pseudomonas • neuropeptides • immunohistochemistry