MACROPHAGES AND B-1 CELL CYTOKINES PROFILE IN AN IN VITRO MODEL OF TOLERANCE TO LPS

BARBEIRO D. F.(1); BARBEIRO H. V.(1), SORIANO F.G.(1)

(1)LIM 51 - Emergency Medicine Department.  University of São Paulo, Brazil

In this work we aimed to characterize the production of cytokines and nitric oxide (NO) by macrophages and B-1 cells using an in vitro model of tolerance to LPS.  Cell cultures were obtained by peritoneal lavage of Xid (B-1 cells free) and Balb/c mice. Macrophages (MA), macrophages + B-1 (FA), and B-1 cells were maintained in culture  (106cells/mL) and the media was changed daily. Three groups were studied: GC- control; G1- LPS (10mg/mL) added in the last 24h of cells culture; G2- LPS (10mg/mL) added in the last 48h of cells culture. Cytokines (pg/ml) and NO2 were measured in the supernatant. We found high levels of TNF-a in the G1 for B-1 (97±47), MA (1877±404) and FA (871±129) when compared to GC (34±3; 117±23; 51±7 respectively) and G2 (29±2; 65±5; 63±2 respectively). B-1 cells IL-6 production was increased in G1 (88±15) compared to GC (5±1) and G2 (14±2). For FA, the groups were also markedly different: GC (50±20) < G1 (142±8) < G2 (341±29). No differences were found for MA. IL-10 concentration in G1 was increased for FA (210±26) and MA (236±47) when compared to GC (99±20; 85±29 respectively) or G2 (99±5; 106±19 respectively). B-1 showed higher levels in the G1 (168±24) and G2 (172±22) groups, when compared to GC (72±7). In FA, we observed an increase in NO2 (mM) quantification when G2 (4±0,1) was compared to GC (3,7±0,05). In MA, G1 (4,8±0,2) and G2 (4,5±0,2) were higher than GC (3,8±0,5). No difference was found for B-1. Our data suggest that the three cell populations develop tolerance to LPS in vitro and that B-1 cells can modulate the role of macrophages to LPS, reducing TNF-a production and sustaining high IL-10 production, after the second LPS challenge.

KEY WORDS: Tolerance, LPS, Macrophages, B-1 cell

CORRESPONDENCE TO:

Denise Frediani Barbeiro, Avenida Dr. Arnaldo, 455, sala 3134. Faculdade de Medicina, USP, São Paulo, (SP),  LIM 51. CEP 01246-903. Email: denisedna@hotmail.com