Monocytosis and accelerated activation of lymphocytes in C1q-deficient autoimmune-prone mice

Abstract

C1q deficiency has been shown to accelerate spontaneous autoimmunity in mice. We studied the time course of activation of monocytes and lymphocytes in autoimmune and non-autoimmune mice in the presence or absence of C1q as a disease accelerator. Autoimmune MRL\Mp.C1qa-\- and non-autoimmune C57BL\6.C1qa-\- mice were analysed at various time points between 6 and 33 weeks of age and compared to strain- and age-matched C1q-sufficient controls. Splenic and peritoneal leucocytes were analysed by flow cytometry and plasma levels of immunoglobulin M (IgM), total IgG, IgG subclasses and IgM autoantibodies were measured. Both C1q-deficient strains had significantly more splenic monocytes than their controls at all time points analysed. In addition, MRL\Mp.C1qa-\- but not C57BL/6.C1qa-\- mice developed splenic hypercellularity starting at about 12-17 weeks old, had signs of accelerated CD4+ T-cell activation and showed a marked increase in splenic plasma cells and total serum IgM levels from about 22 weeks of age. The accelerated CD4+ T-cell activation was not due to a direct inhibitory effect of C1q on T cells. These data show that C1q deficiency causes splenic monocytosis together with accelerated T-cell activation in an autoimmune-prone mouse strain.

Figure 1

Total splenic and peritoneal cell numbers in MRL/Mp versus MRL/Mp.C1qa^−/− mice at different time points between 6 and 33 weeks of age. In both groups of mice the data at each time point are expressed as mean ± SEM (**P < 0·05, ***P < 0·01).

Figure 2

Time course of relative numbers of splenic monocytes (CD11b^high CD16/32⁺ Ly6c⁺) in MRL/Mp.C1qa^−/− (b) and C57BL/6.C1qa^−/− (c) compared to their strain-matched wild-type controls. The monocyte population was isolated by gating on CD11b⁺ cells (R1), followed by selection of CD16/32⁺Ly6c⁺ cells (R2) as represented in (a). The data are expressed as mean ± SEM (*P < 0·1, **P < 0·05, ***P < 0·01).

Figure 3

Relative numbers of activated CD4⁺ T cells in spleens of MRL/Mp versus MRL/Mp.C1qa^−/− mice as characterized by loss of CD62L (a) and up-regulation of CD44 (b). Representative dotblots are shown to demonstrate how the above populations were gated during the analysis. The data are expressed as mean ± SEM (*P < 0·1, **P < 0·05, ***P < 0·01).

Figure 4

Activation of CD4⁺ splenic lymphocytes from MRL/Mp and MRL/Mp.C1qa^−/− mice after stimulation with Con A in the absence (closed symbols) or presence (open symbols) of 50 µg/ml human C1q. The level of CD4⁺ T-cell activation was assessed by measuring the percentage of CD4⁺ T cells expressing CD69 after 17 hr stimulation (a) and the percentage of cells that have divided one or more times after 65 hr stimulation (b). Data are given as mean ± SEM. All experiments were repeated at least twice.

Figure 5

Mean fluorescence intensity of CD44 on B220⁺ lymphocytes (a) and relative numbers of splenic plasma cells (CD138⁺) (b) in MRL/Mp and MRL/Mp.C1qa^−/− mice. Representative dotblots are shown to demonstrate how the above populations were gated during the analysis. The selected cells were defined by the regions R3 and R4, respectively. The data are expressed as mean ± SEM (*P < 0·1, **P < 0·05).

Figure 6

Plasma concentrations of total IgM in MRL/Mp and MRL/Mp.C1qa^−/− mice measured by ELISA as described in Materials and Methods. The data are expressed as mean ± SEM (*P < 0·1, **P < 0·05, ***P < 0·01).