General conditions
The normal saline (NS) treated MRL/lpr mice began to develop skin lesion at 16 weeks of age. The skin lesion was overt at 20 weeks of age (Figure 1). In contrast, MRL/lpr mice treated by 10mg/kg CRIg/FH twice weekly presented less hair loss.There were two mice died in normal saline treated MRL/lpr mice and one died in oral prednisone treated MRL/lpr mice before 20 weeks age..
CRIg/FH reduce proteinuria and protect renal function in MRL/lpr mice
The level of proteinuria in either NS treated or prednisone treated MRL/lpr mice were significantly elevated after 16 weeks of age (Figure 2A), whereas protein levels in urine of MRL/lpr mice treated with CRIg/FH were still not significantly elevated by 20 weeks of age. After 18 weeks of age, the level of proteinuria in all dosage groups of CRIg/FH treated MRL/lpr mice were significantly lower than that of the NS or GC treated mice (P<0.05, Figure 2A).
Treatment with CRIg/FH significantly reduced serum creatinine and urea nitrogen levels in MRL/lpr mice. At 16 weeks of age, serum creatinine levels in all CRIg/FH treated MRL/lpr mice were significantly lower than those in either NS or prednisone treated MRL/lpr mice (P<0.05, Figure 2B). At 20 weeks, serum creatinine levels in CRIg/FH treatment groups maintained to be significantly lower than that in NS treated MRL/lpr mice (P<0.05). However, comparing to prednisone treated MRL/lpr mice, significant decrease in serum creatinine level was only observed in CRIg/FH treatment with highest dose (10 mg/kg twice weekly, P<0.05, Figure 2B).
Similarly, serum urea nitrogen levels in MRL/lpr mice treated with CRIg/FH were all significantly lower than those in the NS treated MRL/lpr mice at 16 weeks of age (P<0.05, Figure 2B). In addition, serum urea nitrogen level in CRIg/FH treatment with higher dosage (10 and 5 mg/kg twice weekly) were significantly lower than that in the prednisone treated MRL/lpr mice (P<0.05, Figure 2B). At 20 weeks of age, the effect of CRIg/FH to maintain serum urea nitrogen at low levels in MRL/lpr mice was observed in higher dosage (10 and 5 mg/kg twice weekly) however not in the lowest dose (2 mg/kg) group (Figure 2B), .
CRIg/FH blocks complement activation in MRL/lpr mice
At 16 weeks, the levels of serum complement C3 and C4 in MRL/lpr mice treated with CRIg/FH at all dose was significantly higher than that in NS treated MRL/lpr mice (Figure 3). The difference in C3 was maintained at 20 weeks age mice only between MRL/lpr mice treated with highest dose CRIg/FH (10mg/kg twice weekly) and NS treated MRL/lpr mice (Figure 3). Nevertheless, the levels of serum C3 and C4 in CRIg/FH treated mice was significantly lower than that in C57BL healthy mice at both 16 and 20 weeks age (Figure 3). The serum levels of C4 was significantly lower in higher dose (10 and 5 mg/kg twice weekly) of CRIg/FH treated MRL/lpr mice when compare to either NS or prednisone treated MRL/lpr mice, at both week 16 and 20. The significance of C4 decreasing is not identified in lowest dose of CRIg/FH (2 mg/kg twice weekly) when compared to prednisone treated MRL/lpr mice (Figure 3).
Similarly, the activation of autoimmunity was significantly lower in MRL/lpr mice treated with CRIg/FH. At 16 weeks, the serum A-ds-DNA level in MRL/lpr mice treated with CRIg/FH at all doses was significantly lower than that in NS treated MRL/lpr mice or GC treated MRL/lpr mice (Figure 3). The effect continued at 20 weeks measurement (Figure 3). Nevertheless, the levels of serum A-ds-DNA was still significantly higher in CRIg/FH treated MRL/lpr mice than that in normal control mice at both 16 and 20 weeks (Figure 3)
CRIg/FH improves lupus nephropathy in MPL/lpr mice
The H&E staining of mice kidney at 20 weeks of age showed that MRL/lpr mice had significant nephropathy in those treated by NS than CRIg/FH (Figure 4A). In MRL/lpr mice treated with highest dose CRIg/FH (10 mg/kg twice weekly), there were 7 mice categorized as LN II, 1 mice LN III, whereas all mice treated with NS were scored as LN IV-G. The activity index in MRL/lpr mice treated with CRIg/FH were significantly lower than that in NS treated lupus mice (Table 1). The activity index of MRL/lpr mice treated with highest dose of CRIg/FH was significantly lower than that of prednisone treated MRL/lpr mice (P<0.05). Nevertheless, the nephropathy index of activity was not statistically different between lowest dose of CRIg/FH treatment (2mg/kg twice weekly) and prednisone treatment. The chronic index of MRL/lpr mice treated with higher dosage of CRIg/FH (10 and 5 mg/kg twice weekly) was the same as that in prednisone treated MRL/lpr mice.
The deposition of complement MAC, C1q, and C3d in all CRIg/FH treatment groups were significantly reduced compared with NS treated mice. Nevertheless, the deposition of immunoglobulin IgM and IgG were not significantly improved in CRIg/FH treatment groups. (Figure 4B)