Qihuang Jianpi Zishen Granules improves renal damage in MRL/lpr mice by inhibiting B cell differentiation via the AIM2/Blimp-1/Bcl-6 axis

doi:10.12122/j.issn.1673-4254.2025.11.02

Abstract

Abstract:

Objective To investigate the efficacy of Qihuang Jianpi Zishen Granules (QJZ) for inhibiting renal B cell differentiation in MRL/lpr mice and explore its underlying mechanism. Methods Thirty 8-week-old female MRL/lpr mice were randomly divided into model group, QJZ group, prednisone (Pred) group, QJZ+Pred group, and AIM2 inhibitor group (n=6), with 6 8-week-old female C57BL/6 mice as the normal control group. After treatments with normal saline, QJZ, Pred, or AIM2 inhibitor for 8 weeks, the mice were examined for urinary total protein-to-creatinine ratio (TPCR) and albumin-to-creatinine ratio (ACR), serum creatinine (Cr) and blood urea nitrogen (BUN) levels, and renal histopathology (with HE, Masson, and PAS staining) and ultrastructural changes (with electron microscopy). ELISA, immunohistochemistry, immunofluorescence staining and flow cytometry were used to detect blood levels of anti-dsDNA antibodies, cytokines and chemokines, renal deposition of complement components C3 and C4, renal expressions of AIM2, CD19, CD27 and CD138, and changes in splenic B lymphocyte subsets. The effect of QJZ on the AIM2/Blimp-1/Bcl-6 signaling axis was examined using Western blotting. Results QJZ treatment significantly improved Cr, BUN, TPCR and ACR in MRL/lpr mice, ameliorated renal pathologies, reduced the expressions of ds-DNA, BAFF, IL-21, CXCL12, CXCL13, C3 and C4, and increased IL-10 levels. QJZ significantly downregulated renal expressions of the key B-cell transcription factors Blimp-1 and XBP-1, upregulated Bcl-6 and PAX5 expressions, inhibited B-cell differentiation, and lowered the expressions of AIM2, CD27, CD138 and CD69. Inhibition of AIM2 similarly reduced renal Blimp-1 and XBP-1 expressions, increased Bcl-6 and PAX5 levels, suppressed B-cell differentiation, decreased IgG production, reduced C3 and C4 deposition, and alleviated renal pathology in MRL/lpr mice. Conclusion QJZ inhibits B cell differentiation and alleviates renal damage in systemic lupus erythematosus possibly by suppressing the AIM2/Blimp-1/Bcl-6 signaling pathway.

Key words: Qihuang Jianpi Zishen Granules, systemic lupus erythematosus, renal damage, B cells, AIM2/Blimp-1/Bcl-6

Lili CHENG, Zhongfu TANG, Ming LI, Junjie CHEN, Shuangshuang SHANG, Sidi LIU, Chuanbing HUANG. Qihuang Jianpi Zishen Granules improves renal damage in MRL/lpr mice by inhibiting B cell differentiation via the AIM2/Blimp-1/Bcl-6 axis[J]. Journal of Southern Medical University, 2025, 45(11): 2297-2308.

Figures/Tables 10

Fig.1 Comparison of renal function among different groups of MRL/lpr mice. A: Expression levels of ACR in MRL/lpr mice in each group. B: Expression levels of TPCR in each group. C: Expression levels of Scr in each group. D: Expression levels of BUN in each group. *P<0.05, **P<0.01.

Fig.2 Comparison of ds-DNA antibody, cytokine and chemokine levels in MRL/lpr mice in each group. A: Levels of ds-DNA in each group. B: Levels of IL-10 in each group. C: Levels of BAFF in each group. D: Levels of IL-21 in each group. E: Levels of CXCR-12 in each group. F: Levels of CXCR-19 in each group. **P<0.01.

Fig.3 Effect of QJZ on renal pathology in MRL/lpr lupus mice. A: Results of HE, PAS and Masson staining of the kidneys in each group (Scale bar=50 μm). B: Comparison of pathological AI scores of the kidneys in each group. C: Comparison of pathological CI scores of the kidneys in each group. *P<0.05, **P<0.01.

Fig.4 Comparison of ultrastructural changes in renal glomeruli in each group under transmission electron microscopy (Scale bar=2 μm).

Fig.5 Comparison of C3 and C4 deposition levels in the kidneys of MRL/lpr mice among the groups. A: Immunohistochemistry for detecting expressions of C3 and C4 in the kidneys in each group (×100). B: Average fluorescence intensity of C3 in the kidneys in each group. C: Average fluorescence intensity of C4 in each group. *P<0.05, **P<0.01.

Fig.6 Comparison of expression levels of key transcriptional proteins involved in B-cell differentiation in each group. A: Relative protein expression levels of Blimp-1 in each group. B: Relative protein expression levels of Bcl-6 in each group. C: Relative protein expression levels of XBP-1 in each group. D: Relative protein expression levels of PAX-5 in each group. E: Protein bands in Western blotting in each group. **P<0.01.

Fig.7 Spleen index of the spleen tissues in each group of MRL/lpr mice. *P<0.05, **P<0.01.

Fig.8 Percentages of B cells in each group of MRL/lpr mice.

Fig.9 Expression levels of CD19, CD138, IgG and AIM2 in the renal tissues of MRL/lpr mice in each group (Immunofluorescence staining; Scale bar=50 μm). A: CD19 expression levels in each group. B: CD138 expression levels in each group. C: IgG expression levels in each group. D: AIM2 expression levels in each group.

Fig.10 Effect of AIM2 inhibitor on renal damage in MRL/lpr mice. A: Relative protein expression levels of Blimp-1 in each group. B: Relative protein expression levels of Bcl-6 in each group. C: Relative protein expression levels of PAX-5 in each group. D: Relative protein expression levels of XBP-1 in each group. E: Western blotting bands in each group. F: Results of HE, Masson, and PAS staining in each group (×200). G: Expression of IgG in each group (×200). H: Level of B cells in each group. I: Expression of C3 and C4 in each group (×200). **P<0.01.

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