Background/Purpose

G protein-coupled receptors (GPCRs), including chemokine receptors on leukocytes, signal through G protein Gβγ subunits. An important target of Gβγ is phosphoinositide 3 kinase γ (PI3Kγ), a major isoform of PI3K whose receptor-dependent activation relies primarily on the Gβγ subunit. We hypothesized that Gβγ inhibition may be an effective treatment for lupus as it will prevent PI3Kγ activation which is upstream of T and B cell survival and the migration and activation of innate immune cells. Here, we evaluated the effects of gallein, a small molecule novel Gβγ inhibitory compound (Lehmann 2008, Mol Pharmacol 73: 410), in lupus prone mice.

Methods

18 week old NZB/NZW mice were treated daily with 20 mg/kg or 35 mg/kg gallein (intraperitoneal injection) or vehicle (n=8 per group) for 15 weeks. For therapeutic intervention, mice with established disease (26 weeks old) were treated for 4 weeks with gallein at 35 mg/kg.  Lymphocytes were enumerated by flow cytometry, auto-reactive antibody secreting cells (ASC) were quantitated by ELISpot, kidney inflammation was evaluated by histology (H&E, immunofluorescence), and renal function was assessed by monitoring changes in proteinuria. To determine the effect of gallein on cell migration, BM neutrophils or Jurkat T cells were stimulated in the presence of fMLP (250nM) or CXCL2 (30 or 100 ng/ml). 

Results

In a prevention model, gallein therapy significantly abrogated the progression of proteinuria (35 mg/kg: p=0.0001, 20mg/kg p=0.0116). There was a significant dose dependent reduction in the number of effector T cells (20 mg/kg: p=0.0495, 35 mg/kg: p=0.0001) and central memory T cells (35 mg/kg: p=0.0029) in spleens of lupus prone mice. Mice treated with a high dose of gallein (35 mg/kg) had a significant reduction in the number of T follicular helper cells (p=0.0092) and germinal center B cells (p=0.0411). Although auto-reactive IgG ASC and dsDNA specific ASC were not reduced in the spleen, mice treated with gallein (35 mg/kg) had a reduction in IgG⁺ and dsDNA⁺ ASC in the BM (p=0.0029 and 0.0571). Strikingly, both doses of gallein caused a marked reduction in the number of dsDNA⁺ ASC in the kidneys (20 mg/kg: p=0.0103, 35 mg/kg: p=0.0308). Treated mice also had a significant reduction in glomerular, interstitial inflammation (p=0.01 and 0.03 respectively), perivascular inflammation (35 mg/kg: p<0.0001), and IgG deposition. Treatment of mice with established disease also resulted in a significant reduction in the numbers of IgG⁺ (p=0.0173) and dsDNA⁺ASCs (p=0.0031) in the kidneys and reduced proteinuria (p=0.0236). Gallein inhibited the migration of WT murine neutrophils and cultured Jurkat T cells in response to fMLP and CXCL12, respectively. Similarly, in vivo administration of gallein in NZB/NZW mice resulted in a less efficient migratory response of BM isolated neutrophils in response to CXCL2. 

Conclusion

These results suggest that Gβγ inhibition with gallein may modulate the generation of PCs in GC reactions as well as alter the kidney inflammatory milieu and PC survival niche in lupus, possibly by influencing the influx and function of inflammatory cells.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/inhibition-of-g-protein-%ce%b2%ce%b3-signaling-inhibits-nephritis-in-lupus-prone-mice/