Toll-Like Receptor 4 Stimulation Triggers Crescentic Glomerulonephritis by Multiple Mechanisms Including a Direct Effect on Renal Cells (1)
Toll-like receptors (TLRs) are a family of pattern recognition receptors that play a central role in the cellular response to microbes. TLR4 recognized lipopolysaccharide (LPS), otherwise known as Gram-negative endotoxin. The autologous phase of nephrotoxic nephritis is due to an immune response to the foreign glomerular bound antibody, which acts as a planted antigen. It is a more clinically relevant model of glomerulonephritis as the pathology shows proliferation, macrophage and T cell infiltration, and thrombosis and crescent formation, which are all features found in human disease.
Disease was induced by using protocol A in wild-type and TLR4-deficient mice. There were more glomerular macrophages, crescents, and thrombosis in wild-type mice compared with TLR4-deficient mice. There was also evidence of a functional effect as serum creatinine was higher in wild-type mice. There was no difference in albuminuria at day 7, and there was higher albuminuria in TLR4-deficient mice at day 14, which just reached statistical significance.
The peak response of the immune response to sheep IgG was seen at day 5, and there was an increase in all four IgG subclasses in wild-type compared with TLR4-deficient mice. The difference in IgG1 and IgG2c was maintained at later time points. To further assess any change in Th1 or Th2 skewing, we restimulated splenocytes with sheep IgG at the end of this experiment and measured interleukin-4 and interferon-γ in supernatants. Interleukin-4 and interferon-γ in supernatants of splenocytes were no significant differences.
In induced disease by using protocol B in wild-type C57BL/6 mice, no significant differences were seen between mice given LPS or control. Antigen-specific levels of all IgG subclasses were similar in both groups.
The distribution of staining did not follow the capillary wall as expected, and this was probably due to the large amount of thrombosis seen. Administration of LPS led to more glomerular crescents, increased glomerular thrombosis, greater numbers of CD68+ macrophages, but no difference in glomerular CD4+ cells. Albuminuria data were only obtained from five mice per group due to technical issues. There was no difference in albuminuria, which was severe in both groups. However, serum creatinine was significantly higher in mice given LPS compared with control.
In the setting of LPS administration after disease induction, the lack of renal cell TLR4 protects from crescent formation and thrombosis in the wild-type → TLR4−/− mice. There was significantly more thrombosis and crescent formation in the TLR4−/− → wild-type mice. There were no differences in albuminuria when either wild-type → wild-type and wild-type → TLR4−/− mice, or TLR4−/− → TLR4−/− and TLR4−/− → wild-type mice were compared. In contrast, differences in serum creatinine mirrored the changes seen in glomerular thrombosis and crescent formation demonstrating an important functional consequence of these morphological changes.
TLR2 stimulation augments the nephritogenic immune response without causing a change in the Th1 versus Th2 balance. There are previous reports of TLR stimulation of intrinsic renal cells causing kidney disease. A bone marrow chimera approach has defined a role for both TLR2 and TLR4 on renal cells in ischemia reperfusion injury, and also shown a role for renal cell TLR4 in a model of infection. In contrast, LPS-induced acute kidney injury in mice was not dependent on renal cell TLR4, as evidenced by studies with transplanted TLR4-deficient and wild-type kidneys. TLR3 has been found on mesangial cells and implicated in the pathogenesis of murine lupus, although in vivo evidence of a role was not determined through the use of bone-marrow chimeras.