Spontaneous recovery from early glomerular inflammation is associated with resistance to anti-GBM glomerulonephritis: tolerance and autoimmune tissue injury

Abstract

Different susceptibility to anti-GBM glomerulonephritis (GN) among animal strains has been reported. Using our rat model for T cell-mediated anti-GBM GN, this study initiated an investigation on the mechanism related with GN susceptibility. Anti-GBM GN was induced either through immunization with the nephritogenic T cell epitope pCol(28-40) from Col4alpha3NC1 or through the transfer of specific T cells. WKY rats were highly susceptible to GN while immuno-compatible LEW rats were GN-resistant. GN-resistance in LEW rats was not associated to the immune response to pCol(28-40). First, both strains mounted a Th1 T cell response to pCol(28-40) with identical specificities; transfer of T cells from LEW to WKY rats induced glomerular injury. Second, co-transfer of antibody from WKY to LEW failed to induce GN. Time-course studies revealed that LEW rats did develop T cell-mediated inflammation in glomeruli at early stages similar to WKY rats, as evidenced by histopathology, proteinuria, CD4(+) T cell infiltration in glomeruli, and glomerular expression of inflammatory molecules. However, glomerular inflammation in LEW rats was transient followed by a full recovery. Thus, GN-resistance in LEW rats was due to its ability to contain early T cell-mediated autoimmune glomerular damage. Our model may reveal a potential tolerance mechanism after autoimmune tissue damage has been initiated.

Figure 1. LEW and WKY rats are immuno-compatible

A. Mixed lymphocyte reactions between rat strains as indicated; LEW vs LEW serves as negative control, while LEW vs SD a positive control; the responsiveness is expressed as ³H-thymidin incorporation. B. Skin allograft rejection; days of rejection are counted from day of grafting until the graft is completely rejected; inset shows a LEW skin graft in WKY at 4 weeks.

Figure 2. LEW rats mount a pathogenic T cell response to pCol(28-40) similar to WKY rats

A. T cell response to pCol(28-40) in LEW (●) and WKY rats (○) after immunization; the responsiveness of T cells is expressed as ³H-thymidin incorporation. B. A T cell line from LEW or WKY rats responds to pCol(28-40) at the presence of thymocytes from WKY rats . C. A T cell line from LEW (■) responds to a set of pCol(28-40) with single substitution in almost identical pattern to those of WKY (□). D. RT-PCR detection of IL-4 or INF-γ in a T cell line from LEW rats after stimulation with pCol(28-40). E. Glomerular injury in pCol(28-40)-specific T cell recipients as indicated; severity of glomerular injury is expressed as glomerular injury score. F. Proteinuria in pCol(28-40)-specific T cell recipients as indicated; transfer of WKY's T cells to WKY recipient serves as a positive control; albumin is indicated by an arrow.

Figure 3. Antibody response in LEW rats immunized with pCol(28-40)

A. Circulating antibody activity to pCol(28-40) from immunized LEW or WKY rats; sera from CFA immunized WKY were used as a negative control; B. Reactivity of circulating antibody from immunized WKY or LEW to a set of pCol(28-40) with single substitution. Antibody activities are expressed as normalized OD. C. Indirect immunofluorescence shows absence of anti-GBM antibody in circulation of LEW and WKY rats; SR-13 serves as a positive control. D. Direct immunofluorescence shows absence of GBM-bound IgG in an immunized LEW rat.

Figure 4. Renal histopathology of LEW or WKY rats after immunization with pCol(28-40)

A. Kidney section of an immunized LEW rats at 40 days post immunization shows normal glomeruli. B. Kidney section from WKY shows glomeruli with fibrous crescent. C. A group of kidney sections under high magnification from the immunized rats as indicated; lymphocyte infiltration is notable in LEW at day 20 (arrows); crescent lesion (arrow heads) is shown in glomerulus of WKY at day 30.

Figure 5. Transient glomerular inflammation in LEW rats after GN induction with pCol(28-40)

A. Summary of renal pathology of different animal groups; severity of glomerulonephritis is expressed as glomerular injury score (0-100). B. Summary of albuminuria in the immunized rats at an early or later stage as indicated; albuminuria is expressed as mg/dl.

Figure 6. Flow cytometry detection of a transient CD4⁺ T cell-mediated glomerular inflammation in LEW rats after GN induction with pCol(28-40)

Leukocytes were isolated from glomeruli of WKY or LEW rats at indicated times and stained with anti-CD4 and anti-CD8 antibodies.

Figure 7. Glomeruli of LEW rats express inflammatory molecules similar to those of WKY rats at an early stage after GN-induction

Glomerular expression of inflammatory molecules as indicated. Only one representative array is shown. Arrows indicate the same set of genes, which includes Bmp2, caspase-1, IL-4Rα, MIP-2α, Frag1, TGF-β1i4, Il-1R1.