Our previous studies have shown that streptococcal IgG Fc receptors (FcR) act to elicit circulating anti‐IgG as well as renal glomerular deposition of IgG in rabbits immunized with group A streptococci (GAS). In order to study if other FcR‐positive bacteria might have similar effects, rabbits were immunized with either group G streptococci (GGS; strain G148) or Staphylococcus aureus (strain Cowan I) for two periods of 8 and 6 weeks, respectively. At the end of immunization, circulating anti‐IgG was found in 6 of 20 (30%) and 4 of 19 (21%) animals receiving G148 and Cowan I, respectively, compared to all 28 receiving FcR‐positive GAS strains of types M1, M4, M15 or M22 (p < 0.05 for both comparisons); furthermore, anti‐IgG appeared earlier and at higher levels in the GAS groups. Weak glomerular IgG deposits occurred in 5 out of 10 (50%) and 2 out of 8 (25%) animals immunized with G148 and Cowan I, respectively. In contrast, all 11 rabbits examined, given GAS of types M1 or M15, displayed heavy deposits. None of four control animals immunized with either of two FcR‐negative strains, GAS type T27 or group B streptococci (GBS) type la, exhibited any renal IgG deposits or circulating anti‐IgG. Renal tissue materials from rabbits immunized with any of the four FcR‐positive GAS strains showed strong inflammatory and degenerative glomerular changes, compatible with the picture seen in acute poststreptococcal glomerulonephritis (APSGN). Only transient renal changes were found in those rabbits immunized with G148 or Cowan I, or the controls injected with the FcR‐negative strains, GAS type T27 or GBS. Thus, only the FcR‐positive GAS strains showed capacity to induce high levels of anti‐IgG, pronounced tissue deposition of IgG as well as irreversible glomerular changes. Our experimental data suggest that streptococcal IgG FcR activity might play an important role in triggering APSGN.

Our previous studies have indicated that the IgG-binding M-family proteins (IgGBP) of group A streptococci may be involved in eliciting experimental acute poststreptococcal glomerulonephritis (APSGN) in the rabbit. These surface proteins were also found to trigger production of anti-IgG, which might conceivably act to enhance renal deposition of immune complexes (IC). In the present study, a clinical isolate of serotype M22 (strain AL168), an isogenic double mutant deficient for both the IgGBPs Mrp and Emm, as well as mutants deficient in only one of the proteins were tested for capacity to induce glomerulonephritis. Streptococci to be used for injecting rabbits were heat-killed. Surface-bound IgG was removed by 1 M KSCN and cells were then repeatedly washed in PBS before use. Rabbits were injected intravenously with 109 cells three times a week for 8 weeks and, following one month of rest, for another 6 weeks. Deposits of IgG and C3 as well as induced chemokines TNF-alpha, IL-1beta and IL-6 were traced in cryostat sections using specific antibodies and appropriate peroxidase-labelled anti-antibodies. In four rabbits immunized with the double mutant strain, no deposits were found, and as examined by TEM, only subtle and transient renal changes were observed. In contrast, the original strain AL168 induced pronounced inflammatory and degenerative glomerular changes in all four rabbits injected, and deposits of TNF-alpha, IL-1beta and IL-6 were found in mesangial and endothelial cells. Similar deposits and glomerular changes were seen in all eight rabbits injected with the mrp-emm+ mutant and in four out of seven animals receiving the mrp+emm- mutant. There was a highly significant correlation between high levels of circulating anti-IgG and development of APSGN. These results confirm an important role of streptococcal IgGBP in triggering experimental APSGN as earlier proposed by our group.

Burova, L. A., Koroleva, I. V., Ogurtzov, R. P., Murashov, S . V., Svensson, M-L. & Schalen, C. Role of streptococcal IgG Fc receptor in tissue deposition of IgG in rabbits immunized with Streptococcus pyogenes. APMIS 100: 567-574, 1992.Induction of anti-IgG during hyperimmunization of rabbit with Streptococcus pyogenes (group A streptococci; GAS) was previously shown to require the presence of IgG Fc receptors (FcR) in the vaccine strain. In the present work, we examined whether streptococcal FcR activity might also be of importance for heart and kidney deposition of IgG, known to occur in poststreptococcal sequelae as well as during experimental immunization of animals. Each of three IgG-binding (GAS types M1, M12 and M22) and two non-binding (GAS type T27 and S. agalactiae (GBS) type Ia) streptococcal strains were used for intravenous immunization of rabbits during two periods of eight and six weeks, respectively, separated by an interval of one month. Before use, vaccine strains were treated with KSCN and carefully washed in order to remove any surface-bound immunoglobulins. No deaths occurred among injected rabbits. No tissue deposition was elicited by the GAS type T27 or the GBS strain. In contrast, the strains of types M1, M12 and M22 all induced deposits of IgG in kidney and heart tissue, beginning during the first immunization period. In two tested animals, receiving GAS of types MI or M22, circulating immune complexes containing anti-IgG antibodies were also detected. Finally, serum autoantibodies reacting with preparations of heart and kidney, but not lung or liver, were demonstrated in each of six animals receiving M1 or M22, reaching maximum levels during reimmunization; such antibodies were not evoked by the two strains not binding IgG. Our results suggest that, in GAS with capacity for non-immune binding of IgG, triggering of anti-IgG acted to enhance tissue deposition of IgG or immune complexes in immunized rabbits. Furthermore tissuespecific antibodies were elicited only by the IgG-binding strains and occurred comparatively late during immunization, suggesting that those antibodies might have been triggered due to the exposition of hidden kidney and heart determinants.

The pathogenesis of acute poststreptococcal glomerulonephritis (APSGN), a major nonsuppurative complication of group A streptococcal (GAS) throat or skin disease, remains unclear. During the years, various theories based on certain streptococcal extracellular factors, as well as immunological mimicry between streptococci and renal tissue, have been forwarded. We earlier reported that many clinical GAS isolates with documented nephritogenic capacity show non-immune binding of monomeric or aggregated IgG. Moreover, in a rabbit model of APSGN we obtained evidence for an important role of streptococcal IgG Fc binding proteins (IgGFcBPs) belonging to the M family surface proteins; thus, hyperimmunization by whole IgGFcBP-positive streptococci was shown to induce renal glomerular changes with deposition of IgG and complement C3, resembling the picture recorded in human APSGN. These typical renal changes were always preceded by the appearance of circulating anti-IgG antibodies. In the present work, using the same rabbit model, each of two purified IgGFcBPs, isolated from type M22 GAS, were found to elicit glomerular degenerative damage comparable to that caused by whole bacteria, as well as formation of anti-IgG. In addition, the induction by whole streptococci (type M1) of experimental APSGN was inhibited by the i.v. administration of purified human or rabbit IgG Fc, but not Fab, fragment, supporting the importance of Fc-mediated mechanisms in causation of glomerulonephritis. We propose that anti-IgG antibody, induced by streptococcal IgGFcBP, facilitated renal accumulation of IgG-containing complexes, which in turn triggered complement deposition and proinflammatory cascades. Further studies on the possible beneficial effect of IgG Fc fragment in APSGN should be of interest.

No abstract

In a rabbit model, we have previously reported evidence for a pathogenic role of streptococcal IgG Fc-binding proteins (IgGFcBP) in poststreptococcal glomerulonephritis (PSGN). These proteins, of the M protein family, were shown to trigger anti-IgG production and enhance renal deposition of IgG and/or immune complexes (ICs), with resulting activation of complement and cytokine cascades. In the present study, type M12/emm12, group A streptococci (GAS) were found often to bind artificial ICs, viz. peroxidase-anti-peroxidase rabbit IgG (PAP) or tetanus toxoidanti-tetanus human IgG (TAT), rather than monomeric IgG. Animals injected with each of four IC binding clinical isolates (from patients with scarlet fever or PSGN) showed pronounced inflammatory and degenerative glomerular changes, morphologically similar to human PSGN, with membrane thickening and IgG and complement C3 deposition, as well as secretion of IL-6 and TNF-a by mesangial and endothelial cells. In contrast, non-binding strains (two from asymptomatic carriers and one from a PSGN case) failed to trigger any renal changes. Only the IC binding strains induced elevated titres of anti-IgG. Though the streptococcal binding component(s) has not been demonstrated, the selective binding of ICs by type M12/emm12 strains appears important for the well-known, marked nephritogenic potential of this GAS type.

Streptococcus pyogenes (group A Streptococcus; GAS) is an important gram-positive extracellular bacterial pathogen responsible for a number of suppurative infections. This micro-organism has developed complex virulence mechanisms to avoid the host's defenses. We have previously reported that SDSC from GAS type M22 causes endothelial-cell dysfunction, and inhibits cell adhesion, migration, metabolism, and proliferation in a dose-dependent manner, without affecting cell viability. This work aimed to isolate and characterize a component from GAS type M22 supernatant that suppresses the proliferation of endothelial cells (EA.hy926). In the process of isolating a protein possessing antiproliferative activity we identified arginine deiminase (AD). Further study showed that this enzyme is most active at pH 6.8. Calculating Km and Vmax gave the values of 0.67 mmol·L(-1) and 42 s(-1), respectively. A distinctive feature of AD purified from GAS type M22 is that its optimum activity and the maximal rate of the catalytic process is close to neutral pH by comparison with enzymes from other micro-organisms. AD from GAS type M22 suppressed the proliferative activity of endothelial cells in a dose-dependent mode. At the same time, in the presence of AD, the proportion of cells in G0/G1 phase increased. When l-Arg was added at increasing concentrations to the culture medium containing AD (3 μg·mL(-1)), the enzyme's capacity to inhibit cell proliferation became partially depressed. The proportion of cells in phases S/G2 increased concomitantly, although the cells did not fully recover their proliferation activity. This suggests that AD from GAS type M22 has potential for the suppression of excessive cell proliferation.

Heat‐killed group A streptococci of types M12 and M22 were used for intravenous immunization of 60 rabbits. Presence of Fc‐receptors binding monomeric IgG in M22 and their absence in M12 has been demonstrated previously. The strains were cultured in either Todd‐Hewitt broth supplemented with 10% horse serum (TH‐H) or in IsoVitaleX, a synthetic medium, supplemented with 10% autologous rabbit serum (IV‐R). A mortality rate of 38% was noted in the first‐mentioned group whereas no mortality was observed in the second group. Sera were examined for agglutination of red blood cells coated with either human or rabbit IgG. Antisera to strains cultured in TH‐H regularly displayed anti‐IgG antibodies reacting with human as well as rabbit IgG; anti‐rabbit IgG titres of anti‐M22 sera were significantly higher than those of anti‐M12 sera. Antisera to strains grown in IV‐R displayed high levels of antibodies reacting with human but not with rabbit IgG. The levels of anti‐human IgG did not differ between anti‐M12 and anti‐M22 sera.