The pathogenesis of autoimmunity in New Zealand mice.

The Pathogenesis BySteven INDEX WORDS: Autoimmunity; systemic lupus erythematosus.

of Autoimmunity Yoshida,

New Zealand

James J. Castles, and M. Eric Gershwin mice;

T

HE CENTRAL THEME underlying the structure and function of the immune system is self/nonself discrimination. This function was a crucial factor in the evolution of intercellular cooperation and the development of multicellular and colonial life forms. Indeed, extant marine invertebrates have recognition systems reminiscent of those of the distant past. In the mammalian immune response this property has reached its greatest complexity. Through a network of interrelated pathways, the immune system responds to the intrusion of living or nonliving foreign matter with an appropriate response directed toward elimination from the host animal. The capacity to generate and maintain a record of the nature of the invading entity also results in faster responses to subsequent contacts. A complex network needs numerous sites of checks and balances to function properly; inherently, this presents many opportunities for imbalances in the system. Few conditions demonstrate this as clearly as systemic lupus erythematosus (SLE), in which the immune system loses its ability to distinguish foreign material from self. In this regard, murine models of autoimmune disease has been helpful in the understanding of this illness.’ Although significant volumes of data describing the many humoral and cellular anomalies of these mice have been generated in the past 3 decades, the primary cause of disease has yet to be determined. The problems of clarifying the etiology of their disorders have been great

From the Division of Rheumatology, Allergy and Clinical Immunology, Department of Internal Medicine, School of Medicine, University of California, Davis. Steven Yoshida, PhD; Postgraduate Research Immunologist. Division of Rheumatology; James J. Castles, MD: Professor of Medicine, Division of Rheumatology; M. Eric Gershwin, MD: Professor of Medicine. Chief. Division of Rheumatology. Supported by NIH grant CA 20816. Address reprint requests to M. Eric Gershwin, MD, University of California, Division of Rheumatology, Allergy and Clinical Immunology, TB 192, Davis, CA 95616. D 1990 by W.B. Saunders Company. 0049-0172/90/l 904-0002%5.00/O 224

in New Zealand Mice

because the study of autoimmunity is closely wedded, philosophically, to that of basic immunity. A major emphasis of both disciplines is to develop an understanding of the principles of self/nonself discrimination. This review of New Zealand mice focuses primarily on recent reports that have most influenced present concepts on disease and provided the most promising directions for future research. GENERAL

LIFE HISTORY

The average life spans for male and female New Zealand black (NZB) mice are 467 and 43 1 days, respectively (Table 1). Their truncated survival times are attributable to an autoimmune hemolytic anemia which appears between 1 and 5 months of age. This anemia is caused by anti-erythrocyte antibodies that first become detectable at 3 months of age and reaches a frequency of 100% by about 1 year. Natural thymocytotoxic autoantibodies (NTA) which bind to the T-lymphocytes of mice have also been detected in sera of NZB mice. The appearance of NTA precedes that of anti-red blood cell (RBC) antibodies and reaches 100% incidence by 3 months of age. The crossings of NZB with New Zealand White (NZW) or SWR mice, hereafter designated the B/W F, and SNFl hybrid mice, result in animals which develop a spontaneous disease similar to human SLE. These hybrid mice usually die of glomerulonephritis, following the deposition of immune complexes. At approximately 5 months of age, deposits of immunoglobulin G (IgG), particularly IgG2a, and C3 are detectable in significant amounts in the glomerular mesangium of B/W F, mice. Immunoglobulins eluted from B/W F, kidneys include those specific for single-stranded (ss) and double-stranded (ds) DNA. The mean life spans of male and female B/W F, mice are 439 and 280 days, respectively. Although there is some overlap in the antigenic specificities of B/W F, and NZB autoantibodies, B/W F, mice are noted mostly for their antibody activity to nuclear antigens. Antinuclear autoantibodies (ANA) are first detectable at 6 months of age in females and reach 100% incidence by 9. Male mice have a some-

Seminars in Arthritisand Rheumatism, Vol 19, No 4 (February), 1990: pp 224-242

MURINE LUPUS

225

Table 1. General Characteristics

of Autoimmune

New Zealand Mice Parameter

SPA’F,

NZB

Life span Male

467.

439

Female

431

260

Prominent

Natural

autoantibodies

thymocytotoxic

Anti-ss, -dsDNA

Anti-erythrocyte

Anti-histone

Anti-ssDNA

Anti-RNA

Anti-gp70

Anti-gp70

Yes

Yes

Yes

Yes

Slightly female

Strongly female

Hemolytic anemia

Renal failure

Accelerated thymic degeneration Tissue infiltration by lymphoid cells Sex predominance Primary cause of death

*Mean life span in days.

what slower disease progression; ANA appear at 7 months and become universally prevalent by approximately 1 year. Of great interest in both NZB and B/W F, mice is the accelerated degeneration of the thymus. Indeed, the abnormal atrophy of thymic cortices in several strains of lupus mice and the premature deterioration of NZB thymic epithelial cells have been described.2’ IMMUNOGLOBULINS:

AUTOANTIBODIES

AND IDIOTYPY

There are four known anti-erythrocyte responses in NZB mice. Three of these antigens are cryptic and can be revealed by RBC treatment with bromelin. Although the naturally exposed antigen (antigen X) is the major target of hemolytic autoantibodies, the nature of this X autoantigen has not yet been characterized. The anti-erythrocyte response may be directed toward membrane phosphatidyl choline.4 Recently, a monoclonal antibody derived from NZB mice specifically bound to N-glycolylneuraminic acid-containing GM2 ganglioside.’ This ganglioside is commonly found in mouse liver and erythrocytes, but is also located in some human tumor tissues. Another report also described an NZB monoclonal antibody with activity similar to pathogenic anti-X antibodies6 Further studies on the characteristics of such monoclonal autoantibodies may lead to the identification of erythrocyte antigens and perhaps clues to the initiating signals of autoimmunity.

The other prominent autoantibody of NZB mice is NTA. These IgM immunoglobulins react with virtually all T cells in a pattern similar to anti-theta antibodies. Although the identity of the target antigen(s) is controversial, NTA activity may lean toward immature and suppressor T cells.‘,s Also, in this context, a recent study of sera from NZB mice used immunoblotting to identify two noncovalently linked molecules unique to the surfaces of lymphoid cells.g Because the heterogeneity of the NTA response increases the difficulty in characterizing the T cell antigens involved, monoclonal NTA will be extremely helpful in this regard. The development of two NZB-derived monoclonal antibodies with cytotoxic activity to T cells has been reported. One showed antigenic cross-reactivity between thymic and gut tissue. The antigen was characterized as an 88,000 M, glycoprotein, unrelated to Ly-I, Lyt-2, or L3T4, and considered a possible novel T cell differentiation marker.“*” The other had a widespread binding pattern more reflective of NTA and NZB sera.12 Pathogenic anti-DNA autoantibodies of B/W F, and SNFl mice possess characteristics that are distinct from other anti-DNA populations. They are especially reactive to dsDNA; NZB mice have circulating antibodies to ssDNA, yet do not suffer from severe glomerulonephritis. The affinity of binding to DNA may also be a factor of the severity of disease. Anti-DNA antibodies are known to cross-react with other molecules such as cardiolipin which share a similar sugar-phosphate backbone structure. However, Smeenk et alI3 report little cardiolipin cross-reactivity of monoclonal antibodies with high affinity for dsDNA. Antibodies with high affinity for DNA were fairly specific to SLE, whereas low-affinity antibodies to DNA were not.i4*i5 Therefore, although low-affinity antibodies to DNA may be indicative of SLE, they are probably not responsible for immune complex deposition in this disease. Renal disease in B/W F, mice is also associated with a pronounced age-dependent isotype switch among the anti-dsDNA antibodies from IgM to IgG, particularly IgG2a and IgG2b. The pathogenic nature of IgG2a may be due to its relatively long half-life and its ability to activate complement. The details of this IgM-to-IgG2 switching are not known; however, exposure to

226

various T-dependent and T-independent antigens resulted in a preference of isotype switching to IgG2 in several lupus mouse models, including of IgG2a the B/W F,. l6 The predominance among antichromatin autoantibodies in SLE mice was also recently observed.” Viral infections may stimulate the production of IgG2a preferentially’s perhaps through the action of interferon gamma (IFN-y).19 Finally, studies on SNFl mice have shown that nephritogenic IgG antibodies to DNA are cationic in charge.20 This characteristic appears to enhance the deposition of immune complexes,21 perhaps because of the presence of anionic sites in the glomeruli of the kidney.22 Within the context of NZB autoimmunity, the interrelationships between the immunoglobulin variable-region genes, the idiotopic characteristics of antibodies, and their binding affinities to autoantigens should be discussed. Idiotopic similarities among anti-DNA antibodies in B/W F, and SNFl mice have been reported20*23; however, idiotopes were shared among antibodies to similar, but not identical, antigens such as DNA and poly (adenosine diphosphate-ribose).24 Common idiotopic structures have also been seen among autoantibodies from SLE and normal mouse strains with even more divergent antigenic specificities. 25*26Finally, a large idiotypic diversity has been shown among B/W F,-derived monoclonal antibodies specific for a similar RNA antigenic determinant. 27These latter results show the limitations of correlations that can be drawn between idiotypy and antigen specificity. Of interest are the observations that some monoclonal murine autoantibodies and conventional antibodies have shown binding to more 26,28 The pattern of antigenic than one antigen. reactivity and repertoire development of spleen cells from young NZB mice is also similar to those of normal mice,29 suggesting that autoreactivity is a normal function of the ontogeny of the immune system. But in animals genetically predisposed to autoimmunity, the structural and functional status of autoantibodies may change with age. The affinity of antibodies to selfantigens may increase, with corresponding decreases in both reactivity to multiple antigens3’ and idiotypic diversity,31 as the autoimmune response matures. Tsubata et a132 present data suggesting that B-cell precursors for high-affinity

YOSHIDA, CASTLES, AND GERSHWIN

anti-DNA antibodies in B/W F, animals are not eliminated as efficiently during maturation as in normal mice; a similar defect in clonal selection has been reported for NZB mice.33 These observations are like the earlier discussion on the nonpathogenic nature of autoantibodies with low affinity for antigen and suggest that they have some role in the ontogeny of pathological autoimmunity and perhaps normal immunity.34 Finally, whether pathogenic autoantibodies are selected from a limited number of immunoglobulin gene families or from a pool of many different gene families is debatable.35,36 Idiotypic restriction in the autoantibody response has been used to support the first position. However, without a clearer picture of the early stages of SLE and the changes that occur during its progression, especially when it concerns the immunogens that initiate the ANA response and the antigens that maintain it, labeling any particular family of Vu genes as the primary source of pathogenic autoantibodies will be difficult. Also, evidence for somatic mutation in the generation of autoantibodies3’g3* implies that selection pressures can converge antigenic specificities among antibodies from diverse genetic origins. B LYMPHOCYTES

Although the B lymphocyte is one of the most well studied of all cell types, its role in autoimmunity, other than as the direct source of autoantibodies, is still unclear. Nevertheless, one characterization of NZB B cells is a tendency toward hyperactivity (Table 2). B-lineage cells from NZB mice exhibited a precocious developmental process, that led to unnaturally high quantities of mature B cells and the depletion of B-cell precursors at early ages. 39 This production of mature B cells was also reflected by the increased numbers of spontaneous and mitogen-induced B-cell colonies obtained from NZB spleens‘te741 as well as elevated, uninduced. in vitro antibody production by spleen cells from NZB and B/W F, mice.42343 The accelerated establishment of IgM’ NZB B cells in long-term bone marrow cell cultures also demonstrated their rapid developmental activity.44 The ability to induce unresponsiveness to specific antigens is also a functional measure of cellular activity. Both NZB and B/W F, B cells have been shown not to be rendered tolerant to low-epitope-density trinitrophenyl (TNP) conju-

MURINE LUPUS

Table 2. Lymphocyte

227

Abnormaliiiea

in New Zealand Mice

B cells Spontaneous in vitro Ig production Hypergammaglobulinemia Increased numbers of colony-forming units-B cells, clonable B cells Early depletion of bone marrow B cell precursors Elevated numbers of Ly- 7 + B cells Increased responsiveness to antigens, mitogens, T-cell signals Defects in tolerance induction Intracellular accumulation of J chains and C terminal frsgments of p chains Increased c-myc oncogene expression Decreased ability to generate T-suppressor cells T cells Decreased suppressor activity Decreased autologous mixed lymphocyte reaction Defects in tolerance induction Enhancement of autoantibody production following transplantation into normal hosts

gates4’ Also, a comparison of B/W F, and BALB/c cell lines disclosed an almost 50-fold higher dose requirement of dinitrophenyl-mouse gamma globulin (DNP-MGG) to induce unresponsiveness in B/W F, cells.& Although the reasons for such T cell-independent nontolerance are not known, it has been suggested that the density of cell surface receptors for antigen or their binding affinities are lower than those on normal cells or that intracellular signal transduction may be defective in the B cells of autoimmune animals. Additional evidence for B-cell hyperactivity includes increases in responsiveness to accessory cell signals.47-49In addition, the overexpression of the c-myc oncogene” and the cytoplasmic accumulation of two proteins, possibly J chains and C terminal fragments of chains,51 are also characteristic of activated NZB B cells. Reports on the association of Lyt-1 (a cell surface protein once thought unique to T lymphocytes) with murine B-cell lymphomas, and the homologous Leu- 1a with human chronic lymphocytic leukemia, led to the discovery of such an expanded population of splenic B cells in NZB mice.53 This has spurred ongoing interest in their possible contributions to autoimmunity. Many morphological and functional characteristics distinguished the Ly-I + from the Ly-1 - B cell. The Ly-I + cells expressed this molecule at a lower density than T cells, and although they are not known to carry other T-cell markers, they

had an increased surface IgM/IgD ratio and present Mac- 1.54Recently, evidence for a possible “sister population” of Ly-I -, Mac- 1’ B cells has been found in an examination of immunodeficient RIIIS/J mice.55 In normal mice, Ly-I + B cells constituted a minor (1% to 2%) portion of splenic and peripheral IgM+ B cells.56 The largest population of Ly-I+ B cells was found in the peritoneal cavity, where 50% of the surface (s) IgM+ B cells of mice 2 to 3 weeks of age were Ly-I +; the frequency decreases to 30% in adults. Also, approximately 20% of peritoneal Ly-l+ B cells expressed X light chains.” Athymic nude mice had normal levels of Ly-1 + B cells, whereas immunodeficient mice were normal for splenic Ly-I+ B cells but had none present in their peritoneums.56‘58 The activated status of these cells has been noted by their elevated levels of c-myc transcripts and the ease of producing immortal, but nontumorigenic, Ly-1 a B cell lines.59 The induction of peritoneal cells into the S phase by phorbol esters in the absence of comitogen6’ and nuclear immunofluorescent staining experiment$l has also demonstrated the high activation state of Ly-l+ B cells. Another characteristic of Ly-l+ B cells is their ability to regulate lymphoid cell functions. For example, their reported helper activity toward Ly-l- B cells may be due to the secretion of a nonimmunoglobulin lymphokine.62 Ly-1 + B cells have also been shown to compose virtually all of the B cells present in the shortest-lived animal model of autoimmunity, the motheaten mouse.63 A B-cell maturation lymphokine, of B-cell origin, has also been described64 and the investigators hypothesized that Ly-1 + B cells may have developed the capacity for idiotype-specific helper activity directed toward B lymphocytes. The possible involvement of Ly-l+ B cells in T cellmediated immune suppression has also been reported. 65 Aged NZB mice also possess “hyperdiploid” Ly-l+ B cells that are characterized by additional chromosomes 10, 15, 17, and X and increased surface expression of Ia. The transfusion of NZB hyperdiploid cells into (NZB x DBA/2)F, recipients resulted in the expansion of donor cell populations in recipient spleens, a decrease in endogenous peritoneal Ly-l+ B cells and splenic B cells, an increase in splenic macrophages, and reduced levels of se-

228

rum IgM and anti-ssDNA autoantibodies. There was no evidence of donor cells in recipient peritoneal cavities. Based on these observations, an immunoregulatory role for the hyperdiploid Ly-1 + B cell has been suggested.@j The regulatory activities of Ly-I+ B cells are also made more intriguing by finding Ly-I + B cells in the thymus of normal mice.67 How such cells may migrate to the thymus is not known; however, the presence of unconventional surface antigens on Ly-I+ B cells suggests the possibility of homing to tissues different from those of Ly-IB cell. Interestingly, the peritoneal cavity is thought to drain into the parathymic lymph nodes6* forming a possible route of travel from the peritoneum to the thymus. The developmental stages of the Ly-I+ B lineage have also been under scrutiny. Reconstitution experiments suggest that Ly-l+ and Ly-1 B-cell progenitors are distinct cell types.69 Also, by the coculturing of Ly-I+ B-cell progenitor clones with microenvironmental elements, bone marrow stromal cells have been shown to be capable of inducing Ly-IB lymphocytes and myeloid cells, whereas fetal liver stroma generated both Ly-I+ and Ly-I- B cells.70971 Finally, the recruitment of Ly-I+ B cells from their precursors appeared subject to feedback regulation by mature Ly-I+ B cells.72 Work on the structural features of immunoglobulin from Ly-l+ B cells shows some evidence for a restriction of variable region genes. Two recent studies have shown the exclusive use of a novel V, gene family, V,l 1, by Ly-l+ B cells.73Y74 An analysis of Ly-li B cell lymphomas also showed antigen-selected restriction of V, and V, gene usage; these genes appeared not to have undergone somatic mutation.75 These Ly-I+ B cell lymphomas also demonstrated the ability for isotype switching while maintaining specificity to phosphatidyl choline. Interestingly, some clones possessing DNA contents approaching tetraploidy were able to secrete and express on their surfaces more than one immunoglobulin isotype.76 Ly-l+ B-cell numbers are above normal in NZB mice in both the peritoneum and the spleen (Table 3); hybrid B/W Fl splenic levels are even higher than those for NZB animals. In aged NZB and B/W F, individuals, elevated numbers of Ly-I+ B cells appear in the circulation.56*57*77 In culture, the Ly-I+ B cell has been shown to be


Table 3. Properties

of LY-LC B Cells in New Zealand Mice

Expanded numbers in spleen and peritoneal cavity: higher in B/W F, mice than NZB. Present in the circulation of aged NZ mice. Responsible for the spontaneous release of antibodies in vitro Antibodies tend to be directed toward self-antigens: RBC, thymocytes, DNA. Ly- 7 + B cells tend to be stimulated by self, and not exogenous, antigens. Ly- I+ B-cell functions appear regulated by T cells. Subpopulation in aged NZB mice may be hyperdiploid and have regulatory activities.

responsible for the spontaneous secretion of 1gM characteristic of NZB spleen cells,56,78 and the specificities of these antibodies were directed toward self, including thymocytes, and SSDNA.‘~,~* The in vitro exposure of NZB spleen cells to exogenous antigens resulted in the stimulation of Ly-l- and not Ly-l+ B cells,56978again suggesting that the repertoire of Ly-lf B-cell surface immunoglobulins is directed toward selfantigens. However, it should be noted that antissDNA antibody production may also reside in the Ly-I- spleen cell population.79 A segregation of autoantibody specificities may be discerned among Ly-l+ B cells. Unlike NZB spleen cells, peritoneal Ly-I+ B cells tended to produce antibodies to bromelin-treated mouse RBC (BrRBC).57 Also, splenic Ly-1.1 B cells from B/W F, mice were not associated with the production of anti-dsDNA autoantibodies. This compartmentalization of antibody specificities was also noticed with hybridomas derived from NZB and BALB/c mice” in which recognition of BrRBC were primarily because of peritoneal cells, and responses to DNA and cytoskeletal proteins were found among spleen-derived hybridomas. The antibodies to RBC were relatively monospecific, in contrast to the polyspecific antiDNA/cytoskeleton antibodies, the latter being suggestive of the natural, low affinity form discussed previously. A study of xid (X-linked immunodeficiency)-congenic lines of motheaten mice showed a reduced number of Ly- 1 + B cells accompanied by decreased serum IgM, antissDNA and anti-BrRBC autoantibody levels; interestingly, NTA titers remained elevated.*’ Despite the evidence for hyperactivity and spontaneous in vitro immunoglobulin production by NZB Ly-1 + B cells, these characteristics may not be inherent to these cells, but possibly under regulatory control. The maintenance of anti-

229

MURINE LUPUS

erythrocyte antibody production by the peritoneal B cells of normal C3H mice was shown to require the presence of T-cell or macrophage factors.82 Also, the ability to readily clone autoantibody-producing hybridomas from lipopolysaccharide (LPS)-stimulated BALB/c cells suggests that precursors of autoreactive B cells were present but simply not activated.” Studies on an Ly-1 + B-cell clone have also shown a T-cell requirement for antigen-induced differentiation to antibody production; interestingly, the T-cell signals were proved replaceable by monoclonal antibodies to I-E but not I-A.83 The significance of the relationship between the Ly-I+ B cell and autoimmunity is also reflected by recent studies on the human homolog, Leu-l B cells. These cells synthesized rheumatoid factor and anti-ssDNA autoantibodiess4 and composed approximately 50% of the B cells in human fetal spleen but were rare in adult spleen,85 expressed Mac-l and p150,95 surface antigens,86 and constituted a major portion of the B lymphocytes following bone marrow transplantation.87 EVIDENCE FOR MICROENVIRONMENTAL INFLUENCES IN AUTOANTIBODY

PRODUCTION

Numerous studies on autoimmune strains of mice have demonstrated abnormalities in virtually all leukocyte lineages.88 These observations suggest that problems among non-B cells capable of shaping B-cell activity were involved in generating the pathological response to self. Several studies have been performed to estimate the impact of a nonautoimmune environment on NZB B cells. First, the transfer of purified, NZB splenic B cells into NZB.xid/xid mice increased the number of anti-ssDNAproducing splenic fragments in the recipients, thereby showing that mature, autoreactive NZB B cells alone could transfer autoimmunity to an NZB environment deficient in host B cells.8g A succeeding study then reported that following the transplantation of NZB or DBA/2 splenic non-T cells into NZB.xid and DBA.xid hosts, the rate of B-cell expansion correlated more closely with the host environment than the source of the donor cells; both donor cell types proliferated more readily in NZB.xid animals.gO*glThe investigators therefore conclude that the ,B-cell environment is capable of influencing the autoim-

mune response. It should be emphasized that these experiments transferred splenic B cells from autoimmune mice. The presumed mature and activated status of these cells implies that these recipient animals were models for the maintenance, but not the cause, of autoimmunity. Also, although donor cells were probably responsible for the increase in the anti-ssDNA IgM response, the investigators acknowledge the possible induction of endogenous B cells by the donor NZB cells. Another host that may be used to alleviate this problem is the severe combined immunodeficiency (SCID) mouse.g2 These animals are genetically unable to generate mature lymphocytes but are thought to have a normal microenvironment and reconstitution capability. Data on soluble effector molecules also suggest that a network of environmental factors are involved in guiding the autoimmune response in NZB mice. Humoral factors that enhanced the maturation of immediate B-cell precursors were found in the serum of young NZB mice.g3 Amniotic fluid from NZB mice was also shown to contain high interleukin-1 (IL-l) activity upon separation from the inhibitory alphafetoprotein; colony-stimulating activity-l was reduced instead.g4 In B/W F, mice, IFN-y aggravated their autoimmune response whereas antibodies to IFN-yg5 and replacement therapy with recombinant tumor necrosis factor delayed disease development. g6 Elevations in serum acute-phase proteins were also associated with the onset and severity of disease in NZB and B/W F, mice.” Ahmed et a1,g8 recently reported that estrogen increased the ratio of Ly-I + B cells committed to autoantibody production, perhaps through a direct hormonal effect on B cells. Nutrition is also. an influential factor in autoimmunity.” Studies have reported the beneficial effects of fish-derived lipids on inflammatory .processes. The basis for these responses is probably that fish oils (eicosapentaenoic acid) are poorer substrates for the production of arachidonic acid than vegetable oils (linoleic acid) and have a greater tendency toward the formation of immunoinhibitory types of leukotrienes and prostaglandins. Changes in dietary lipid might have altered the membrane characteristics of lymphocytes, such as fluidity. Dietary zinc deprivation has also been shown to lessen the severity of disease in NZB and B/W F, mice.iee Although


230

the mechanism of zinc-deprivation effects is not known, zinc-finger formation of DNA-binding proteins”’ or protein kinase Cl’* may be involved. Experimental supplementation of low selenium doses in rodents increased immune responsiveness. lo3 The mode of action appeared to be as a component of glutathione peroxidase. The enhancement of antioxidant activity may have reduced the damaging effects of cellular oxidizing agents, such as those produced during the respiratory burst of phagocytic leukocytes. In addition, action on the hydroperoxides of unsaturated fatty acid results in sparing of unsaturated fatty acids with a beneficial influence on autoimmune conditions. lo4Finally, the feeding of caseinfree diets to B/W F, animals led to a decrease of the isotype switching of anti-dsDNA antibodies and glomerular deposition of immune complexes.lo5 In the context of nutrition, the contributions of gut-associated immunity to the diseases of NZ mice may be important. Although this subject has not been vigorously pursued, some information is available. Peyer’s patch cells from NZB mice did not spontaneously secrete immunoglobulins in vitro, as did NZB splenocytes, and their IgA to IgG ratio was much lower than those from DBA/2 rnice.lM The mesenteric lymph nodes of B/W F, mice also had subnormal lymphocyte numbers and functions.“’ T-CELL DEPENDENCY OF THE AUTOANTIBODY RESPONSE

Indirect evidence showed that the autoimmune responses in NZB and B/W F, mice were related to T-helper activity (Table 4). These included demonstrations of the dependency of NZB and B/W F, B cells on helper signals for proliferation Table 4. Evidence for the T-Cell Dependency Pathogenic

Autoantibody

of

Production in New Zealand Mice

Treatment of mice with anti-CD4 antibodies reduced disease severity in B/W F, mice. ANA similarities in NZ mice and GVH disease were noted. NZB T cell transplantation into DBA/Z hosts resulted in increased autoantibody levels. T cells required for in vitro anti-dsDNA autoantibody production, and IgM-to-IgG switch. T cells required for in vitro synthesis of cationic autoantibodies by B/W F, and SNFl cells. Studies on NZ T-cell clones confirmed T-cell involvement in autoantibody production.

and immunoglobulin production,47*10* the improved condition of B/W F, mice following treatment with anti-CD4 antibodies,“’ the heightened numbers of terminal deoxynucleotidyl transferase-positive cells in B/W F, thymuses,“’ and the appearance of antibodies to nuclear antigens with chronic graft-versus-host (GVH) disease.“‘*“* More directly, enhanced autoantibody production when NZB T cells were cotransplanted with DBA/2 B cells into DBA/xid hosts113 and the NZB T-cell dependency of in vitro autoantibody secretion by NZB and DBA/2 B cells have been shown.114*115 Sekigawa et al’uj also describe an age-related, isotype-specific regulatory mechanism for this response that was dependent on specific subsets of CD4+ and CD8+ T cells. Splenic L3T4+ T cells from 2-month-old B/W F, mice tended to upregulate IgM production, whereas L3T4+ T cells from 7-month-old mice augmented IgG production. Although B cells capable of producing anti-DNA IgM were recovered from young B/W Fl mice, IgG producers were found only in the older animals. Datta et al”’ also document the need for both T and B lymphocytes to be derived from aged B/W F, mice in order to induce anti-dsDNA IgG synthesis. They report that T cells enriched for both CD4+ CD8- and CD4- CD8- populations were able to provide the necessary differentiation signals to produce cationic anti-dsDNA IgG. The charged nature of these antibodies and their specificity for DNA were thought to be separate phenomena because cationic antibodies evident in the cultures of spleen cells of 1-year-old NZB mice were not specific for DNA, and anti-DNA IgG antibodies induced by LPS treatment were not cationic. Studies with autoreactive T-cell clones were also used to clarify the T dependency of autoantibody production. Naiki et al”’ describe a T-cell clone derived from a B/W F, mouse that enhanced the in vitro production of IgM and IgG anti-ssDNA antibodies by young and old B/W F, splenic B cells, respectively. Additionally, adoptive transfer of this cell clone into aged B/W F, animals increased their serum levels of IgM, IgG2a, and IgG2b antibodies specific for ssDNA. The frequency of B/W F, autoreactive T cells capable of releasing IL-2 also increased with age.“’ These T-helper clones may be able to

MURINE LUPUS

affect B-cell differentiation through antigenspecific (antigen plus Ia requirement), as well as antigen-nonspecific (Ia only), pathways.12’ An NZB T-cell clone has also been studied and its ability to enhance B-cell proliferation and differentiation described.i2’ A report by Sainis and Datta12’ describes culture characteristics for SNFl cells similar to those found for B/W F, mice. Like NZB splenocytes, cultured SWR cells did not result in the release of cationic, antiDNA IgG. Importantly, I-A restriction was involved in these studies of T-cell clones. SUPPRESSOR NETWORKS

T-cell help in the induction of autoantibody production may be complemented by an agedependent loss of suppressor mechanisms. Numerous studies involving the coculture of various lymphocyte populations have been performed to analyze how autoantibody production is regulated down. The in vitro response to mouse RBC by splenocytes from Coomb’s positive NZB mice was reported to be suppressed when cocultured with splenocytes from young, but not old syngeneic animals.lz3 This suppression was specific for mouse RBC and the cells responsible for this suppression carried the Lyt-2 marker.1143124Furthermore, age-related, class-specific suppression moderated anti-DNA antibody production by B/W F, splenocytes; Lyt-2+ cells from young mice tended to inhibit anti-DNA IgG production, whereas cells from aged mice suppressed IgM production.116 Lyt-2+ suppressor T cells were also responsible for reducing the in vitro synthesis of autoantibodies by splenocytes from young BALB/c and C57B1/6 mice,12’ thereby confirming the presence of autoreactive B-cell clones in all mouse strains but reinforcing the notion that the expression of autoimmunity is partially controlled by suppressor T cells. Also, based on such experimentals, Moore and Calkins124 concluded that because of the ability of young cells to suppress the autoreactive tendencies of old cells, “hyperactive” B cells were probably responsive to T-cell-mediated suppression. The ability of helper or contrasuppressor T cells to mask the effects of suppressor T cells in aged NZB mice has been hypothesized.‘23Y’24The in vitro enhancement of autoantibodies by a population of Lyt-2- T cells may have been

231

caused by contrasuppressor T cells.114Laskin et alli’ submit that NZB contrasuppressor cells may have been responsible for the incapability of unfractionated DBA/2 spleen cells to suppress autoantibody production during in vitro coculture. The inability of young NZB thymocytes to inhibit autoimmune hemolytic anemia in loweek-old syngeneic animals126 may have been caused by the establishment of host contrasuppressor activity. Suppression by the idiotype network might be defective in NZB mice. Cohen and Eisenberg12’ reported anti-idiotypic activity to anti-erythrocyte autoantibodies in (NZB x CBA)F, but not NZB mice. A B/W F,-derived, monoclonal antiDNA antibody was reported capable of inducing an anti-idiotype response in B/W F, animals, followed by a decrease in disease severity; however, untreated animals did not have similar auto-anti-idiotypic activity.12s These observations support the idea that anti-idiotypic antibodies, which normally suppress autoimmune responses, were not functioning properly in autoimmune mice. This weakness in NZB mice may be a general trait not limited to autoantigens. An inability to regulate down the anti-TNP response was also seen in NZB mice.129 Interestingly, this deficiency in generating an antiidiotypic response was comparable to that of athymic mice, suggesting that this humoral abnormality is thymus-linked. Anti-idiotypic antibodies may also enhance an autoimmune response, as seen in B/W F, animals.24 In this example, rabbit anti-anti-DNA immunoglobulin increased the DNA-binding affinity of B/W F, autoantibodies by altering their DNA binding sites. The idiotype network may also regulate the immune response through the generation of T-suppressor function. In vitro studies have demonstrated the need for accessory cells in generating suppressor activity,130*t31and an adoptive transfer experiment has shown a defect in the ability of NZB B cells to induce T-suppressor activity, implying a possible role for the idiotype network in this abnormality.132 NON-B/T CELLS

The functions of NZB-derived non-B/T cells have also been examined, although not as extensively as lymphocytes. The production of prostaglandin E (PGE) by B/W F, peritoneal mac-

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rophages were reported decreased,133 and a possible IL-1 defect was noted in NZB splenic dendritic cells.134 Natural killer (NK) activity was also altered in autoimmune mice. An assay for NK activity against young thymocyte targets provided evidence of increased resistance to lysis by NZB and B/W F’ thymocytes while NZB spleen cells showed decreased killing ability’35; this suggested a possible mechanism by which autoreactive T cells may escape elimination. Conversely, target killing by liver NK cells from B/W F’ mice was abnormally high and increased with age 136;this was concluded to be a product of immune complexes and foreign antigen clearance by the liver. MAJOR HlSTOCOMPATlBlLlTY

COMPLEX

Correlations between the presence of particular major histocompatibility complex (MHC) characteristics and the incidence of autoimmune disease are not uncommon in people and animals. In addition, in vitro systems for the examination of autoantibody production consistently recognized the requirement for MHC compatibiliAnother important demonstration of the ty. 118-120 relationship of Ia gene products and autoimmunity was disease remission in B/W F, animals following treatment with anti-I-A’ antibodies (Table 5),13’ a possible consequence of a direct effect of anti-Ia antibodies on B cells.13*Perhaps the mechanism involved in such B-cell inactivation may be found in a report by Vaickus et a113’ in which monoclonal antibodies to HLA class 2 molecules were reported to induce a cytostatic influence on HLA-DR+ cells by cross-linking class 2 molecules. Table 6. MHC Involvement

in the Disease of

Another indication of MHC involvement in autoimmunity is the production of autoantibodies during GVH reactions.113112In experimental GVH, hybrid mice are injected with spleen or lymph node cells from one of the parental strains. The ensuing response is thought to be due to donor cell recognition of allogeneic H-2 molecules on host tissues. In fact, the lymphoid cell infiltration of NZB tissues were thought to resemble GVH disease.* However, the role of MHC in SLE may be somewhat more stringent than what is required for inducing GVH disease. In an experimental GVH model for SLE, Portanova et a114’reported that severe renal disease required an association with a particular Ia haplotype (H-zb) but ANA production was not required. Although the MHC has been the primary histocompatibility region examined, increasingly attention is being focused on the minor histocompatibility (MIH) or minor lymphocyte-stimulating (MIS) antigens.14’ This poorly described system for self-recognition, which is actively involved in tissue rejection and mixed lymphocyte cultures, may also be involved in autoreactivity. In fact, increased NZB T-cell reactivity to MIH antigens has been reported.14* Another series of experiments provided supporting information on the participation of H-2 in B/W F’ autoimmunity. By backcrossing B/W F’ mice with parental strains, a correlation was detected between H-2 heterozygosity (H-2’/Hzd) and the development of murine systemic 1upus.‘43,‘USimilar backcross experiments in the SNFl animal model also demonstrated a link between autoimmune lupus and the I-A- and T cell receptor (TcR)-chain genes of the normal SWR parental strain.14’ THYMIC INFLUENCES ON AUTOIMMUNITY

New Zealand Mice MHC restriction observed during in vitro autoantibody production. Administration of anti-la antibody reduced autoantibody levels in viva. Similarities in characteristics of murine lupus and GVH dis8858. Haterozygosity at the H-2 is linked to SLE in B/W F, and SNFl mica. Genetic studies suggest linkages of autoimmunity genes to the MHC. Possible aberrant la expression on apithalial cells, including thymic tissues, may invite abnormal MHC-mediated T-call development.

The thymus is a lymphoid organ of central importance in the development of T cells. Abnormalities in this tissue will have an enormous impact on the homeostasis of the immune system and the regulation of an immune response. The NZB model of autoimmunity is characterized by distinct thymic problems (Table 6). The NZB mice, as in several known lupus mouse strains, show accelerated thymic atrophy.’ An extensive histological analysis of NZB and B/W F, thymuses was also performed by de Vries and Hijmans.* They reported high lymphoid cell

MURINE LUPUS

233

Table 6. Summary of the Thymic Characteristics

of

New Zealand Mice Early involution and degeneration of epithelial structure. Infiltration by plasma cells. Reduced proliferative ability of thymic epithelial cells in culture. No increase in KLl ’ epitheliel cells following in viva hydrocortisone treatment. Studies on athymic/thymectomized

mice and thymic reconati-

tution suggested that thymuaea from young animals have normalizing effects on autoimmunity, whereas thoae from older animals have detrimental effects. la density on thymic epithelial cell monolayera may influence the SMLF characteristics of thymocytea following coculture with thymic cell monolayera.

densities in the vascular spaces of the medulla followed by plasma cell infiltration in both NZB and B/W F, mice. Patterns of epithelial degeneration differed between these two animal models. While drastic reductions in NZB medullary epithelial cells were seen within a month after birth, neonatal B/W F, mice displayed hyperplasia instead. With age, this cell population in B/W F, animals increased, accompanied by the development of Hassall’s corpuscles and other epithelial cell aggregates. At approximately 6 months of age, decreases in lymphocyte infiltration were noted, followed by epithelial degeneration and the appearance of lymphoid follicles. Subsequent studies also reported the inability of NZB, and the reduced ability of B/W F,, thymic epithelial cells to proliferate in vitro3 and identified the infiltrating lymphoid cells in B/W F, thymuses as plasma and immunoglobulin-positive B cells.‘46 Unlike BALB/c controls, the ratio of dull Thy-l+, Lyt-l+, dull PNA+ to bright Thy-l +, Lyt-I +, bright PNA+ thymocytes increased with age in the B/W F, hybrids. Thymic degeneration is a relatively early characteristic of autoimmune development in NZB mice and can be observed at a younger age than detectable circulating NTA.3 One means of measuring thymic influences on autoimmunity is thymic removal, either by thymectomy or the development of athymic nude mice. The results of such studies have been variable and include data suggesting that thymectomy increases, decreases, or does not influence surviva1.489’473’48 Thymic engraftments have also been used to study thymic influences in NZ mice. Following neonatal thymectomy of B/W Fi mice, reconstitution with thymuses from 2-week-old

syngeneic mice was able to prevent the otherwise characteristic acceleration of disease, whereas thymuses from lo-week-old animals would not.i4’ Finally, the transfer of neonatal B/W nu/+ thymuses into B/W nu/nu mice resulted in a disease pattern comparable to that of euthymic B/W F, animals.48 These studies do tend to support a function for the thymus in disease development and, in particular, the negative properties of those from older autoimmune mice. The interpretation of data based on thymectomy or nude mice studies will also need to consider recent reports of T cells bearing yS receptors, cell populations that may develop via an extrathymic pathway.‘49 Thymic influences on the severity of autoimmune disease have also been studied in the context of gender and sex hormones. In general, female NZB and B/W F, animals have shorter life spans than their male counterparts. Older (~24 weeks) B/W F, mice demonstrate a decrease in responsiveness to T cell mitogens in females but not in males.15’ Androgen therapy improves the survival of aged male B/W F, mice, along with a greater proliferative responses of spleen cells to phytohemagglutinin (PHA).“’ Sex hormones are known to effect immunity by their actions on the thymus’52 and this may be one way gender influences the autoimmune response in B/W F, animals. Recently, thymosin treatment of castrated male B/W F, mice was shown to prevent the disease-accelerating effects of castration1s3; the authors suggest that castration may have increased the rate of thymic involution, a process that is reduced by thymosin. How thymic abnormalities influence NZB and B/W F, disease is not clear; however, thymic differences have been noted between autoimmune and nonautoimmune strains of mice. Thymic epithelial cells expressing the KLl marker increased in number following in vivo administration of hydrocortisone into normal mice. This increase was not seen in NZB animals.‘54 Thymic epithelial cell monolayers from normal and young B/W F, mice elevated Thy-l expression on the surfaces of spleen cells from nude Swiss mice and increased their responsiveness to Con A; NZB and old B/W F, thymic epithelia did not have this effect. Thymic epithelial cells of BALB/ c origin were also able to normalize NZB spleen (but not lymph node) cells’ ability to respond to


234

Con A as well as reduce their proliferative capacity in unirradiated BALB/c micea Another series of experiments centered on the function of bone marrow pre-T cells following coculture with thymic epithelial cells to characterize NZB thymic abnormalities. As an indicator of thymocyte function, the syngeneic mixed lymphocyte (SMLR) reaction was used. Lower responses of NZB thymocytes, regardless of the source of stimulator cells, and depressed IL-2 activity in the SMLR culture supernatants of NZB cells were noted. NZB thymic epithelial cell cultures also displayed lower IL-l activity and Ia expression, and higher PGE2 and PGE3 levels in comparison to normals.155*‘56An extension of this study showed that treatment of NZB thymic epithelial cell cultures with interferon-y was able to increase Ia expression on these cells. Preincubation of NZB thymocytes on these treated monolayers resulted in increased response activity, in the SMLR.“’ These studies suggest that the functional abnormalities in the T-cell compartment of NZB mice may have been partially because of improper education, an outcome possibly related to la density on thymic reticuloepithelial cells. GENETICS

The immunologic problems associated with NZB mice are generally recognized as manifestations of underlying genetic disorders. Although the nature of these disorders is not clear, information on the genetic influences of NZB autoimmunity shows that multiple genes are involved in disease expression and these genes are dominant or codominant. Recombinant inbred lines have been used to study the genetics of NZB autoimmunity. Datta et al”* report that B-cell hyperactivity, autoantibody production, and abnormal autologous mixed lymphocyte reactions were independent phenomena. Information on the control of NTA and anti-ssDNA antibody production by separate single genes as well as multi-gene regulation of anti-erythrocyte antibody synthesis has also been presented by Raveche et a1,‘59 who have also reported associations of NTA production with increased serum IgM levels and splenomegaly with chromosomal hyperdiploidy. The results of a further studynj’ propose that interactions between two groups of nonlinked genes underlie the

immunopathologic characteristics of NZB mice. The first group was associated with the expression of polyclonal B-cell hyperactivity, CFU-S abnormalities, endogenous mouse leukemia virus (MuLV), and NTA. The other was related to the production of antibodies to MuLV, ssDNA, and erythrocytes, and tolerance defects to bovine gamma-globulin. There were no observed associations with H-2. The results of studies by Chused et a1i61show that increases in splenic null cells were governed by a single recessive gene, an involvement of three genes in increased plasma cell production, and the role of either of two separate genes for heightened Ly-l + B cell numbers. The abnormal enlargement of Lyt-2+ T cells has also been shown to be inherited as a semidominant trait, possibly linked to B-cell activation (perhaps a reflection of B-cell involvement in T-suppressor function); an additional pair of genes may be important in the expression of renal disease. Overall, these data have not identified specific chromosomal sites for these autoimmunity genes, and so studies on well-characterized genes have been instituted. For the most part, NZB immunoglobulin genes are normal and have no inherent inclination toward a pathological autoimmune response.16* But the allotypy of Igh constant regions has been linked to the electric charge of anti-ssDNA antibodies and their quantity of production,‘63 and Knight and Adams’64 suggest that NZB and B/W F, disease may be linked to immunoglobulin genes. Examinations of T-cell receptor genes also uncovered deletions in the P-chain loci of both the NZW and SWR strains of mice’453’65;however, the relevance of such findings to the pathogenesis of SLE has not been resolved. The relationship of the endogenous viral protein, gp70, with glomerulonephritis has also been assessed with variable results. Proteinuria was reported proportional to the levels of serum gp70 in the B/W F, mouse mode1,‘66 whereas SNFl studies report no link between gp70 and histologically identifiable glomerulonephritis’67; the latter group did show a correlation between gp70 and lymphomas.‘68 The most consistent association of autoimmunity characteristics with a specific genetic locus has been seen with histocompatibility. Knight and Adams’64 report a close linkage of one NZB

235

MURINE LUPUS

nephritis gene to the H-2 and MIH connections to one nephritis and one hemolytic anemia gene. Backcross experiments on the B/W F, model provided information on H-2 associations with NTA, anti-DNA antibodies, anti-gp70 antiFurthermore, both bodies, and proteinuria. 166~169 parental strains were thought to contribute two dominant genes each to the anti-dsDNA response, one which was linked to chromosome 17’43;the NZW genes were believed responsible for the IgM-to-IgG switch. Kotzin and Palmer’@ also reported a requirement of H-2 heterozygosity for anti-dsDNA IgG production in B/W F, animals; they suggested that a relevant gene may rest approximately 10 centi Morgans (CM) from the MHC of the NZW (H-Y) parent. Heterozygosity for I-A was also necessary for the full expression of glomerulonephritis in SNFl mice.14’ Finally, a possible abnormality in the tumor necrosis factor-a locus, which maps to the H-2, was discovered in NZW mice.96 Certain generalizations on the diseases of NZB and NZB-derived hybrid mice can be made based on the preceding genetic studies, and there is some agreement with the cellular and humoral abnormalities seen in these animals. In NZB mice, polyclonal B-cell hyperactivity appears linked to endogenous stem cell hyperactivity, increased numbers of Ly-I+ B cells, and IgM hypergammaglobulinemia. These characteristics may be most closely associated with NTA production, less so with anti-erythrocyte antibodies, and minimally with the anti-DNA response. Linkage to H-2 is not seen, although an involvement of NZB chromosome 4 is evident. Alternatively, in B/W F, animals, the anti-DNA response is not strongly linked to hypergammaglobulinemia, but is influenced by chromosome 17 (H-2) of both parental strains and chromosome 6 of NZW mice’609170; this H-2 association may be a reflection of the l-cell dependency of murine lupus. CONCLUSION

There are several key directions that are currently of importance in understanding the pathogenesis of autoimmunity in NZ mice. They include (1) a clarification of the niche held by natural autoantibodies in the development of pathologic autoimmunity, (2) the role of regulatory T cells in the autoantibody response, (3) the regulatory functions of B cells, (4) the impor-

tance of the MHC in guiding reactivity to self, (5) the influence of thymic degeneration on autoimmunity, (6) the identification of immunogens that participate in the induction of disease, and (7) the genetic bases underlying these considerations. The search for a primary cause of the autoimmunity in New Zealand mice has been hampered by the large number of immunologic aberrations in these animals, each of which could conceivably be connected to other elements of the immune system. Because of the network nature of the immune system, a defect in one component will probably result in the generation of problems in others. The disease of NZ mice has in the past been compared with that of an aged immune system, especially in regard to thymic involution, the accelerated development of bone marrow B cells, the loss of tolerance of self-antigens, and associations with immune deficiencies. However, NZB autoimmunity may also be an example of the arrested development of the immune system. It is interesting that some of the components of autoimmunity, including natural autoantibodies and Ly-I+ B cells, are present during the development of healthy mice. Self-recognition is thought to be an essential part of the normal ontogeny of immunity, perhaps as a means of expanding the immune repertoire before generalized contact with exogenous antigens. For example, the multispecificity of natural autoantibodies may be of some benefit in protecting the host from microbial invasion before the maturation of an adaptive immune system. Pathological autoimmunity may be an exaggeration of this otherwise normal physiological process. Allied to this idea are proposals that Ly-I+ B cells and T cells with yS receptors may be more primitive than their respective counterparts, Ly-l- B cells and ~$3 TcR T cells, in terms of phylogeny as well as ontogeny.‘71*‘72For example, Ly-I + B cells display surface characteristics that suggest a cell that appeared before the full segregation of leukocyte lineages that we now assume compose the majority of cells in a normal, mature mammalian immune system. The possible extrathymic development of yS TcR T cells in murine lupu~‘~~ and Ly-I+ B cells suggests that more thorough examinations of the developmental and comparative aspects of immunity are needed to discover many relevant characteristics of the immune

236


system, essential for effectively dealing with autoimmune diseases. The data thus far argue for antigen-driven, T-cell dependent mechanisms in overt autoimmunity. T lymphocytes have generally been thought of as central to the regulation of an immune response and homeostasis of the immune system, but research is now uncovering aspects of the B cell suggesting that it may be as important to immune regulation. Networking with other cells via idiotype interactions, the secretion of lymphokines, and antigen presentation are now gaining more credence as legitimate B-cell functions and thereby increase the known range of B-cell behaviors. For example, B cells were recently reported to release IL- 1173and can now be named possible antigen presenters. Thus, B-cell activities, other than their production of autoantibodies, should be investigated as elements of autoimmunity. Autoreactive B cells are believed to exist throughout the life of an animal, although their potential for autoantibody production may be greatest during ontogeny (ie, natural autoantibodies). Such cells may be especially important in the autoimmune response as presenters of antigens to T cells and as constituents of the idiotype network. Therefore, they could possibly direct B and other cell types toward selfreactivity. Such a mechanism may be operating in lupus mice, expressing accelerated thymic degeneration and lacking other regulatory components that normally keep autoreactive B cells in check. One might consider the possibility of B cells, perhaps derived from populations which produce natural autoantibodies or those of the Ly-I+ lineage, in bending the T-cell repertoire toward autoreactivity. This may be of particular interest when also considering the antigenpresenting capabilities of B cells,1739174the reported influence of B cells on T-cell development,130’175 and the possible lack of proper T-cell education in murine lupus. The feedback of stimulatory signals from such T cells to B-cell populations may result in a reinforced, mature autoimmune response. Such a scenario could be

one explanation for the observation of both polyclonal and T cell/antigen-mediated mechanisms in the disease of NZB mice. Studies on the antigen presenting capabilities of autoreactive B cells and the selectability of the T-cell repertoire of NZ mice may be illuminating. Isotype switching in the autoimmune response may also be indicative of certain T- and B-cell interactions. The predominance of IgG2a suggests that interferon-y may be involved; the secretion of this lymphokine is thought to be primarily a function of Thl, rather than Th2 cells. Interestingly, a review of the properties of cytotoxic T cells shows that in terms of lymphokine production, they resembled Th 1 more closely than Th2 cells.‘76 Cytotoxicity may also be a prominent characteristic of cells which express y6 T-cell receptors.“’ Recently, B cells have been reported to have the ability to provide the necessary signals for the in vitro generation of cytotoxic T cells.“s Large numbers of immature pre-T cells are known to be in the spleens of NZB mice.“’ In all, the production of IgG2a may be a reflection of changes in the ratios of T-cell types present in NZ mice, a possibility that should be investigated. This in turn may result from thymic abnormalities and the inability of this organ to accommodate proper T-cell education, as well as the possibility for B-cell-influenced T-cell development in extrathymic sites. The proposition that T cells in NZ mice may be more malleable than those of normal strains underlies the above discussion of B-cell influences on T-cell development and function. The possibility of defective T-cell education and development in NZ mice is supported by observations that the properties of NZB thymic epithelial cells appear abnormal before the appearance of all the major autoantibodies, as well as thymectomy and reconstitution studies reporting thymic influences on autoimmunity. Relatedly, the greater likelihood for adults rather than children to develop GVH disease following bone marrow transplantation may be due to thymic involutions that would have presumably occurred in adults.‘*’

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Autoimmunity and tumor resistance. 3-Methylcholanthrene tumorigenesis in New Zealand Black mice.

Genetic studies of autoimmunity and retrovirus expression in crosses of New Zealand black mice. II. The viral envelope glycoprotein gp70.

Genetic studies of autoimmunity and retrovirus expression in crosses of New Zealand black mice I. Xenotropic virus.

Autoimmunity in the pathogenesis of hypertension.

Acquisition of autoimmunity genes by New Zealand mice is associated with natural resistance to infection by mycobacteria.

Duke-Elder Lecture: new concepts on the role of autoimmunity in the pathogenesis of uveitis.

Sialoadhesin deficiency does not influence the severity of lupus nephritis in New Zealand black x New Zealand white F1 mice.

The Potential Roles of Bisphenol A (BPA) Pathogenesis in Autoimmunity.

Autoimmunity to chaperonins in the pathogenesis of arthritis and diabetes.

Exploring autoimmunity in the pathogenesis of abdominal aortic aneurysms.

Autoimmunity: an underlying factor in the pathogenesis of hypertension.

The organization of radial glial fibers in spontaneous neocortical ectopias of newborn New Zealand black mice.

Modeling EBV infection and pathogenesis in new-generation humanized mice.

Age-related decline in the antibody response to E. coli lipopolysaccharide in New Zealand Black mice.

A new beginning in autoimmunity?

Avian malaria in New Zealand.

Bovine photosensitization in New Zealand.

Medical research in New Zealand.

Letter: Duovirus in New Zealand.

Asthma deaths in New Zealand.

Asthma deaths in New Zealand.

Outcome measurement in New Zealand.

Physician assistants in New Zealand.

Avian reoviruses in New Zealand.