Expression of liver fatty acid binding protein alters growth and differentiation of embryonic stem cells.

Although expression of liver fatty acid binding protein (L-FABP) modulates cell growth, it is not known if L-FABP also alters cell morphology and differentiation. Therefore, pluripotent embryonic stem cells were transfected with cDNA encoding L-FABP and a series of clones expressing increasing levels of L-FABP were isolated. Untransfected ES cells, as well as ES cells transfected only with empty vector, spontaneously differentiated from rounded adipocyte-like to fibroblast-like morphology, concomitant with marked reduction in expression of stage-specific embryonic antigen (SSEA-1). These changes in morphology and expression of SSEA-1 were greatest in ES cell clones expressing L-FABP above a threshold level. Immunofluorescence confocal microscopy revealed that L-FABP was primarily localized in a diffuse-cytosolic pattern along with a lesser degree of punctate L-FABP expression in the nucleus. Nuclear localization of L-FABP was preferentially increased in clones expressing higher levels of L-FABP. In summary, L-FABP expression altered ES cell morphology and expression of SSEA-1. Taken together with the fact that L-FABP was detected in the nucleus, these data suggested that L-FABP may play a more direct, heretofore unknown, role in regulating ES cell differentiation by acting in the nucleus as well as cytoplasm.

Sterol carrier protein-2 localization in endoplasmic reticulum and role in phospholipid formation.

Although sterol carrier protein-2 (SCP-2; also called nonspecific lipid transfer protein) binds fatty acids and fatty acyl-CoAs, its role in fatty acid metabolism is not fully understood. L-cell fibroblasts stably expressing SCP-2 were used to resolve the relationship between SCP-2 intracellular location and fatty acid transacylation in the endoplasmic reticulum. Indirect immunofluorescence double labeling and laser scanning confocal microscopy detected SCP-2 in peroxisomes > endoplasmic reticulum > mitochondria > lysosomes. SCP-2 enhanced incorporation of exogenous [(3)H]oleic acid into phospholipids and triacylglycerols of overexpressing cells 1.6- and 2.5-fold, respectively, stimulated microsomal incorporation of [1-(14)C]oleoyl-CoA into phosphatidic acid in vitro 13-fold, and exhibited higher specificity for unsaturated versus saturated fatty acyl-CoA. SCP-2 enhanced the rate-limiting step in microsomal phosphatidic acid biosynthesis mediated by glycerol-3-phosphate acyltransferase. SCP-2 also enhanced microsomal acyl-chain remodeling of phosphatidylethanolamine up to fivefold and phosphatidylserine twofold, depending on the specific fatty acyl-CoA, but had no effect on other phospholipid classes. In summary, these results were consistent with a role for SCP-2 in phospholipid synthesis in the endoplasmic reticulum.

Sterol carrier protein-2 alters high density lipoprotein-mediated cholesterol efflux.

Although sterol carrier protein-2 (SCP-2) participates in the uptake and intracellular trafficking of cholesterol, its effect on "reverse cholesterol transport" has not been explored. As shown herein, SCP-2 expression inhibited high density lipoprotein (HDL)-mediated efflux of [(3)H]cholesterol and fluorescent 22-(N-(7-nitrobenz-2-oxa-1, 3-diazol-4-yl)amino)-23,24-bisnor-5-cholen-3b-ol (NBD-cholesterol) up to 61 and 157%, respectively. Confocal microscopy of living cells allowed kinetic analysis of two intracellular pools of HDL-mediated NBD-cholesterol efflux: the highly fluorescent lipid droplet pool and the less fluorescent pool outside the lipid droplets, designated the cytoplasmic compartment. Both the whole cell and the cytoplasmic compartment exhibited two similar kinetic pools, the half-times of which were consistent with protein (t(b)(12) near 1 min) and vesicular (t(d)(12) = 10-20 min) mediated sterol transfer. Although SCP-2 expression did not alter cytoplasmic sterol pool sizes, the rapid t(b)(12) decreased 36%, while the slower t(d)(12) increased 113%. Lipid droplets also exhibited two kinetic pools of NBD-cholesterol efflux but with half-times over 200% shorter than those of the cytoplasmic compartment. The lipid droplet slower effluxing pool size and t(d)(12) were increased 48% and 115%, respectively, in SCP-2-expressing cells. Concomitantly, the level of the lipid droplet-specific adipose differentiation-related protein decreased 70%. Overall, HDL-mediated sterol efflux from L-cell fibroblasts reflected that of the cytoplasmic rather than lipid droplet compartment. SCP-2 differentially modulated sterol efflux from the two cytoplasmic pools. However, net efflux was determined primarily by inhibition of the slowly effluxing pool rather than by acceleration of the rapid protein-mediated pool. Finally, SCP-2 expression also inhibited sterol efflux from lipid droplets, an effect related to decreased adipose differentiation-related protein, a lipid droplet surface protein that binds cholesterol with high affinity.

Expression of fatty acid binding proteins is altered in aged mouse brain.

Brain membrane lipid fatty acid composition and consequently membrane fluidity change with increasing age. Intracellular fatty acid binding proteins (FABPs) such as heart H-FABP and the brain specific B-FABP, detected by immunoblotting of brain tissue, are thought to be involved in fatty acid uptake, metabolism, and differentiation in brain. Yet, almost nothing is known regarding the effect of age on the expression of the cytosolic fatty acid binding proteins (FABPs) or their content in brain subfractions. Electrophoresis and quantitative immunoblotting were used to examine the content of these FABPs in synaptosomes in brains from 4, 15, and 25 month old C57BL/6NNia male mice. Brain H-FABP and B-FABP were differentially expressed in mouse brain subcellular fractions. Brain H-FABP was highly concentrated in synaptosomal cytosol. The level of brain H-FABP in synaptosomes, synaptosomal cytosol, and intrasynaptosomal membranes was decreased 33, 35, and 43%, respectively, in 25 month old mice. B-FABP was detected in lower quantity than H-FABP. More important, B-FABP decreased in synaptosomes, synaptic plasma membranes, and synaptosomal cytosol from brains of 25 month old mice. In contrast to H-FABP, B-FABP was not detectable in the intrasynaptosomal membranes in any of the three age groups of mice. In conclusion, expression of both H-FABP and B-FABP was markedly reduced in aged mouse brain. Age differences in brain H-FABP and B-FABP levels in synaptosomal plasma membranes and synaptosomal cytosol may be important factors modulating neuronal differentiation and function.

Expression and intracellular processing of the 58 kDa sterol carrier protein-2/3-oxoacyl-CoA thiolase in transfected mouse L-cell fibroblasts.

Although the sterol carrier protein 2 (SCP-2) gene encodes for two proteins, almost nothing is known of the function and potential processing of the larger transcript corresponding to the 58 kDa sterol carrier protein-2/3-oxoacyl-CoA thiolase (SCP-x), in intact cells. L-cell fibroblasts transfected with cDNA encoding for the 58 kDa SCP-x protein had a 4.5-fold increase in SCP-x mRNA transcript levels. Western blot analysis showed SCP-x protein expression reached 0.011% of total protein, representing a 4.1-fold increase over basal levels. Surprisingly, the 13.2 kDa SCP-2 protein also increased 2-fold in the transfected cells. This was consistent with part of the 58 kDa SCP-x being proteolytically processed to 13.2 kDa SCP-2 as there was no evidence of an mRNA transcript corresponding to a 13.2/15.2 kDa gene product in the transfected L-cell clones. Confocal immunofluorescence microscopy of transfected L-cells showed that SCP-x/SCP-2 co-localized in highest concentration with catalase in peroxisomes, but significant amounts appeared extra-peroxisomal. Overexpression of SCP-x significantly altered cholesterol uptake and metabolism. Uptake of exogenous [3H]cholesterol and total cholesterol mass were increased 1.9- and 1.4-fold, respectively, in SCP-x expressors. Although cholesterol ester mass was unaltered, incorporation of exogenous [3H]cholesterol and [3H]oleic acid into cholesteryl esters increased 2.3- and 2.5-fold, respectively. These results from intact cells suggest the 13.2 kDa SCP-2 can arise from the larger SCP-2 gene product and indicate a role for the 58 kDa SCP-x protein in cholesterol uptake and intracellular cycling.

Spontaneous and engineered mutant mice as models for experimental and comparative pathology: history, comparison, and developmental technology.

During the last half-century pathologists have explored the biologic mechanisms associated with inherited human and veterinary diseases by using inbred and inbred mutant (spontaneous) strains of mice. The first successful gene transfer to mice by pronuclear injection of the herpes simplex virus thymidine kinase gene and rabbit and human beta-globulin genes was achieved in the early 1980s. This accomplishment was followed a few years later with the creation of a mouse bearing a disrupted hypoxanthine phosphoribosyl transferase (hrpt) gene (targeted mutation based on ES cell blastocyst injection). Since then, hundreds of genetically engineered models of biomedical importance have been created. The unprecedented scale and scope of development of engineered models present great opportunities as well as experimental challenges to the investigator. The aim of the present review is to provide a framework of information on engineered mouse models from the perspective of experimental and comparative pathology research. Sections include: 1) a brief historical account of the development of mouse models of disease, with increasing progression of genetic refinement as represented by inbred (spontaneous) and congenic (targeted) mutant strains of mice; 2) a synopsis of spontaneous and targeted mutations, with anecdotal examples of expression of individual genes and interactions between multiple mutant genes; 3) selected examples of targeted mutations of interest to developmental and cancer biologists and immunologists; 4) an overview of the technology of development of transgenic mice; and 5) an introduction to on-line database resources of current multi-species genomic information.

Cellular differentiation and I-FABP protein expression modulate fatty acid uptake and diffusion.

The effect of cellular differentiation on fatty acid uptake and intracellular diffusion was examined in transfected pluripotent mouse embryonic stem (ES) cells stably expressing intestinal fatty acid binding protein (I-FABP). Control ES cells, whether differentiated or undifferentiated, did not express I-FABP. The initial rate and maximal uptake of the fluorescent fatty acid, 12-(N-methyl)-N-[(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-octadec anoic acid (NBD-stearic acid), was measured in single cells by kinetic digital fluorescence imaging. I-FABP expression in undifferentiated ES cells increased the initial rate and maximal uptake of NBD-stearic acid 1.7- and 1.6-fold, respectively, as well as increased its effective intracellular diffusion constant (Deff) 1.8-fold as measured by the fluorescence recovery after photobleaching technique. In contrast, ES cell differentiation decreased I-FABP expression up to 3-fold and decreased the NBD-stearic acid initial rate of uptake, maximal uptake, and Deff by 10-, 4.7-, and 2-fold, respectively. There were no significant differences in these parameters between the differentiated control and differentiated I-FABP-expressing ES cell lines. In summary, differentiation and expression of I-FABP oppositely modulated NBD-stearic acid uptake parameters and intracellular diffusion in ES cells.

Beta 2-microglobulin-deficient mice are protected from hypergammaglobulinemia and have defective antibody responses because of increased IgG catabolism.

The goal of this study was to determine whether class I proteins play an important role in the regulation of Ig and to elucidate the mechanism(s) involved. We analyzed the phenotype imposed by a null allele of beta 2-microglobulin (beta 2m). Serum Ig levels of several mouse strains showed a beta 2m dependence that was most evident in mice genetically predisposed to develop chronic systemic lupus erythematosus, was preferential to IgG isotypes, and was greatly exaggerated in aging mice that normally develop hypergammaglobulinemia. Beta 2m-deficient mice, regardless of genetic background, also displayed a substantial reduction of specific Ab in response to a prototypic T cell-dependent Ag and a prototypic T cell-independent 2 Ag. This reduction could be accounted for by a selective diminution of Abs of the IgG class. Therefore, class I proteins play a considerable role in the regulation of Ig. The beta 2m dependence could not be explained by class I-dependent immunoregulatory cells (CD8+ cells, NK1.1+ T cells, or conventional NK+ cells) or by the transfer of maternal IgG into the prenatal/neonatal mouse made possible by the beta 2m-dependent Fc receptor (FcRn). However, a beta 2m-dependent increase in the half-lives of IgG, presumably conferred by lifelong FcRn expression, was observed in all mice regardless of genetic background and age. We conclude that FcRn-mediated protection of IgG from catabolism is a generic mechanism that best explains the lifelong beta 2m dependence of Ig in both normal and pathologic situations.

Acyl-CoA binding proteins: multiplicity and function.

The physiological role of long-chain fatty acyl-CoA is thought to be primarily in intermediary metabolism of fatty acids. However, recent data show that nM to microM levels of these lipophilic molecules are potent regulators of cell functions in vitro. Although long-chain fatty acyl-CoA are present at several hundred microM concentration in the cell, very little long-chain fatty acyl-CoA actually exists as free or unbound molecules, but rather is bound with high affinity to membrane lipids and/or proteins. Recently, there is growing awareness that cytosol contains nonenzymatic proteins also capable of binding long-chain fatty acyl-CoA with high affinity. Although the identity of the cytosolic long-chain fatty acyl-CoA binding protein(s) has been the subject of some controversy, there is growing evidence that several diverse nonenzymatic cytosolic proteins will bind long-chain fatty acyl-CoA. Not only does acyl-CoA binding protein specifically bind medium and long-chain fatty acyl-CoA (LCFA-CoA), but ubiquitous proteins with multiple ligand specificities such as the fatty acid binding proteins and sterol carrier protein-2 also bind LCFA-CoA with high affinity. The potential of these acyl-CoA binding proteins to influence the level of free LCFA-CoA and thereby the amount of LCFA-CoA bound to regulatory sites in proteins and enzymes is only now being examined in detail. The purpose of this article is to explore the identity, nature, function, and pathobiology of these fascinating newly discovered long-chain fatty acyl-CoA binding proteins. The relative contributions of these three different protein families to LCFA-CoA utilization and/or regulation of cellular activities are the focus of new directions in this field.

beta2-microglobulin dependence of the lupus-like autoimmune syndrome of MRL-lpr mice.

MRL-lpr/lpr mice develop a distinctive immunologic disease characterized by accumulation of unusually large numbers of T cells in the peripheral lymphoid organs. Most of the accumulating T cells express an alpha beta-TCR but are peculiar in that they express neither CD4 nor CD8 co-ligands. Concurrent with lymphoaccumulation of such double negative (DN) T cells, MRL-lpr/lpr mice develop a lethal systemic lupus erythematosus-like autoimmune syndrome. This study focuses on the role of MHC class I molecules in this latter pathologic process. Highly backcrossed class I molecule-deficient MRL and MRL-lpr mice carrying a functionally defective allele of the gene beta 2-microglobulin (B2m) were produced. Class I deficient MRL-lpr/lpr mice demonstrated a substantial reduction in DN T cells, confirming other reports indicating that most DN T cells arise from progenitors positively selected on MHC class I molecules. Significantly, class I-deficient MRL-lpr/lpr mice also demonstrated a diminution of every autoimmune disease indicator analyzed including hypergammaglobulinemia; autoantibodies including anti-DNA, anti-Smith antigen, and rheumatoid factor; and glomerulonephritis. The results indicate that class I-dependent T cells are crucial not only for the development of DN T cells, but for multiple features of the MRL-lpr/lpr systemic lupus erythematosus syndrome. Moreover, the pattern of hypergammaglobulinemia suggests that the requirement for MHC class I proteins is restricted temporally to later stages of the disease.

Kinetic and organ-specific patterns of cytokine expression in acute graft-versus-host disease.

Although many cytokines have been previously implicated in graft-versus-host disease (GVHD), no study to date has comprehensively evaluated their expression over time or in different tissues affected by GVHD. Using a semi-quantitative reverse transcriptase-PCR technique and a murine model of acute GVHD, we have evaluated the expression levels of mRNA for a wide range of cytokines in spleen, gut and liver tissues at weekly intervals after bone marrow transfer. The earliest cytokine responses seen were increases in IL-2, IL-10, IFN-gamma, MIP-1 alpha and TNF-alpha in the spleen, suggesting a primarily Th1 pathway. Other cytokines (IL-1 alpha, IL-10 and MIP-1 alpha) were persistently elevated in GVHD mice, but were variable depending on the tissue. These data demonstrate that a wide range of cytokines are involved in the GVHD response and that their kinetic pattern of expression is different in various affected tissues.

Single and combined humoral and cell-mediated immunotherapy of Pneumocystis carinii pneumonia in immunodeficient scid mice.

Homozygous mutant scid/scid (severe combined immunodeficiency) mice (referred to as scid mice) lack both specific humoral and cell-mediated immune functions and are exemplary in vivo models for analysis of host-parasite relationships. In our colony, scid mice routinely and predictably develop spontaneous Pneumocystis carinii pneumonia (PCP) with high morbidity. Previous studies have identified both T cells (specifically, CD4+ cells) and antibody as independent mechanisms of effective anti-P. carinii resistance; however, CD4+ T cells also cause an often fatal hyperinflammatory reaction. The current study has explored the optimal application of these immune components for conferring protection against P. carinii. Anti-P. carinii hyperimmune serum was highly effective at reducing the number of P. carinii organisms in early, intermediate, and advanced stages of PCP and was capable of increasing the mean life expectancy of P. carinii-infected scid mice by more than threefold if provided on a continuing basis. When a short course of hyperimmune-serum therapy was provided prior to transfer of P. carinii-sensitized normal lymphocytes, scid mice were rendered permanently free of P. carinii without the pathological sequelae of the hyperinflammatory reaction. These findings are discussed in the contexts of mechanism and clinical relevance.

Lethal exacerbation of Pneumocystis carinii pneumonia in severe combined immunodeficiency mice after infection by pneumonia virus of mice.

Mice homozygous for the mutant allele scid (severe combined immunodeficiency) have been described as excellent models for Pneumocystis carinii (Pc) pneumonia (PCP), a major health problem in patients with acquired immune deficiency syndrome (AIDS) and other immunodeficiency states. Other microorganisms have been shown to infect AIDS patients simultaneously with Pc, but whether one opportunist is able to directly influence the pathogenicity of another has not been determined previously. We have deliberately coinfected scid mice (with extent Pc infection) with a variety of primarily pneumotropic viruses and bacteria and have identified pneumonia virus of mice as causing a dramatic increase in the density of Pc organisms and the morbidity due to PCP in immunodeficient scid mice. This finding has clinical significance in the management of PCP, in that the identification and treatment of coinfecting pneumotropic pathogens may be as important as treatment targeted at Pc. A search for other synergistic (or antagonistic) microorganisms and determination of their mechanism(s) of action in altering the progression of PCP is indicated.

Multiple mechanisms of tumorigenesis in E mu-myc transgenic mice.

Transgenic mice bearing a c-myc oncogene under control of the immunoglobulin heavy chain enhancer (E mu-myc mice) reproducibly develop and die from tumors of the B lymphocyte lineage (J.M. Adams, A.W. Harris, C.A. Pinkert, L.M. Corcoran, W.S. Alexander, S. Cory, R.D. Palmiter, and R.L. Brinster, Nature (Lond.), 318: 533-538, 1985; W.Y. Langdon, A. W. Harris, S. Cory, and J.M. Adams, Cell 47: 11-18, 1986; A.W. Harris, C.A. Pinkert, M. Crawford, W.Y. Langdon, R.L. Brinster, and J.M. Adams, J. Exp. Med., 167: 353-371, 1988; reviewed in S. Cory and J.M. Adams, Annu. Rev. Immunol., 6: 25-48, 1988). Analysis of lymphocytes obtained by serial sampling of peripheral blood from individual hemizygous (E mu-myc/0) and homozygous (E mu-myc/E mu-myc) transgenic mice indicates that proliferation in the original host and transplantability into histocompatible recipients are distinct properties that can be acquired independently and in either order. These two types of transgenic mice differ in that homozygous mice have about one-fourth the life span of hemizygous mice and develop polyclonal, non-transplantable tumors in comparison to the oligoclonal, highly transplantable malignancies seen in hemizygous animals. In conclusion, the overall concept of malignancy is best viewed as an aggregate of the separable parameters of cellular proliferation, clonality, tissue invasiveness, metastasis, and (experimental) transplantability. The E mu-myc transgenic mouse represents an attractive model in which to investigate the multistep nature and alternative pathways of tumorigenesis.

Both immunity and hyperresponsiveness to Pneumocystis carinii result from transfer of CD4+ but not CD8+ T cells into severe combined immunodeficiency mice.

The opportunistic pathogen Pneumocystis carinii (Pc) is considered to be the leading cause of morbidity in patients with AIDS. It is important, therefore, to determine the immunological mechanisms of resistance to Pc. We have taken advantage of the lack of both T and B lymphocytes in severe combined immunodeficiency (scid) mice to determine the critical factors in resistance to spontaneously acquired Pc pneumonia. Using adoptive transfer of unfractionated or fractionated lymphocyte subsets or hyperimmune serum from congenic normal donors, we have demonstrated that effective immunity to Pc results from the action of CD4+ but not CD8+ T cells (in the absence of antibody) or from humoral immunity (in the absence of T cells). However, responses of CD4+ T cells (but not antibody) to already well-established burdens of Pc are often accompanied by a fatal hyperinflammatory reaction. The activity of CD4+ T cells against Pc thus illustrates a broadly applicable principle that T cell immunity represents a critical balance between consequences beneficial and harmful to the host.

Differences defined by bone marrow transplantation suggest that lpr and gld are mutations of genes encoding an interacting pair of molecules.

Homozygosity for either of the lymphoproliferation (lpr) or generalized lymphoproliferative disease (gld) mutations of mice causes the development of systemic lupus erythematosus-like autoimmune syndromes that are characterized by severe lymphadenopathy and highly elevated serum immunoglobulin levels. Although the mutations are nonallelic, analysis of homozygous lpr/lpr and gld/gld mice on the same strain background has indicated that the pathology and severity of the autoimmune syndromes induced by these mutations are indistinguishable. To explain this, it has previously been suggested that lpr and gld may represent mutations in molecules involved in sequential steps of an intracellular metabolic pathway of T cells. We have now investigated the behavior of both lpr and gld in a variety of bone marrow chimeras and have found that functional differences between lpr and gld become apparent after bone marrow transfer. Transfer of lpr/lpr bone marrow to irradiated congenic +/+ recipients caused the development of a graft-vs.-host-like lymphoid wasting syndrome, whereas transfer of gld/gld bone marrow to +/+ recipients resulted in development of a gld-like autoimmune syndrome. Additionally, gld/gld hosts behaved like +/+ hosts irrespective of the genotype of the donor bone marrow, whereas lpr/lpr hosts behaved unlike +/+ hosts when reconstituted with either lpr/lpr, gld/gld, or +/+ bone marrow. These are the first clear differences between these two mutations yet described. Our studies indicate that the molecule altered by the gld mutation is expressed only by bone marrow-derived cells, whereas the molecule altered by the lpr mutation is expressed by both bone marrow-derived cells and by one or more peripheral radioresistant cell populations. To reconcile these differences with the fact that homozygous lpr/lpr and gld/gld mice are indistinguishable, we suggest an alternative model for the relationship between the lpr and gld mutations in which the two molecules affected represent an interacting ligand-receptor pair expressed by different cells.

Bone marrow transplantation from mutant lpr/lpr mice. Functional abnormalities rather than alloantigenic differences appear to determine the development of a graft-vs.-host-like syndrome.

Transfer of bone marrow (BM) from autoimmunity-prone mice homozygous for the lymphoproliferation (lpr) mutation to irradiated congenic +/+ recipients has previously been shown to result in a syndrome similar to chronic graft-vs.-host (GVH) disease. It has been suggested that this syndrome may be due to an antigenic difference caused by the lpr mutation itself or to antigenic differences at loci closely linked to the lpr locus (Theofilopoulos, A. N. et al., J. Exp. Med. 1985. 162:1; Mosbach-Ozmen, L. and Loor, F., Thymus 1987. 9:197). However, the results presented here indicate that alloantigenic differences do not play a role in this syndrome. Instead, the chronic disease observed in lpr/lpr----(+/+) BM chimeras appears to develop as a result of a functional defect associated with the lpr mutation which is expressed shortly after transfer of lpr/lpr BM to irradiated recipients. This defect causes an increase in the levels of serum IgG1 and IgG2, which peak at 4-5 weeks post-transfer and then decline to normal levels by 9-10 weeks post-transfer. Inflammation similar to that observed in classic GVH reactions accompanies excess IgG production in congenic +/+ recipients but not in lpr/lpr recipients of lpr/lpr BM. We demonstrate that the GVH-like response occurring in lpr/lpr----(+/+) chimeras is dependent on mature T cells, but that either lpr/lpr or (+/+) T cells can support this reaction. These results suggest that transfer of lpr/lpr BM to normal mice causes immunoregulatory disturbances which lead to nonspecific activation of T cells. We speculate that lpr/lpr BM causes a GVH-like reaction in +/+ recipients but a systemic lupus erythematosus-like syndrome in lpr/lpr recipients because of intrinsic differences in the +/+ and lpr/lpr host environments. Considering these findings, the lpr/lpr----+/+ GVH model may be useful for analysis of factors capable of inducing undesirable reactions in clinical BM transplantation between nominally histocompatible donors and recipients, in addition to being informative about the nature of the lpr mutation itself.

Spontaneous Pneumocystis carinii pneumonia in immunodeficient mutant scid mice. Natural history and pathobiology.

The opportunistic pathogen Pneumocystis carinii (Pc) poses a major clinical health problem in individuals with immune deficiency, including those patients with human immunodeficiency (HIV)-associated acquired immune deficiency disease (AIDS). Heretofore, in vivo investigations of the biology of Pc and pathogenesis of pneumocystosis have generally employed steroid-induced immune suppression with antibiotic prophylaxis and protein deprivation. This approach has many drawbacks, chief among them being the widespread, multiple interacting effects caused by the inducing agents. Athymic (nude) mice and rats have been used, but are less than ideal, as the immune defect primarily affects T lymphocytes. This article describes the natural history, pathobiology, and environmental effects on Pc pneumonitis in nonaxenically housed mice homozygous for the autosomal recessive mutation 'severe combined immunodeficiency' (scid), which almost totally lack both cell-mediated and antibody-mediated immune functions. The predictability, unequivocal expression, high morbidity, and well-defined genetic basis make scid/scid mutant mice the model of choice for in vivo studies of spontaneous pneumocystosis.

Effect of xid on autoimmune C3H-gld/gld mice.

The xid gene was introduced into C3H-gld/gld mice to determine its effects on the development of autoimmune disease. C3H-gld/gld.xid mice were compared with C3H-gld/gld mice for the development of lymphadenopathy, surface phenotype of lymph node (LN) cells, c-myb oncogene RNA production, serum immunoglobulin (Ig) levels, and autoantibody production. In addition, C3H-gld/gld and C3H-lpr/lpr mice were examined for serum Ig and autoantibody levels. The results showed that the xid gene had no effect on either the development of the severe lymphadenopathy characteristic of C3H-gld/gld mice or the phenotype of the Ly-2-, L3T4-, Ly-5(B220)+ T-cell subset that is expanded in the LN and spleens of these mice. Similarly, xid did not affect the high levels of c-myb oncogene RNA expression by C3H-gld/gld LN and spleen cells. By contrast, the xid gene caused a significant reduction in serum IgM but not IgA levels and almost completely ablated the generation of both IgM and IgG anti-ssDNA antibodies and anti-dsDNA antibodies. These data suggest that the xid gene can dramatically decrease the B-cell manifestations of autoimmunity in gld homozygotes without affecting their abnormal T-cell expansion. Comparisons of age-matched C3H-gld/gld and C3H-lpr/lpr mice showed that they had similarly elevated serum IgM and IgA levels and anti-ssDNA and anti-dsDNA antibody levels providing further evidence that gld and lpr produce parallel defects in C3H mice.