Paracrine stimulation of human renal fibroblasts by proximal tubule cells

Paracrine stimulation of human renal fibroblasts by proximal tubule cells. BACKGROUND: Interstitial fibrosis strongly predicts the degree and progression of renal failure in human renal disorders. Since active fibrosis tends to initially occur in a peritubular distribution, the possibility that human proximal tubule cells (PTC) relay fibrogenic signals to neighboring cortical fibroblasts was examined in vitro. METHODS: Cell proliferation (cell counts and thymidine incorporation), total collagen synthesis (proline incorporation), matrix metalloproteinase (MMP) activity (gelatin zymography), and autocrine secretion of insulin-like growth factor-I (IGF-I) were measured in primary cultures of human cortical fibroblasts cocultured with PTC or exposed to PTC-conditioned media (PTCCM). RESULTS: Cell numbers and thymidine incorporation rates were increased in cortical fibroblasts cocultured with PTC (136.4+/-7.3% and 119.3+/-8.2% of control values, respectively, P < 0.05) or incubated in PTC-CM (114.0+/-5.9%, P < 0.05 and 146.7+/-13.3%, P < 0.05, respectively). PTC-CM stimulated cortical fibroblast collagen synthesis (13.5+/-1.0% vs. 10.8+/-0.7%, respectively, N = 24, P < 0.05) and MMP-2 and MMP-9 secretion. Cortical fibroblast secretion of IGF-I binding protein-3 (IGFBP-3), which in turn modulates the autocrine and paracrine actions of IGF-I, was enhanced in the presence of PTC-CM compared with control (1162.2+/-94.2 vs. 969.1+/-58.9 ng/mg protein/day, P < 0.05), but no change was observed in cortical fibroblast secretion of IGFBP-2 (260.9+/-38.8 vs. 290.9+/-36.6 ng/mg protein/day, P = NS) or IGF-I (56.7+/-6.6 vs. 57.0+/-6.8 ng/mg protein/day, P = NS). Human PTC secreted transforming growth factor-beta1 (TGF-beta1) and the AB heterodimer of platelet-derived growth factor (PDGF-AB) in a time-dependent fashion and the augmentation of cortical fibroblasts mitogenesis, collagen synthesis and IGFBP-3 secretion induced by PTC-CM was replicated by exogenous TGF-beta1 and PDGF. Furthermore, the stimulatory effects of PTC on cortical fibroblasts were potentiated in transiently acidified PTC-CM (which activated latent TGF-beta1), and were abrogated by neutralizing antibodies specifically directed against TGF-beta1 and PDGF-AB. Cortical fibroblasts in turn released a soluble factor(s) into cortical fibroblast-conditioned media that reciprocally stimulated PDGF-AB production by PTC (4.79+/-1.55 vs. 0.78+/-.06 ng/mg protein/day, P < 0.05). CONCLUSIONS: PTC modulate the biological behavior of neighboring cortical fibroblasts in the human kidney through paracrine mechanisms, which include the production and release of PDGF-AB and TGF-beta1. Renal insults that result in proximal tubule injury may perturb this paracrine interaction, thereby culminating in excessive fibroblast proliferation and interstitial fibrosis.     

Dual pathways for organic anion secretion in renal proximal tubule

Transport on the "classical" organic anion system in renal proximal tubule is specific, active, Na-dependent, and ouabain sensitive. Here we review recent studies using intact teleost proximal tubules and laser scanning confocal microscopy which show that the secretion of large organic anions, such as, fluorescein-methotrexate (FL-MTX, Mw 923 Da) is handled by a separate and distinct organic anion transport system. In contrast to the classical system, FL-MTX uptake into cells and secretion into the tubular lumen was ouabain insensitive and largely Na-independent. KCN did not affect cellular uptake but abolished secretion into the lumen. PAH and probenecid, potent inhibitors of transport on the classical system, were weak inhibitors of FL-MTX transport. Uptake and secretion of FL-MTX were inhibited by micromolar concentrations of other organic anions (MTX, folate, bromocresol green, bromosulfonphthalein). FL-MTX secretion into the lumen was inhibited by leukotriene C4 and cyclosporine A, neither of which affected transport of the model substrate for the classical system, fluorescein. Thus, FL-MTX secretion is specific, but largely Na-independent and ouabain-insensitive. Both the basolateral and luminal steps in FL-MTX transport differ from those associated with fluorescein and P-aminohippurate secretion.     

Predominant expression of monoamine oxidase B isoform in rabbit renal proximal tubule: regulation by I2 imidazoline ligands in intact cells

Previous studies have shown that a subpopulation of the catecholamine-degrading enzymes monoamine oxidase (MAO) A and B holds a previously unknown regulatory site, the I2-imidazoline binding site (I2BS). In the present work, we characterized the isoforms of monoamine oxidases expressed in the rabbit renal proximal tubule, defined their relationship with I2BS, and investigated the ability of I2BS ligands to inhibit enzyme activity in intact cells. Two findings indicate that MAO-B is the predominant isoform expressed in the renal proximal tubule cells: 1) Western blot performed with an anti-MAO-A/MAO-B polyclonal antiserum revealed a single 55-kDa band corresponding to MAO-B; 2) enzyme assays showed an elevated MAO-B activity ([14C]beta-phenylethylamine oxidation: Vmax = 1.31 +/- 0.41 nmol/min/mg protein), whereas MAO-A activity was only detectable ([14C]5-HT oxidation: Vmax = 80.3 +/- 19 pmol/min/mg protein). Photoaffinity labeling with the I2BS ligand [125I]2-(3-azido-4-iodophenoxy)-methylimidazoline revealed a single 55-kDa band, which indicates that MAO-B of the renal proximal tubule cells holds the I2 imidazoline binding site. [3H]Idazoxan binding studies and enzyme assays showed that, in intact cells, I2BS ligands bind to and inhibit MAO-B. Indeed, the increase in the accessibility of intracellular compartment by cell permeabilization did not enhance [3H]idazoxan binding, which indicates that, in intact cells, intracellular I2BS are fully occupied by imidazoline ligands. In addition, enzyme assays showed that incubation of proximal tubule cells with imidazoline ligands leads to a complete, dose-dependent inhibition of MAO activity. These data show the predominant expression of MAO-B in rabbit renal proximal tubule and its regulation by imidazoline ligands in intact cells.     

Effect of intraluminal bicarbonate and chloride on fluid absorption by the rat renal proximal tubule

In order to study mechanisms of fluid transport in the rat renal proximal convoluted tubule, the effects of large variations in intraluminal HCO3- and Cl- concentrations were measured by microperfusion techniques. No differences in rates of fluid transport were found when intraluminal HCO3- was varied from 4 to 30 mEq/liter and Cl- from 146 to 120 mEq/liter. Inhibition of H+ secretion with benzolamide had no effect on fluid absorption when little or no HCO3- was present in the lumen, but did reduce fluid transport when 25 mEq of HCO3- was present. If several different mechanisms are responsible for proximal fluid transport, such as nonelectrogenic active NaHCO3 transport, passive chloride diffusion and active sodium transport linked to H+ secretion, the above observations imply that they all operate at approximately the same rate, since the dominant driving force would have been different with each perfusion solution. The data seem more compatible with the view that active sodium transport is the major driving force for fluid absorption in the proximal tubule, that this is not linked to H+ secretion and that anions modify the rate of absorption only to the degree that they are able to accompany sodium across the epithelium. An additional observation was that absorption of isotonic NaCl was very slow in short segments of tubule, as compared to HCO3--containing perfusion solutions. Although the mechanism is uncertain, these data suggest that a finite amount of intraluminal HCO3- is necessary for optimal proximal fluid transport.     

Inhibition of proximal convoluted tubule transport by dopamine

BACKGROUND: Dopamine can produce a natriuresis and diuresis independent of changes in renal hemodynamics. However, previous studies have failed to demonstrate an inhibition of transport by dopamine in intact proximal convoluted tubules. METHODS: Rabbit proximal convoluted tubules were perfused in vitro with an ultrafiltrate-like solution and bathed in a serum-like albumin solution. RESULTS: In the present study, the addition of 10-5 M dopamine to the lumen or bath of proximal convoluted tubules perfused in vitro had no effect on transport. In proximal convoluted tubules, addition of 10-6 M bath norepinephrine increased the rate of volume absorption from 0.65 +/- 0.08 to 0.93 +/- 0.08 nl/mm. min (P < 0.01). Addition of 10-5 M luminal dopamine in the presence of bath norepinephrine inhibited the rate of volume absorption to 0.72 +/- 0.10 nl/mm. min (P = 0.01). The inhibition in the rate of volume absorption by luminal dopamine in the presence of bath norepinephrine was completely blocked by the DA1 antagonist, SCH 23390. The DA1 agonist luminal 10-5 M fenoldopam also inhibited volume absorption in the presence of bath norepinephrine, but the DA2 agonist luminal 10-5 M quinpirole was without effect. Bath 10-5 M dopamine had no effect on volume absorption in the presence of bath norepinephrine. CONCLUSION: Dopamine has no direct epithelial action on the proximal convoluted tubule. However, luminal dopamine antagonizes the stimulation in transport produced by norepinephrine. These studies suggest that luminal dopamine may play a role to modulate sodium transport in the presence of renal nerve activity.     

Protein kinase C isozyme expression and down-modulation in growing, quiescent, and transformed renal proximal tubule epithelial cells

Renal alpha-protein kinase C (PKC) is rapidly down-modulated modulated in animals treated with the renal toxin and tumor promoter, folic acid (Dong et al., Cancer Res., 53: 4542-4549, 1993). To further explore the role of PKC isozymes in renal growth and carcinogenesis, we compared phorbol ester receptor and PKC isozyme content, distribution, and regulation in primary and oncogene-altered rat renal proximal tubule epithelial cells (RPTE) in culture. Immunoblot analysis and RNase protection assays indicated that RPTE expressed at least four PKC isozymes, alpha, delta, epsilon, and zeta. Total phorbol ester receptors were decreased in primary proliferating, E1A-immortalized, and SV40-transformed RPTE compared to primary quiescent RPTE. The decrease in PDBu binding was largely due to a specific decrease in alpha-PKC protein content to approximately 50% of the level in quiescent RPTE. Degradation rates and message levels were compared to determine the mechanism for the decrease in alpha-PKC. Whereas alpha-PKC message levels in quiescent and proliferating primary RPTE were comparable, alpha-PKC degradation was increased in proliferating cells. These results indicate that the decreased alpha-PKC content was due largely to increased turnover. Phorbol ester stimulated the rate of degradation, thus demonstrating a link between degradation rate and PKC activation. These results suggest that the increased basal degradation rate in proliferating and oncogene-altered cells reflects an increase in activity of PKC in these cells.     

Induction of rabbit renal mixed-function oxidases by phenobarbital: cell specific ultrastructural changes in the proximal tubule

Administration of phenobarbital (60 mg/kg) daily for 4 days to male rabbits resulted in induction of renal cytochrome P-450 (3.5-fold) and a corresponding increase in ethoxycoumarin-O-deethylase and benzphetamine-N-demethylase activity (17- and 4-fold, respectively). Kidney weight to body weight ratio and renal ethoxyresorufin-O-deethylase were not affected by phenobarbital pretreatment. Numerous focal areas of proliferation of smooth endoplasmic reticulum (SER) were evident in proximal tubule cells from phenobarbital treated rabbits while proximal tubular cells from control rabbits had only small and sparcely located aggregates of SER. Phenobarbital-induced SER proliferation was specifically localized to the S3 segment of the proximal tubule. Proliferation was not observed in S2 cells of the proximal tubule, cells of Henle's loop, distal tubules, or collecting tubules in rabbits pretreated with phenobarbital. These data demonstrate the biochemical heterogeneity of cell types within the proximal tubules of rabbits. Furthermore, induction of mixed-function oxidases specifically in S3 cells of the proximal tubule may be of toxicological significance in the metabolic activation of certain nephrotoxicants.     

Immunologic characterization of plasma membranes from the renal proximal tubule of the dog

Plasma membrane fractions from the brush border (BBM) and antiluminal (ALM) surfaces of the dog's renal proximal tubule cell were separated using free-flow electrophoresis. Rabbits immunized with BBM rapidly produced antibody, but rabbits immunized with ALM did not respond. Indirect immunofluorescence and immunoferritin studies showed that the antibody reacts with the brush border of the proximal tubules in the normal kidney of the adult dog. It also reacts with the surface membranes of certain other absorptive and secretory epithelia, such as gall bladder, small intestine, epididymis, and lacrimal gland. The antibody has affinity for the membrane maltase without affecting its catalytic activity, but does not appear to have affinity for the membrane alkaline phosphatase or the high affinity binding site for phlorizin present in the BBM. Polyacrylamide electrophoresis of solubilized BBM showed approximately 37 protein bands and four glycoproteins. We conclude that the proximal tubule cell is immunologically polarized with respect to the distribution of antigenic proteins, and that the BBM is highly antigenic. The antigenic components appear to be high molecular weight glycoproteins present in the glycocalyx.     

Voltage and cosubstrate dependence of the Na-HCO3 cotransporter kinetics in renal proximal tubule cells

The voltage dependence of the kinetics of the sodium bicarbonate cotransporter was studied in proximal tubule cells. This electrogenic cotransporter transports one Na+, three HCO3-, and two negative charges. Cells were grown to confluence on a permeable support, mounted on a Ussing-type chamber, and permeabilized apically to small monovalent ions with amphotericin B. The steady-state, di-nitro-stilbene-di-sulfonate-sensitive current was shown to be sodium and bicarbonate dependent and therefore was taken as flux through the cotransporter. Voltage-current relations were measured as a function of Na+ and HCO3- concentrations between -160 and +160 mV under zero-trans and symmetrical conditions. The kinetics could be described by a Michaelis-Menten behavior with a Hill coefficient of 3 for HCO3- and 1 for Na+. The data were fitted to six-state ordered binding models without restrictions with respect to the rate-limiting step. All ordered models could quantitatively account for the observed current-voltage relationships and the transinhibition by high bicarbonate concentration. The models indicate that 1) the unloaded transporter carries a positive charge; 2) the binding of cytosolic bicarbonate to the transporter "senses" 12% of the electric field in the membrane, whereas its translocation across the membrane "senses" 88% of the field; 3) the binding of Na+ to the cotransporter is voltage independent.     

Increased functional load on mouse kidney proximal tubule epithelial cells causes changes in nucleolar 3-D architecture

Ultrastructural 3-D analysis of nucleolar architecture and Ag-NOR protein distribution in mouse kidney-cortex proximal-tubule epithelium has been performed. A principal scheme of structural changes of the nucleolus and organization of its components during the intensification of pre-rRNA synthesis (dynamic model of a nucleolus) based on computer spatial modelling has been advanced. According to the nucleolar composition, three groups of cells, which differ from each other by rRNA synthesis, are defined in normal kidney. Most nephron proximal-section cells (about 52%) are characterized by lower activity of RNA synthesis. Such kind of cells are defined as group I (nucleolar diameter 0.7-1.5 microm) and always contain resting, ring-shaped or close to ring-shaped dense nucleoli, which have 2 or 3 fibrillar centers. Nucleoli of group II cells (about 37%, nucleolar diameter 1.5-2.5 microm) have a higher level of activity, contain 4-7 fibrillar centers, and their structural organization is close to reticulated forms due to the first indications of vacuolar network (identified as prereticulated nucleoli). The most active cells of group III (about 11%, nucleolar diameter 2.5-3.5 microm) include cells with typical reticulated nucleoli with a well expressed vacuolar network and numerous fibrillar centers (18-22). Increased functional load of the epithelium caused by unilateral nephrectomy and diuretic (4-chlor-H [2-furylmethyl] 5-sulphamyl-antranic acid) injection changed the proportion of the different cell groups: group I decreased (about 25%), whereas groups II and III increased (about 8% and 17%, respectively). The increase of nucleolar activity first causes a deformation of the individual fibrillar centers as well as complication and growth of their surface. Further, a progressive fragmentation of the fibrillar centers and the growth of their total volume is observed. The complication and growth of the total volume of Ag-positive zones is another indication of the nucleolar activation. The vacuolar system develops by a gradual fusion of small isolated cavities into a united vacuolar network. Nucleoli with 2-7 fibrillar centers are considered to be intermediate forms reflecting successive stages of its activation or inactivation: from the resting ring-shaped nucleolus via transient stages of increasing functional activity to the active reticulated nucleoli and vice versa. The observed differences in the nucleolar ultrastructure are regarded as evidence of the functional heterogeneity of cell populations within one functional segment of nephron.     

Shiga toxin-1 regulation of cytokine production by human proximal tubule cells

BACKGROUND: Interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor (TNF) levels are elevated in kidneys of patients with post-diarrheal hemolytic uremic syndrome (D+HUS) and may contribute to renal dysfunction. The renal cellular sources of these inflammatory cytokines in D+HUS are largely unknown, however, the proximal tubule has emerged as a potentially important candidate. Since Shiga toxin-1 (Stx-1) has been implicated in the genesis of D+HUS, we examined the effect of Stx-1 on cytokine production by human proximal tubule cells. METHODS: Stx-1 cytotoxicity, protein synthesis inhibition, and effect on IL-1, IL-6, and TNF protein release and mRNA levels were determined. The effect of another protein synthesis inhibitor, cycloheximide (CHX), on these parameters was also evaluated. RESULTS: Stx-1 greatly increased TNF release and mRNA levels while CHX, at concentrations that produced similar inhibition of protein synthesis, had no effect on TNF production. In contrast, Stx-1 and CHX caused comparable elevations in IL-1 release and mRNA accumulation. Stx-1 and CHX also stimulated IL-6 mRNA accumulation, but only at concentrations that either were cytotoxic or substantially blocked protein synthesis. Finally, lipopolysaccharide, which is likely to be elevated in the circulation of patients with D+HUS, had no effect alone, but synergized with Stx-1 to increase IL-1 production. CONCLUSIONS: These results indicate that Stx-1 stimulates proximal tubule inflammatory cytokine production and that this effect is due partially to nonspecific induction of mRNA levels as well as activation of Stx-1-specific mechanisms.     

Multiple gene duplication and expression of mouse bcl-2-related genes, A1

Atrial reentrant tachycardia (ART) was ablated in an anatomically guided approach. Five patients with ART underwent 2 linear incisions without careful pace or activation mapping. One line was from an atrial activation site earlier than P wave onset to the nearest fixed anatomic conduction barrier, i.e., the inferior vena cava or coronary sinus ostium. The other line was made just above or closely crossed the first line vertically. Mean application time was 29 +/- 19 minutes, and the application energy was 14,001 +/- 12,322 joules. Mean follow-up after ablation was 15 +/- 10 months. Three patients underwent electrophysiologic study three months after and sustained ART was not induced. All patients were free of sustained tachycardia events without antiarrhythmic drugs during the postoperative clinical course. Although anatomically guided ablation for ART requires much time and energy, it is easily and effectively done without careful activation or pace mapping, and is indicated if ablation using activation mapping or entrainment technique fails to cure the ART.     

Colloidal silica-coated tissue culture dishes for primary cell cultures: growth of rabbit renal proximal tubule cells

The use of colloidal silica as a substratum for primary cultures of differentiated cells has significant advantages over classic tissue culture polystyrene. In this report, the growth and the level of expression of differentiated function of primary rabbit renal proximal tubule (RPT) cell cultures on colloidal silica is examined, using hormonally defined serum-free medium. Primary RPT cells grew to confluence more rapidly on colloidal silica than on tissue culture polystyrene (TC+). Moreover, following three passages, the RPT cells increased in number threefold more than parallel cultures on TC+. The morphology of primary RPT cells on colloidal silica were found by means of transmission electron microscopy to possess a polarized morphology with a brush border, and differentiated markers were retained even after passaging, including the Na+/glucose cotransport system and Glut 7.     

Distinct interleukin-1beta-converting enzyme family proteases mediate cisplatin- and staurosporine-induced apoptosis of mouse proximal tubule cells

Interleukin-1beta converting enzyme (ICE) family proteases (caspases) are known to be implicated as important effectors of apoptotic pathways. The purpose of this study was to elucidate the role of ICE family proteases in apoptosis of mouse cells derived from the terminal proximal tubule (S3) treated with cisplatin, an anti-tumor drug, or staurosporine, a protein kinase C inhibitor. For this purpose, we measured the activities of ICE family proteases and examined the effects of tetrapeptide and viral ICE family protease inhibitors on the activities of ICE family proteases in and the degree of apoptosis of S3 cells treated with cisplatin and staurosporine. RT-PCR analysis revealed that S3 cells as well as mouse kidney express mRNA for ICE and CPP32, an ICE family protease. Results of enzymatic analysis, determination the degree of DNA fragmentation and cytotoxicity test suggest that CPP32 mediates cisplatin-induced apoptosis of S3 cells, whereas ICE family proteases other than CPP32 mediate staurosporine-induced apoptosis of S3 cells. In conclusion, distinct ICE family proteases mediate apoptosis of mouse proximal tubule cells depending on the stimuli to which the cells are exposed.     

Intrinsic differences in various segments of the proximal convoluted tubule

Until recently it has not been possible to compare directly the function of superficial and juxtamedullary nephrons. The present studies, using in vitro microperfusion, were designed to examine whether functional differences exist between proximal convoluted tubule segments of superficial and juxtamedullary nephrons. Electrophysiological studies showed that major differences exist between the relative chloride and sodium permeabilities of these segments. In the 1st mm of the superficial proximal convoluted tubule, the permeability to sodium was greater than that to chloride, whereas in the 2nd mm of the superficial proximal convoluted tubule and all later segments, the permeability to chloride was greater than that to sodium. The juxtamedullary proximal convoluted tubule was found to differ from the superficial proximal convoluted tubule in two respects: first, the relative permeabilities to chloride and sodium did not differ in the various segments of the juxtamedullary proximal convoluted tubule; second, the permeability to sodium was greater than to chloride throughout. When perfused with a solution lacking glucose and amino acids, the superficial and juxtamedullary convolutions exhibited the same transepithelial potential change, a reversible decrease to less than -- 1 mV. It thus appears that in both convolutions there exists electrogenic sodium transport coupled to the transport of these organic solutes. This differs from pars recta of both of these nephrons, which have been shown to exhibit electrogenic sodium transport independent of organic solutes. However, when perfused with a solution lacking glucose and amino acids but also containing high chloride and low bicarbonate concentrations, the superficial convolution developed a significantly more positive potential than the juxtamedullary. This difference reflects greater relative chloride permeability in the superficial proximal convolution. These studies show that intrinsic functional differences exist between proximal convoluted tubules obtained from the superficial and juxtamedullary nephron populations.     

Isolation, mapping, and functional expression of the mouse X chromosome glycerol kinase gene

Glycerol kinase (Gyk) participates in the metabolism of endogenously derived and dietary glycerol. Deficiency of the human enzyme activity is an X-linked recessive disorder with a clinical picture varying from childhood metabolic crisis to asymptomatic adults incidentally identified by hyperlipidemia screening (pseudohypertriglyceridemia). Gyk is a member of a small group of kinases termed ambiquitous enzymes that are found in the cytosol or as membrane-bound enzymes associated with the voltage-dependent anion channel of the mitochondrial outer membrane. It was recently reported that in humans there are X-linked and autosomal copies of Gyk sequences, both apparently functional genes and processed pseudogenes. To understand the role of Gyk in normal metabolism and the variable clinical features seen with Gyk deficiency, we have characterized the mouse Gyk gene. We present the sequence of a full-length mouse Gyk cDNA that is alternatively spliced in brain. The Gyk gene was mapped to the mouse X chromosome by both fluorescence in situ hybridization and an interspecies backcross panel, demonstrating conservation of synteny with dmd. To confirm the functional identity of the cDNA, transient transfection of the cDNA into COS7 cells was shown to cause a marked elevation in glycerol kinase activity.     

Host modifier genes affect mouse autoimmunity induced by the lpr gene

A defect in apoptotic signal transmission through CD95 is an essential genetic mechanism for lymphoproliferation and autoimmunities in lpr or gld mice. However, disease manifestations are largely affected by the host genetic background. To identify and map such host genes modifying lpr gene effect, ie, the lpr modifier (Lprm) genes, 82 MRL/lpr x (MRL/lpr x C3H/lpr) F1 mice were subjected to immunopathological and genetical analyses. High-grade vasculitis and glomerulonephritis among backcross mice were observed in separate groups of mice. Microsatellite analysis revealed that there were two host genes affecting the occurrence of vasculitis, Lprm1 (chromosome 4) and Lprm2 (chromosome 3). A recessive MRL allele at Lprm1 enhanced vasculitis to occur in both sexes, whereas that of Lprm2 inhibited its development selectively in females. Genotype combinations of these two genes explained the severity of vasculitis in crosses of MRL/lpr and C3H/lpr mice and also the vasculitis-prone recombinant inbred strain McH5/lpr. A recessive MRL allele at Lprm3 (chromosome 14) suppressed glomerulonephritis. The weight of the spleen was increased by a recessive MRL allele at Lprm4 (chromosome 5) yielding a logarithm of odds score of 2.02 in a quantitative trait locus analysis. In contrast, the weight of axillary lymph nodes was increased by a recessive MRL allele at a locus on chromosome 2, but its presence was not supported by the quantitative trait locus analysis. The titer of anti-dsDNA autoantibody was controlled by the locus Lprm5 on chromosome 16, which had an logarithm of odds score of 3.41. Possible candidate genes for Lprm genes deduced from their map locations are discussed and compared with the autoimmunity genes reported thus far. In conclusion, autoimmune disease manifestations by the lpr mutation are affected by multiple host genes separately.