Processing of IgA Aggregates in a Rat Model of Chronic Liver Disease

CLINICAL IMMUNOLOGY AND IMMUNOPATHOLOGY

Vol. 84, No. 2, August, pp. 107–114, 1997Article No. II974357

Processing of IgA Aggregates in a Rat Model of Chronic Liver Disease

Alessandro Amore,* Dario Roccatello,†,1 Giuseppe Picciotto,‡ Steven N. Emancipator,§ Roberto Ropolo,‡Giovanni Cacace,‡ Adolfo Suriani,Ø Bruno Gianoglio,* Luigi Massimino Sena,\ Paola Cirina,*

Gianna Mazzucco,Ø Victor Alfieri,** Giuseppe Piccoli,** Rosanna Coppo,*and Pier Giuseppe De Filippi‡

*Nephrology and Dialysis Department, Regina Margherita Hospital, Turin, Italy; †Immunopathology Center and Renal Clinical Physiopathology,G. Bosco Hospital, Turin, Italy; ‡Nuclear Medicine Department, S. Giovanni Battista Hospital, Turin, Italy; ØDepartment of Biomedical Sciences

and Human Oncology, \Department of Experimental Medicine and Oncology, and **Department of Nephrorology, University of Turin, Italy;and §Institute of Pathology, Case Western Reserve University, Cleveland, Ohio 44106

in control rats injected with aggregated IgG; i.e., MTTHeavy alcohol intake and/or lipotrope-deficient diet values were more prolonged in rats given whiskey or

induced hepatocellular injury and mesangial deposition LD diet than normally nourished rats (P õ 0.01). Theof IgA and often IgG in Lewis rats. The experimental lipotrope-deficient diet and the chronic alcohol abuseanimals showing more severe urinary abnormalities and per se seem to lead to critical changes in hepatic uptakehistologic damage in the glomeruli had increased levels and catabolism of both an IgA and an IgG aggregate,of IgA antibodies to dietary antigens and altered intesti- which could account in turn for the reported appear-nal permeability. Based on human studies, the pro- ance of renal immunoglobulin deposits in this experi-longed circulation of IgA-containing complexes associ- mental model. Due to the comparable delay in removalated with the liver disease could be envisaged as im- of IgA and IgG probes in equally nourished animals, ad-portant for the development of mesangial IgA deposits. ditional factors are likely to be involved in the promi-In order to verify this hypothesis, four groups (G) of nent deposition of IgA. q 1997 Academic PressLewis rats were studied: G1 received thrice a week anintragastric infusion of 1.5 ml/100 g body wt of whiskey;G2 rats were nourished with lipotrope-deficient diet; G3 INTRODUCTIONrats were given both whiskey and LD diet; G4 rats werenourished with regular chow. After 12 weeks, heat-ag- Patients with a variety of chronic liver diseases, in-gregated rat monomeric IgA was labeled with 123I and cluding alcoholic cirrhosis, chronic biliary obstruction,intravenously injected. Three control subgroups of rats, cholangitis, and cirrhosis associated with cystic fibrosisone given whiskey, one nourished with LD diet, and one develop, in up to 60% of cases, mesangial deposits ofwith regular chow, were injected with radiolabeled heat- IgA indistinguishable from those observed in IgA ne-aggregated rat IgG. A large field-of-view digital gamma phropathy (IgAN) (1–9).camera, equipped with an ultra-high-resolution collima- We recently developed an experimental model oftor and interfaced to a dedicated computer, was used to chronic liver disease in rats mimicking the human con-analyze tracer kinetics and fate. The liver was the main dition of alcoholism, by using intragastric delivery oforgan involved in clearance of both test probes. The he- whiskey and/or a lipotrope-deficient (LD) diet (10).patic mean transit (MTT) was 114 { 11 min in G1 (pro- Fatty changes in liver with bridging fibrosis and earlyteinuria of 6.9 { 1.41 mg/day and hematuria ////), 221 nodule formation were observed in rats given both{ 19 min in G2 (proteinuria 9.1 { 0.64 mg/day and hema- whiskey and LD diet. These findings were associatedturia//////), and 230{ 15 min in G3 (proteinuria 9.5{

with the appearance of mild mesangial expansion with0.58 mg/day and hematuria //////). In each case MTTgranular deposits of IgA, C3, and often IgG in the glo-value was found to be significantly prolonged comparedmeruli. An increase in xylose intestinal absorption andto G4 (85 { 4 min). The multiple regression analysisin levels of IgA antibodies specific for dietary antigensshowed that MTT values, proteinuria, and hematuriasuggested that the initiating event could be a height-were significantly correlated (P õ 0.01). Controls hadened intestinal permeability. However, rats receivingtrace amount proteinuria (0.82 { 0.17 mg/day, signifi-whiskey or LD alone had normal intestinal permeabil-cantly lower than for each study group, P õ 0.03) andity and high levels of IgA specific for dietary antigensundetectable hematuria. Similar results were obtainedwithout any evidence of polyclonal IgA activation.These rats showed only moderate hepatic fatty1 To whom correspondence and reprint requests should be ad- changes, but significant amounts of glomerular IgA de-dressed at Divisione di Nefrologia e Dialisi e Centro di Immunopato-posits, hematuria, and proteinuria. Therefore, we hy-logia, Ospedale G Bosco, Piazza del Donatore di Sangue 3, 10154

Torino, Italy. pothesized that abnormal clearance kinetics of IgA and

107 0090-1229/97 $25.00Copyright q 1997 by Academic Press

All rights of reproduction in any form reserved.

AID Clin 4357 / a512$$$221 07-02-97 23:17:20 clinal AP: Clin

108 AMORE ET AL.

IgA-containing immune complexes, due to derange- Access to food was always unrestricted. After 3 months,the rats were analyzed for the organ uptake and catabo-ment of some critical liver pathways, underlies the

pathogenesis in this model. lism of the IgA aggregate probe.A selective and prolonged alteration in complement-

Morphologic Studies of Kidneymediated immune clearance was recently described ina different animal model based on single intraperito- A portion of renal cortex was fixed in Serra fluid,neal injection of ethanol (11). Although this ethanol- processed through graded alcohols and xylenes, andinduced decrease in complement-dependent sequestra- embedded in paraffin. Four-micrometer-thick sectionstion and phagocytosis probably has direct implications were stained with periodic acid–Schiff’s reagent andon immune clearance for infectious agents from the Masson’s trichrome.bloodstream (12), the applicability of this effect to the Snap-frozen tissue was sectioned at 4 mm. A directhandling of macromolecular IgA and IgA-containing method was used for the detection of IgG (fluoresceincomplexes is questionable. isothiocyanate-labeled antisera from U.S. Biochemi-In the present report, we explored the possibility that cals, Cleveland, OH). IgA deposits were detected by ana prolonged circulation of IgA immune complexes, due indirect method with a monoclonal antibody specific forto a defective hepatic sequestration, is the basis of the the a chain of rat IgA (Zymed, San Francisco, CA) and aIgA mesangial deposition in our model. fluorescein isothiocyanate-labeled polyclonal antibody

anti-mouse IgG not cross-reactive with rat IgG (U.S.MATERIALS AND METHODS

Biochemicals).Small (1 mm3) fragments of renal cortex were fixedTest Probes

by immersion in 2.5% glutaraldehyde in 0.1 M sodiumRat monomeric IgA (Zymed, South Francisco, CA), cacodilate (pH 7.3). The tissue was postfixed in OsO4,

diluted at 20 mg/ml, was aggregated by heating at 637C embedded in Epon–Araldite. Ultrathin sections offor 60 min. High-performance liquid chromatography almost three glomeruli for each animal, stained withanalysis before and after heating was performed with uranyl acetate and lead citrate, were evaluated. Ana Perkin-Elmer series 10 LC cromatograph (Perkin- arbitrary score ranging from 0 (absence of detectableElmer Corp, Analytical Instruments, Norwalk, CT), Ul- deposits) to 2/ (finely granular to homogeneous elec-traidrogel 1000-column (Waters–Millipore, Mildford, tron-dense deposits in the mesangium of all lobules ofMA), and Bio-Sil TSK 250-column (Bio-Rad Lab, Mu- all glomeruli) was used by a pathologist (G.M.), whonich, Germany). The eluants were measured at ultravi- blindly examined codified specimens.olet adsorbance at 280 nm by a Uvikon 735 LC detector

Organ Uptake and Catabolism of the(Kontron Instruments, Zurich, Switzerland). The IgAImmunoglobulin Aggregate Probeaggregate was labeled with 123I (Sorin, Saluggia, Italy)

by the chloramine T method. One milligram of proteinA large field-of-view (UFO V37,151) Stargem Generalwas labeled with 100 MBq of 123-iodine and 62.5 mg/

Electric digital scintillation camera (Medical System Di-100 g body wt was injected in rat femoral vein. Chroma-vision, Milwauke, WI), equipped with a low-energy/ultra-toscanner controls of labeling efficiency were performedhigh-resolution (4 mm) collimator and interfaced to a ded-before injection.icated computer (Statview), was used for dynamic whole-A control probe of rat IgG (Sigma Chemical Co. Ibody scintigraphic studies of the animals.8015) aggregated by heating at 637C for 30 min at the

The animals, anesthesized with ketamine, wereconcentration of 20 mg/ml was similarly analyzed atplaced in supine position on the camera with both hindhigh-performance liquid chromatography, radiola-legs spread out and fixed with surgical tape.beled, and injected in control animals.

A flow study at 1-sec intervals was obtained for 1min followed by sequential imaging every 30 sec forAnimals240 sec and every 60 sec for 55 min. Radioactivitycounts over heart, liver and, sometimes, bladder wereFour groups of randomized 4-week-old male Lewis

rats (Charles River Breeding Laboratories, Calco, Italy) simultaneously and continuously recorded. The imageswere complemented by computer-generated time–ac-were studied: the first group (14 rats) received intragas-

tric infusions of 1.5 ml/100 g of rat weight of a commer- tivity graphs after manual selection of the region ofinterest. Activity–time curves were obtained by usingcial whiskey (Black Velvet, Black Velvet Distilling Co.,

Toronto, Canada) thrice a week and were nourished a reframe at 1 frame/1 min for 50 min. To determine thedisappearance rate of the test probe from the generalwith regular chow (Piccioni Laboratories, Brescia, Italy);

the second group (13 rats) received an LD diet (Piccioni circulation (heart) a Q BASIC program was employed.It allowed to obtain the fit of the blood clearance curveLabs); the third group (5 rats) received both intragastric

infusions of whiskey and a LD diet; the fourth was the by the peeling method. Each component of the curvewas determined by searching the best fit using the leastcontrol group (14 rats) nourished with regular chow.


109LIVER PROCESSING OF AGGREGATED IgA IN RAT

squares method. Programs were implemented by a se- Organ Uptake and Catabolism of theImmunoglobulin Aggregate Probesnior health physicist (R.R.).and Laboratory Parameters

Analysis of Organ Dynamic CurvesThe static scintigraphies performed to ascertain

As in common tracer techniques involving the rapid whether the test probe administered was comparableinflow of tracer into an organ followed by a more grad- in each rat revealed that 5 rats of whiskey group, 4ual washout, data from the curve of tracer concentra- nourished with lipotrope-deficient diet, 2 given bothtion against time were fitted by means of a function whiskey and LD diet, and 4 normal rats did not receivebased on the form of the distribution: adequate amount of radioactivity. These rats were not

considered for further examination. Evaluated animalsY Å A(t 0 t0)ae0b(t0t0) for t § t0 . included 9 rats given whiskey (6 injected with aggre-

gated IgA and 3 injected with aggregated IgG), 9 ratsgiven LD diet (6 injected with aggregated IgA and 3A fit was carried out using a multiple linear regressioninjected with aggregated IgG), 3 rats given both whis-of Y on the two new variables, X1 and X2 (‘‘g-variatekey and LD diet injected with aggregated IgA, and 10fit’’). g fit was also used to extrapolate the washoutnormal rats (7 injected with aggregated IgA and 3 in-phase of the curve, which was necessary because of thejected with aggregated IgG).effects of recirculation which obscured the later parts

Within a few seconds after injection, aggregated IgAof the washout.reached the liver. Representative imaging and activ-In the general formulation of a g function, the rateity–time profiles in normal and experimental groupsof increase of the organ dynamic graph (reflecting theare shown in Figs. 2 and 3.uptake) is partially dependent on values of a. Most im-

As in other kinetic models in human using aggre-portantly g function allows calculation of the liver meangated immunoglobulins, i.e., aggregated secretory IgAtransit time, reflecting catabolism of the injected probe.of £ 1.5 1 106 daltons (13), cross-linked macromole-cules of ú2 1 106 Da obtained from seric (monomeric)UrinalysisIgA (14) and IgA–IgG aggregates of 1.8 1 106 Da (15,

After urine collection in individual metabolic cages, 16), the time course of radioactivity in the liver of ratsthe amount of proteinuria was evaluated by turbidime- injected with aggregated IgA consisted of three phases:try induced by 3% sulfosalicylic acid and expressed in an initial rapid increase in activity primarily reflectingmg/24 hr, with bovine serum albumin as a standard. blood flow, a slower increase predominantly represent-Microhematuria was detected by Hemastix (Ames, ing extraction of tracer ligand by receptors, and theMiles Inc., Elkhart, IN). eventual decrease expressing lysosomal breakdown.

The hepatic dynamic graph was analyzed by employingStatistics g function. In its general formulation (see Materials

and Methods) the rate of increase of the organ dynamicAn unpaired t test, a linear coefficient of regressiongraph (reflecting the uptake) is partially dependent onand a multiple regression analysis were used. Thevalues of a. Values of a in study groups (whiskey, 0.246mean and the standard error of replicate observations{ 0.08; LD diet, 0.207 { 0.059; and even whiskey andare given in the text.LD diet, 0.127 { 0.22) were not significantly differentfrom controls (0.322 { 0.08). However, each studyRESULTSgroup had values of liver MTT significantly prolongedthan normals (85 { 4 min): whiskey—114 { 11 min,Test ProbesPõ 0.025; LD diet—221{ 19 min, Põ 0.01, comparedboth to normal and whiskey rats; whiskey and LDThe IgA preparation injected, as analyzed by HPLC,diet—230{ 15 min, Põ 0.01 compared to both normalconsisted of a major peak of 11 106 Da (85% of the entireand whiskey rats, but P Å ns compared to LD diet rats.preparation) corresponding to the aggregated form of

Data of heart activity–time profiles were taken asimmunoglobulins, and a minor peak (about 15%) ofrepresentative of blood clearance. Due to initial fluctu-about 1.5 1 105 Da of nonaggregated IgA. The prepara-ations of activity in some cases, the heart dynamic pro-tion proved to be stable over a full range of temperaturesfile was better analyzed in its second part, after thecorresponding to incubations and after 123I labeling.10th minute. The second component of the heart activ-Chromatoscanner analysis indicated a labeling effi-ity curve was assumed to be representative of the dis-ciency ranging between 90 and 93.5% (Fig. 1).appearance of the test probe from the circulation. It isThe IgG preparation consisted of a single peak of 1inversely related to half-time (t1/2 Å log2/l). l values in1 106 Da, i.e., the aggregated form of immunoglobulins.whiskey (0.127 { 0.050 min01), LD (0.087 { 0.055Stability at various temperatures and labeling effi-

ciency was similar to the IgA preparation. min01), and whiskey and LD diet rats (0.072 { 0.015)


110 AMORE ET AL.

FIG. 1. Typical chromatoscanner profile of the test probe showing a labeling efficiency of 91.5%. Experimental conditions were runspeed 12 cm/min (abscissa) and time constant 0.5 sec. In ordinate, counts per minute (cpm) 1 105.

AID Clin 4357 / a512$$4357 07-02-97 23:17:20 clinal AP: Clin


FIG. 2. Composite scintigraphic images of rats injected with 123I-labeled aggregated IgA. Rat 3, given whiskey, was administered asmaller amount of radioactivity and was not included in the study. The other animals included a rat given whiskey (No. 1), a rat given LDdiet (No. 2), and a normal control (No. 4). Rat abdominal anatomy is also shown.

were less negative and t1/2 more prolonged (mean 5.4, blood stream) and a values (expression of liver uptake)(Fig. 4). Detailed data are shown in Table 1.7.9, and 9.5 min, respectively) compared to normal

(0.286 { 0.177 min01, correspondent to a mean t1/2 of In terms of urinary abnormalities, hematuria waspresent in all but one rat maintained on the LD diet2.4 min, P õ 0.05 for each study group). Despite the

lack of statistical significance of the differences of a (//////), in 5 of 6 rats given whiskey (////), andin all 3 rats given both whiskey and LD diet (//////).values among the groups, a significant correlation

(P õ 0.01) was found between l values (representing Hematuria was undetectable in controls. Proteinuriawas significantly higher in each experimental groupthe rate of disappearance of the test probe from the


112 AMORE ET AL.

FIG. 4. Relationship between a values (expression of liver up-take) and l values (expression of the rate of disappearance of theaggregated IgA labeled with 123I from the circulation).

in organs where free iodine could accumulate (tyrod,gut), or in bladder.

Renal Injury

The hepatotoxic effects of the described regimenshave been extensively described elsewhere (10). Immu-

TABLE 1

Liver Uptake and Catabolism and Blood Clearance Rateof IgA Aggregates in Rats

FIG. 3. Typical time–activity curves obtained by simultaneousrecording of activity of liver (right lobe) and heart after injection No. MTT a lof the aggregated IgA labeled with 123I. Compared to normal (top),abnormal hepatic dynamic graphs can be clearly appreciated in the 1 151 0.196 NDrat given whiskey (middle) and especially in the rat given LD diet 2 150 ND 0.066(bottom). 3 100 0.233 0.088

4 99 0.599 0.3615 88 0.248 0.1166 101 0.198 0.1347 189 0.123 0.094(whiskey, 6.90 { 1.41 mg/day; LD diet, 9.1 { 0.64 mg/8 196 0.157 0.074day; whiskey and LD diet, 9.5{ 0.58 mg/day) compared9 235 0.489 0.014to normal (0.82 { 0.17 mg/day, P õ 0.03 for each

10 312 0.228 0.183group). The multiple regression analysis showed that 11 216 0.116 0.085MTT values, proteinuria and hematuria were signifi- 12 183 0.128 0.071

13 219 0.150 0.098cantly correlated (r2 Å 0.78, P õ 0.01).14 211 0.148 0.072Table 2 shows results of g function analysis and15 262 0.083 0.047blood clearance rate in rats given whiskey or LD diet 16 99 0.657 0.644

injected with aggregated IgG. Again a values were 17 80 0.136 0.13418 98 0.226 0.189similar in study groups and normals, while MTT were19 78 0.149 0.136more prolonged in rats given whiskey or LD diet and20 74 0.400 0.261l values less negative in rats given LD diet than con-21 85 0.367 0.289

trols (ANOVA for three groups P õ 0.005 and P õ 22 76 ND 0.3520.05, respectively).

Note. Rats 1–6 were given whiskey, rats 7–12 were nourishedAlpha, MTT, and l values obtained following injec-with LD diet, rats 13–15 were given whiskey and nourished with LDtion of radiolabeled aggregated IgG were similar to val-diet, rats 16–22 were normal animals. MTT, expressed in minutes, isues obtained by injecting aggregated IgA. the mean transit time in the liver; a is the exponent of the classic

Finally, no significant amount of radioactivity (i.e., formulation of the function (see text) regulating the liver uptake; gvalues are expressed in min01. ND, not determined.greater than circulation) was detected during the study



TABLE 2 Light microscopy analysis revealed focal signs of mes-angial proliferation and sclerosis in 2 of 4 cases. LightLiver Uptake and Catabolism and Blood Clearance Rateand electron microscopy examination and immunoflu-of IgG Aggregates in Ratsorescence did not reveal obvious morphologic changes

No. MTT a l in all normal rats with the exception of a case showingfocal signs of glomerulosclerosis.

23 105 19 0.13224 138 35 0.211

DISCUSSION25 143 27 0.18826 225 10 0.06727 229 22 0.176 In the present study, liver uptake and catabolism of28 192 17 0.094 a uniform preparation of heat-aggregated immunoglob-29 87 27 0.278

ulin A were studied in four groups of rats, one receiving30 94 28 0.19331 72 45 0.321 intragastric infusion of commercial whiskey, a second

nourished with an LD diet, a third given both whiskeyNote. Rats 23–25 were given whiskey, rats 26–28 were given LD and LD diet, and a fourth group of animals nourished

diet, rats 29–31 were normal animals.with regular chow.

The combination of LD and alcohol intake was pre-viously found to promote hepatic and renal changes,leading to hepatocellular injury and a secondary formnofluorescence examination showed IgA localized in

the mesangial area corresponding to electron-dense de- of IgA nephropathy (10). The present study focused onthe possible role of an impaired clearance of IgA andposits (1/) in 8 of 9 rats given whiskey and focal mesan-

gial proliferation at the light microscopy in 2. A greater IgA-immune complexes in the pathogenesis of the renalinjury and based on the possibility to apply establishedextent (2/) of mesangial electron-dense deposits to-

gether with positive IgA immunofluorescence (Fig. 5) techniques of analysis of kinetics and fate of an im-munecomplex-like test probe (15, 16) in a well-charac-were evidenced in the 12 rats given LD diet with or

without whiskey examined. Weak mesangial IgG de- terized experimental model (10).An aggregated IgA was labeled with 123I and tracerposits were found in 5 of 9 rats given LD diet, 2 of 3

given LD diet and whiskey, and 3 of 9 given whiskey. kinetics was followed by means of a large field-of-view

FIG. 5. Immunofluorescence micrograph showing representative deposits of IgA in glomeruli from rats maintained on a LD diet. 1400.


114 AMORE ET AL.

2. Abramowsky, C. R., Dahms, B., and Swinehart, G., IgA associ-digital gamma camera equipped with an ultra-high 4-ated glomerular deposits in liver disease. Hum. Pathol. 16,mm resolution collimator. The liver was found to be the1243–1246, 1985.main organ involved in the handling of the aggregate.

3. Bene, M. C., De Korwin, J. D., Hurault De Ligny, B. H., Aymard,Computer-generated time–activity graphs of liver B., Kessler, M., and Faure, G. C., IgA nephropathy and chronicwere analyzed by a mathematical model (16), which liver cirrhosis Am. J. Clin. Pathol. 89, 769–773, 1988.allowed an indirect evaluation of the slope of liver accu- 4. Berger, J., Yaneva, H., and Nabarra, B., Glomerular changes in

patients with cirrhosis of the liver. Adv. Nephrol. 7, 3–14, 1978.mulation (representing tracer uptake) and calculation5. Borghi, M., and Pecchini, F., IgA nephropathy preceding liverof liver transit time (expression of tracer catabolism).

cyrrhosis. Clin. Nephrol. 24, 213–217, 1985.It was shown that LD diet with or without a heavy6. Crowson, C. N., and More, R. H., Glomerulotubular nephrosisalcohol intake and even the alcohol intake alone altered

correlated with hepatic lesions. Incidence and morphology of as-the liver handling of radiolabeled aggregated IgA as-sociated kidney and liver lesions in human autopsy material.

sumed to mimic circulating IgA-immune complexes. Arch. Pathol. 60, 73–84, 1955.These data were consonant with the results of a recent 7. Kalsi, J., Delacroix, D. L., and Hodgson, H. J. F., IgA in alcoholicreport on human cirrhosis (16), showing that removal cirrhosis. Clin. Exp. Immunol. 52, 499–504, 1983.of IgA-containing complexes was impaired. Notably, 8. Nochy, D., and Druet, P., Les glomerulopathies au cours des

maladies hepatiques. Nov. Press. Med. 8, 1595–1599, 1979.liver uptake and catabolism of the aggregated IgA in9. Woodroffe, A. J., IgA glomerulonephritis and liver disease. Aust.rats closely resembled those of the IgA–IgG aggregate

N.Z. J. Med. 11, 109–111, 1981.in patients with liver cirrhosis (16).10. Amore, A., Coppo, R., Roccatello, D., Piccoli, G., Mazzucco, G.,A similar defect in the handling of an aggregated

Gomez-Charri, M., Lamm, M. E., and Emancipator, S. N., Exper-IgG was found as well. This observation was consonant imental IgA nephropathy secondary to hepatocellular injury in-with the immunofluorescence findings of IgG codepos- duced by dietary deficiencies and heavy alcohol intake. Lab. In-ited with IgA in a few cases but indicated that addi- vest. 70, 68, 1994.tional factors were likely to be involved in the promi- 11. Messner, R. P., Meryhew, N. L., and DeMaster, E. G., Selective,

prolonged alteration of complement-mediated immune clearancenent deposition of IgA in this experimental model.after acute exposure of mice to ethanol. Clin. Immunol. Immuno-As previously shown (10) LD diet was more effectivepathol. 70, 73, 1994.than whiskey in inducing fatty changes in the liver.

12. Ruiz, P., Gomez, F., and Schreiber, A. D., Impaired function ofCorrespondingly, the hepatic sequestration was found macrophage Fc receptors in end-stage renal disease. N. Engl. J.to be greatly impaired in LD diet rats. Damaged cells Med. 322, 717, 1990.could express a reduced number of receptors (17). 13. Roccatello, D., Picciotto, G., Coppo, R., Piccoli, G., Molino, A.,

Cacace, G., Amore, A., Amoroso, A., Quattrocchio, G., and Sena,Other defects possibly relate to the mechanisms of in-L. M., Clearance of polymeric IgA aggregates in humans. Am. J.tracellular transport and lysosomal trafficking, criticalKidney Dis. 14, 354, 1989.for both recycling of the receptor (18) and IgA catabo-

14. Rifai, A., Schena, F. P., Montinaro, V., Mele, M., D’Addabbo,lism. Variables such as hepatic blood flow can be con-A., Nitti, L., and Pezzullo, J. C., Clearance kinetics and fate of

sidered, but are less likely. macromolecular IgA in patients with IgA nephropathy. Lab. In-While these results confirmed, in the rat model, a vest. 61, 381, 1989.

recent report in cirrhosis patients (16), it is noteworthy 15. Roccatello, D., Picciotto, G., Ropolo, R., Coppo, R., Quattrocchio,G., Cacace, G., Molino, A., Amoroso, A., Baccega, M., Isidoro, C.,that the liver damage induced in the experimental ratsCardosi, R., Sena, L. M., and Piccoli, G., Kinetics and fate ofwas moderate, suggesting that even mild hepatocellu-IgA–IgG aggregates as a model of naturally occurring immunelar injuries were able to induce an early derangement complexes in IgA nephropathy. Lab. Invest. 66, 86, 1992.

of the immune clearance of Ig aggregates. 16. Roccatello, D., Picciotto, G., Torchio, M., Ropolo, R., Ferro, M., Fran-ceschini, R., Quattrocchio, G., Cacace, G., Coppo, R., Sena, L. M.,

ACKNOWLEDGMENTS De Filippi, P. G., and Piccoli, G., Removal systems of immunoglobu-lin A and immunoglobulin A containing complexes in IgA nephrop-

This study was supported by grants from Ministero dell’Universita athy and cirrhosis patients. Lab. Invest. 69, 714, 1993.e della Ricerca Scientifica e Tecnologica (60%). 17. Morell, A. G., Gregoriadis, G., Scheinberg, I. H., Hickman, J., and

Ashwell, G., The role of sialic acid in determining the survival ofglycoproteins in the circulation. J. Biol. Chem. 246, 1461, 1971.REFERENCES

18. Perez, J. H., Branch, W. J., Smith, L., Mullock, B. H., and Luzio,J. P., Investigation of endosomal compartments involved in en-1. Abramowsky, C. R., and Christiansen, D. M., Secretory immuno-

globulin deposits in children with extrahepatic biliary atresia: docytosis and transcytosis of polymeric immunoglobulin A bysubcellular fractionation of perfused isolated rat liver. Biochem.Studies in a human counterpart of experimental ligation of the

bile ducts. Hum. Pathol. 18, 1126–1131, 1987. J. 251, 763–770, 1988.

Received August 29, 1995; accepted with revision February 25, 1997


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Processing of IgA Aggregates in a Rat Model of Chronic Liver Disease