Enhanced Expression of the CD71 Mesangial IgA1 Receptor in Berger Disease and Henoch-Schönlein Nephritis: Association between CD71 Expression and IgA Deposits
Elie Haddad*,,
Ivan C. Moura,
Michelle Arcos-Fajardo,
Marie-Alice Macher*,
Veronique Baudouin*,
Corinne Alberti,
Chantal Loirat*,
Renato C. Monteiro and
Michel Peuchmaur
*Service de Nephrologie, Service de Sante Publique, EA 3102, Université Paris 7 and Service de Pathologie, Hôpital Robert Debre, Assistance Publique-Hôpitaux de Paris, Paris, France; and INSERM E0225, Faculte de Medecine Xavier Bichat, Université Paris 7, Paris, France.
Correspondence to Dr. Elie Haddad, Service de Nephrologie, AP/HP, Hôpital Robert Debre, 48 Bd Serurier, 75019 Paris, France. Phone: 33-1-40-03-24-67; Fax: 33-1-40-03-24-68;
ABSTRACT. IgA nephropathy (IgA-N) that comprises Berger diseaseand Henoch-Schönlein Purpura (HSP) nephritis is definedby mesangial IgA deposits. Recently, this group has characterizeda new receptor for IgA, the transferrin receptor (CD71), expressedon mesangial cells. To assess whether CD71 was involved in thepathogenesis of IgA-N, its expression was analyzed togetherwith IgA deposits on 16 kidney biopsies from 16 patients withBerger disease (n = 4) or HSP (n = 12). These biopsies werecompared with 17 kidney biopsies of a group of 15 patients (controlgroup) with other glomerulonephritis, including systemic lupuserythematosus, poststreptococcal acute glomerulonephritis, membranoproliferativeglomerulonephritis, steroid-sensitive minimal change nephroticsyndrome, steroid-resistant idiopathic nephrotic syndrome withfocal and segmental glomerulosclerosis, and persistent and isolatedproteinuria with minimal change on kidney biopsy. In this controlgroup, IgA deposits could be observed in eight kidney biopsiesof seven patients. These biopsies were also compared with normalkidney specimens (normal group). In normal kidney, it was foundthat CD71 was linearly expressed on tubular epithelium but waseither not expressed or very dimly in glomeruli. In contrast,CD71 was strongly expressed in 105 of the 107 glomeruli of thekidney biopsies from the IgA-N group. For the control group,it was found that expression of CD71 in glomeruli was correlatedto the presence of IgA deposits. Indeed, among the 87 glomeruliof nine kidney biopsies (eight patients) without IgA fixation,78 exhibited no CD71 expression and nine exhibited a very dimone. On the other hand, all 49 glomeruli of the eight kidneybiopsies (seven patients) in which IgA deposits were detectedexhibited CD71 expression (P < 10-4). Performance of dual-labelingstudies with confocal microscopy on kidney biopsies of IgA-Npatients demonstrated that most of the IgA deposits co-localizedwith CD71. It was also demonstrated that the intensity of theexpression of CD71 was not linked to the intensity of clinicalor biologic findings but to the intensity of cellular proliferationin both IgA-N and control groups. These results show that mesangialCD71 expression is not specific to IgA-N. However, the associationbetween IgA deposits and CD71 expression and their co-localizationin the mesangium provide strong evidence that CD71 is a majorIgA receptor on mesangial cells. Email: elie.haddad@rdb.ap-hop-paris.fr
Henoch-Schönlein purpura (HSP) nephritis and Berger diseaseare both considered as IgA nephropathy (IgA-N). Berger disease,first described by Berger and Hinglais (1), is a mesangial proliferativeglomerulonephritis defined by mesangial deposition of IgA. HSPis a systemic IgA-mediated vasculitis that usually associatesskin lesions and abdominal and joint pains (2). About 30% ofthe HSP patients present with glomerular lesions (3–7)that are characterized, like in Berger disease, by mesangialdeposition of IgA. It has been suggested that Berger diseaseand HSP are pathogenetically linked and that Berger diseasecould be a kidney-restricted form of HSP. Indeed, kidney lesionsare indistinguishable (8), and, in both disorders, many patientshave increased serum IgA levels (9–11) and IgA containingcirculating immune complexes (CIC) (11,12). Whatever the mechanism(s)involved in the pathogenesis of these two diseases, the finalcommon pathway of Berger disease and HSP is represented by mesangialIgA deposition. Although it has been established that IgA couldbind specifically to mesangial cells and induce proliferationand cytokine production (13), the identity of the receptor(s)of IgA on mesangial cells remains unknown. Indeed, none of thefour IgA receptors that have been identified in humans havebeen shown to be expressed on mesangial cells (13–15).These include the IgA Fc receptor (FcRI; CD89) expressed onmyeloid cells that binds IgA1 and IgA2 (16–19), the polymeric-Igreceptor (pIgR) on mucosal epithelial cells (20) that bindspolymeric IgA and IgM, the Fc/µR on B cells and macrophagesthat also binds IgA and IgM (21), and the asialoglycoproteinreceptor (ASGP-R) on hepatocytes that can bind glycoproteins,including IgA1 and IgA2 through terminal galactose or N-acetylgalactosamineon O-linked and some N-linked glycans. Recently, our group hascharacterized a new receptor for IgA1 that, in contrast to FcRI,is not expressed on leukocytes but on mesangial cells (22).This receptor, identified as the transferrin receptor (TfR orCD71), binds IgA1 but not IgA2 (22). We proposed that this receptorcould have IgA1 as a ligand in IgA-N and showed that this receptorwas expressed in the glomeruli of three Berger disease patients(22). To assess whether this mesangial IgA1 receptor was involvedin the fixation of IgA on mesangial cells and whether it wasspecific of IgA-N, we analyzed its expression on kidney biopsiesof 16 children with Berger disease or HSP in comparison withkidney biopsies of 15 children with other glomerular diseasesand with normal kidney specimens from children with nephroblastoma.We also investigated whether the expression of CD71 in IgA-Ncould be correlated to the severity of the nephritis as definedby clinical, biologic, or histopathologic features.
Patients and Tissue Samples
We retrospectively analyzed 16 kidney biopsies of 16 patientswith HSP (n = 12) or Berger disease (n = 4). This group wascalled IgA-N group. These biopsies were compared with 17 kidneybiopsies of 15 patients with systemic lupus erythematosus (n= 6), poststreptococcal acute glomerulonephritis (n = 3), typeI membranoproliferative glomerulonephritis (n = 1), steroid-sensitiveidiopathic minimal change nephrotic syndrome (n = 1), steroid-resistantidiopathic nephrotic syndrome with focal and segmental glomerulosclerosis(n = 1), and persistent and isolated proteinuria with minimalchange lesions (n = 3). This second group was called controlgroup. For these two groups, renal tissue samples were obtainedby percutaneous needle biopsy taken on clinical indicationsfrom children who were treated during the years 1995 to 2001in the Nephrology Unit of Hopital Robert Debre, Paris, France.We also retrospectively analyzed five kidney samples from nephrectomyspecimens from children with nephroblastoma. Sections were performedfar away from the tumoral tissue, and the histopathologic analysisof these sections was considered normal. Each section containedhundreds of glomeruli, but only 50 glomeruli per patient werecounted and analyzed because the aspect was homogenous. Thisgroup was called normal group.
Diagnosis of HSP relied on the association of joint pain, abdominalpain, and purpura with normal platelet counts. All patientswith Berger disease had repeated macroscopic hematuria. In allcases, the diagnosis was confirmed by kidney biopsy showingthe presence of mesangial IgA deposits. The main clinical andbiologic characteristics of these 16 patients at the time ofkidney biopsy are given in Table 1. Diagnoses in control grouppatients were based on typical clinical and biologic findingsand on kidney histology on which light microscopy and immunofluorescencestaining for immunoglobulins and complement components had beenperformed.
Table 1. Clinical and biologic features of patients with Berger disease or HSP nephritis at the time of kidney biopsya
All biopsies were re-analyzed by conventional light microscopy.Patients from IgA-N group were graded according to the classificationof the International Study of Kidney Disease in Childhood (ISKDC)modified by Heaton et al. (23) for HSP nephritis and accordingto the classification proposed by Haas (24) for Berger disease.
In all cases, frozen biopsies were available. These frozen sectionswere analyzed for IgA deposition and CD71 expression. In allcases, glomeruli available on frozen samples were quantifiedand analyzed and compared with glomeruli available on lightmicroscopy.
Immunohistochemical Studies
In each case, one fragment of the renal biopsy was immediatelyfrozen in liquid nitrogen, embedded in Tissue-Teck O.C.T Coump(Miles Scientific, Elkhart, IN) and stored at -80°C afterdiagnostic process was completed.
Immunohistochemistry was performed on serial frozen sectionsto analyze both the IgA deposition and the CD71 expression.
Immunostaining for IgA was performed using a direct immunofluorescencetechnique with an anti-human IgA isothiocyanate fluorescein(FITC)–conjugated antibody (anti-human IgA-FITC, ref F0204;DakoCytomation, France). Briefly, 5-µm cryosections weretransferred onto poly-L-Lysine (Sigma Chemical Co., La Verpillère,France)–coated slides, dried for 60 min at room temperature,fixed in alcool-ether 1:1 for 20 min and then dehydrated inalcool 95°C for 10 min. After rinsing 3 x 5 min in PBS,sections were incubated with anti-IgA FITC-conjugated antibodydiluted 1:10 in PBS for 30 min at room temperature in darkness.The slides were rinsed 3 x 5 min in PBS and mounted in hydrophilicfluorescein medium (Biosys, Compiègne, France). The slideswere examined under epifluorescence with an Aristoplan Leitzmicroscope.
Immunostaining for CD71 was performed using a three-stage immunoperoxidasetechnique with a mouse IgG2a anti-human CD71 monoclonal antibody(clone M-A712; BD PharMingen, Le Pont de Claix, France). Allrinsing and incubations were performed in 0.05 ml/L Tris-HClat pH 7.6 (Tris) at room temperature. Briefly, 5-µm cryosectionswere transferred onto poly-L-Lysine coated slides, dried for60 min at room temperature, fixed in alcool-ether 1:1 for 20min, and then rehydrated in alcool 95°C for 10 min. Afterrinsing 3 x 5 min in Tris, sections were incubated with anti-CD71MoAb diluted to 1:50 (final concentration, 10 µg/ml) for60 min. After rinsing, they were incubated with 1:50 peroxidase-conjugatedsecondary antibody (rabbit anti-mouse IgG, ref P0260; DakoCytomation,France) for 30 min and then with 1:50 peroxidase-conjugatedtertiary antibody (pig anti-rabbit IgG, ref P0217; DakoCytomation,France) for 30 min. Immunoreactive sites were visualized byuse of the Supersensitive kit (Biogenex HK1535K, San Ramon,CA) according to manufacturer’s protocol. Omission ofthe primary antibody and use of an irrelevant MoAb with thesame isotype served as controls.
Co-Localization of CD71 and IgA by Confocal Microscopy
Briefly, each frozen section was incubated with a mouse IgG2aanti-human CD71 monoclonal antibody (1:50 for 1 h), rinsed inTris, and then incubated with an anti-mouse Texas red–conjugatedgoat antibody (1:100 for 1 h), rinsed in Tris, and finally incubatedwith an anti-IgA FITC-conjugated rabbit polyclonal antibody(1:10, 30 min). Omission of the anti-CD71 MoAb was used as anegative control (no Texas red fluorescence). Sections wereexamined with a ZEISS LSM 510 laser scan microscope using ax63 objective with two lasers (argon and helium-neon). The Texasred fluorophore was visualized with the 543-nm excitation wavelenghof a helium-neon laser and FITC with the 488-nm excitation wavelengthof an argon laser.
Quantification of CD71 Expression and Grading of Histologic Findings
The IgA staining was defined as absent or present. The intensityof CD71 expression was classified as absent, low (grade 1),moderate (grade 2), or high (grade 3). This semi-quantitativegrading scale used for CD71 expression was based on the surfaceof glomerular tuft area staining positively for CD71 (see Figures 1 and 2).The glomeruli of frozen samples were assigned as normal,increased mesangial matrix (mesangiopathic glomeruli), or proliferative(endocapillary or extracapillary proliferation).
Figure 1. CD71 expression on normal kidney. (A) Frozen section of the cortex exhibiting a CD71 expression in tubule but no CD71 (grade 0) expression in the mesangial area of glomeruli. (B) Frozen section of the cortex exhibiting a very dim (grade 1) CD71 expression in the mesangial area of glomeruli. Original magnifications: x400 in A; x640 in B.
Figure 2. CD71 expression on IgA nephropathy (IgA-N) kidney biopsy. (A) A mesangiopathic glomerulus (increased mesangial matrix) in IgA-N with a grade 2 CD71 expression. (B) A proliferative glomerulus with a grade 3 CD71 expression. Original magnification, x640.
Figure 3. Co-localization of IgA and CD71 in the mesangium of an IgA-N patient. Samples were analyzed by confocal microscopy. Squares represent focal planes of a floculus in mesangium. Sections were stained with anti-IgA FITC antibody (A) and anti-CD71 Texas red antibody (B) (see Materials and Methods). As shown by the arrow, IgA and CD71 co-localize in the mesangium as reflected by the yellow signal of the merge composite (C).
Figure 4. Kidney biopsy from an IgA-N patient (patient 7) exhibiting various types of glomerular lesions. A representative mesangiopathic glomerulus (increased mesangial matrix) on formalin-fixed section stained with periodic acid-Schiff (PAS)(A) and a representative mesangiopathic glomerulus on frozen section exhibiting a grade 2 CD71 expression (B). A representative glomerulus with pure mesangial proliferation on formalin fixed section stained with PAS (C) and a representative glomerulus with pure mesangial proliferation on frozen section exhibiting a grade 3 CD71 expression (right). Inserts: mesangial IgA deposition on frozen section by immunofluorescence technique. Original magnification, x400.
Statistical Analyses
Data were expressed as frequency. Comparisons used 2 test. Alltests were two-tailed, and P < 0.05 was considered statisticallysignificant. All statistical analyses were performed using SAS8.0 (SAS Inc, Cary, NC) software package for PC computer.
Association between IgA Deposits and CD71 Expression
In the normal kidney, CD71 was found to be expressed on tubularepithelium (Figure 1). In the glomeruli (n = 250), CD71 expressionwas either not detected (grade 0, Figure 1A) or found with avery dim expression (grade 1, Figure 1B). As expected, no IgAdeposits were found in these glomeruli.
In the 16 kidney biopsies of the 16 patients from IgA-N group,CD71 was markedly expressed in the mesangial area in all glomeruli(n = 107). As shown in Table 2, among these 107 glomeruli, CD71expression was high (grade 3) in 51 glomeruli (Figure 2B), moderate(grade 2) in 54 (Figure 2A), and low (grade 1) in only 2. Moreover,mesangial CD71 expression was similar in Henoch-Schönleinnephritis and in Berger disease. As expected, mesangial IgAdeposits were found in all 107 glomeruli of the IgA-N group.
Table 2. Association between IgA deposits and CD71 expression in glomerulia
The expression of CD71 and the presence of IgA deposits werealso tested in the 17 kidney biopsies (136 glomeruli) of the15 patients of the control group. In this group, no IgA depositscould be found in nine kidney biopsies of eight patients (87glomeruli), whereas IgA deposits were found in eight kidneybiopsies of seven patients (49 glomeruli). Review of the medicalrecords of these seven patients with IgA deposits on kidneybiopsy confirmed that none of them could be considered as anatypical presentation of IgA nephropathy. These seven patientswith IgA deposition in control group suffered from systemiclupus erythematosus (n = 5), acute poststreptococcal glomerulonephritis(n = 1), and steroid-resistant idiopathic nephrotic syndromewith focal and segmental glomerulosclerosis (n = 1). The eightother patients from the control group who had no IgA depositson kidney biopsy suffered from systemic lupus erythematosus(n = 1), acute poststreptococcal glomerulonephritis (n = 2),type I membranoproliferative glomerulonephritis (n = 1), steroid-sensitiveminimal change nephrotic syndrome (n = 1), and persistent andisolated proteinuria with minimal change lesions (n = 3). Asshown on Table 2, the expression of CD71 in glomeruli of patientsfrom the control group was strongly associated to the presenceof IgA deposits. Indeed, among the 87 glomeruli without IgAdeposits, 78 exhibited no CD71 expression (grade 0) and 9 exhibiteda low one (grade 1). On the other hand, all 49 glomeruli inwhich IgA deposits were detected exhibited CD71 expression thatranged from grade 2 to grade 3, demonstrating an associationbetween the presence of IgA deposits and a grade 2 or 3 CD71expression (P < 10-4, 2 test). Altogether, among the 156glomeruli with IgA deposits (107 from the IgA-N group; 49 fromthe control group), 154 displayed a moderate or high expressionof CD71 and 2 a low one, whereas among the 87 glomeruli withoutIgA deposits (all of them being from the control group), 78had no CD71 expression and 9 a low one (P < 10-4, 2 test).
Co-Localization of CD71 and IgA in Glomeruli
Five kidney biopsies of IgA-N patients were used for IF analysisby confocal microscopy. Seventeen glomeruli were examined andshowed the same findings. As shown on Figure 3, most of theIgA deposits were co-localized with CD71 in mesangial regionsof glomeruli. This co-localization was observed on each focalplan performed by the confocal microscope, thereby ruling outthe hypothesis that this co-localization was related to an overlappingof the different plans.
Association between CD71 Expression and Severity of the Kidney Disease
To determine whether the intensity of CD71 expression was correlatedwith the severity of the disease in the IgA-N group, level ofCD71 staining was compared with clinical and biologic findingsat the time of biopsy such as serum IgA level, hematuria, proteinuria,renal insufficiency, and arterial hypertension. None of thesecriteria correlated with the intensity of CD71 expression. However,this kind of association was very difficult to assess becauseof the small number of patients and because CD71 expressionwas homogeneous in only 9 of the 16 patients of the IgA-N group,whereas it was very heterogeneous in 7. Indeed, for these sevenpatients, some glomeruli exhibited a high level of CD71 expressionand other glomeruli exhibited a moderate level (Table 3). Wetherefore tested whether the level of CD71 expression was linkedto histopathologic findings. For this purpose, glomeruli onfrozen sections were classified as normal or as showing increasedmesangial matrix or showing proliferative lesions. Histologicanalysis is usually difficult to perform on frozen sections;therefore, data from frozen biopsies were compared with dataobtained on light microscopy. Findings were similar betweenfrozen and fixed sections, except for patient 12 for whom histologicanalysis on fixed sections showed a focal proliferative glomerulonephritiswith less than 50% crescents (grade IIIa according to the classificationof the ISKDC modified by Heaton et al. [23]), whereas all glomeruliobserved on frozen sections displayed proliferative lesions.The observation of a slight discrepancy between fixed and frozensections in only one patient allowed the comparison of the CD71expression and the histologic analysis of frozen sections, glomerulusby glomerulus. By performing this comparison, we found thatthe level of CD71 expression was strongly associated with theintensity of proliferation. Indeed, as shown on Table 3, amongthe 107 glomeruli studied in the IgA-N group, 60 glomeruli showedonly an increased mesangial matrix and 47 showed proliferativelesions, including seven with crescents. Among the 60 glomeruliwith increased mesangial matrix, two glomeruli displayed a grade1 CD71 expression, 54 displayed a grade 2 (Figure 2A), and onlyfour displayed a grade 3. On the other hand, all 47 glomeruliwith proliferative lesions displayed a grade 3 CD71 expression(Figure 2B), demonstrating that the presence of proliferativelesions was associated with a grade 3 CD71 expression (P <10-4, 2 test). Moreover, the seven glomeruli with crescent displayedan enhancement of CD71 expression within the crescents. Withinthese crescents, CD71+ cells were mostly epithelial cells (notshown). As shown on Figure 4, patient 7 is representative ofthe heterogeneity of CD71 expression and of its linkage to theintensity of proliferation. Indeed, among the seven glomeruliavailable on frozen sections in this patient, four glomeruliwith increased mesangial matrix exhibited a low or a grade 2CD71 expression (Figures 4A and 4B) and the three glomeruliwith proliferative lesions exhibited a grade 3 CD71 expression(Figure 4C and 4D).
Table 3. Association between CD71 expression in glomeruli and mesangial proliferation in patients from IgA-N groupa
We also tested whether this association between CD71 expressionand the intensity of proliferation was observed in the 49 glomeruliof the eight kidney biopsies of the control group in which IgAdeposits were detected. As shown in Table 4, among the 23 glomeruliwith proliferative lesions, 16 exhibited a grade 3 CD71 expressionand seven a grade 2. On the other hand, the 24 glomeruli withminimal change lesions and the two glomeruli with segmentalglomerulosclerosis all exhibited a grade 2 CD71 expression,demonstrating that proliferative lesions were associated toa higher frequency of grade 3 CD71 expression as compared withminimal change lesions (P < 10-4, 2 test). Among the 87 glomeruliof the control group without IgA deposits, 18 glomeruli displayedproliferative lesions, including three with crescents (Table 5),and 69 were normal or showed minimal change lesions. Amongthe 18 glomeruli with proliferative lesions, CD71 expressionwas low (grade 1) in nine and was not detected in nine, whereasnone of the 69 glomeruli that were normal or with minimal changelesions were found to express CD71. This suggests that, despitea slight reinforcement of CD71 expression, proliferative lesionsin the absence of IgA deposits do not result in an enhancementof CD71 expression sufficient to make it detectable at a higherlevel than that observed in the normal group.
Table 5. Association between CD71 expression in glomeruli and mesangial proliferation in patients from control group with no IgA deposits on glomerulia
The identification of a mesangial receptor for IgA is a crucialstep for the understanding of the pathogenesis of IgA-N. Wecould show in a previous study that the transferrin receptor(TfR/CD71), known to bind the transferrin itself (25) and tobind the hemochromatosis protein (26), was also an IgA1 receptorexpressed on cultured mesangial cells (22). Moreover, we hadshown in this study that the mesangial cell expression of CD71was enhanced in the glomeruli of three patients with Bergerdisease. To test whether this enhancement of CD71 expressionwas specific of IgA-N, we analyzed the expression of CD71 inkidney biopsies of children with Berger disease or HSP nephritisand compared it with that of children with other glomerulardisease and with that of normal kidney. In normal kidney, CD71was undetectable or expressed at a very low grade in glomeruli.In all 16 IgA-N patients studied, the expression of CD71 inglomeruli was enhanced as compared with that observed in glomeruliof normal kidney, confirming our first results in three adultpatients with Berger disease (22). However, we found that CD71expression in glomeruli was also enhanced in other glomerulonephritis,demonstrating that this expression was not specific of IgA-N.Nevertheless, we could show that CD71 expression was linkedto the presence of IgA deposits (P < 10-4) and that the intensityof CD71 expression was strongly associated with the intensityof cellular proliferation (P < 10-4). Considering this secondobservation, one could hypothesize that CD71 expression is notcorrelated to the IgA deposition but is simply a marker of mesangialproliferation, which is theoretically possible because CD71is known to be a marker of proliferation in other cellular models(27–34). Several observations make the hypothesis thatCD71 is only linked to cellular proliferation very unlikely.First, among the eight patients with other glomerular diseasesfor whom kidney biopsies showed no IgA deposition, one patientsuffered from a grade IVA systemic lupus erythematosus glomerulonephritiswith endocapillary proliferation, two from a post-streptococcalacute glomerulonephritis, and one from a type I membranoproliferativeglomerulonephritis. Despite the proliferative lesions that wereobserved in all 18 glomeruli of these four patients, CD71 expressionwas absent in nine and low (grade 1) in nine. Second, in patients17, 22, and 23 from the control group with IgA deposits, grade2 CD71 expression was observed in 26 of their 33 glomeruli,but none of these 26 glomeruli displayed proliferative lesions.Third, in the IgA-N group, grade 2 CD71 expression was alsoobserved in the glomeruli without any proliferative lesions.All these observations strongly suggest that IgA depositionby itself is associated with an enhancement of CD71 expressionand that CD71 expression is first correlated with IgA deposits.Moreover, we could demonstrate, by performing dual-labelingstudies with confocal microscopy, that most of the IgA depositsdid co-localize with CD71 in the mesangium. This observationis the first one, to our knowledge, showing a co-localizationof IgA deposits with a presumed receptor on mesangial cells.The fact that this co-localization is observed on kidney biopsiesof patients with IgA deposits is much more relevant than ifit was observed on cultured mesangial cells. Altogether, theassociation of IgA deposits and CD71 expression and the demonstrationof co-localization of IgA and CD71 strongly suggests that CD71is a major receptor for IgA1 in mesangium. Interestingly, wecould show on confocal microscopy that most of the IgA moleculesbut not all of them co-localized with CD71. The hypothesis ofanother IgA receptor cannot therefore be ruled out.
Two hypotheses could account for the fact that IgA depositionis linked to an enhancement of CD71 expression. The first hypothesisis that the primary event is an overexpression of CD71 by anunknown mechanism and that this overexpression induces a fixationof IgA. This hypothesis could account for the association ofIgA deposition and CD71 expression in glomerular diseases otherthan IgA-N but is very unlikely in the setting of IgA-N. Indeed,most of the recent studies suggest that the primary event inIgA-N is an abnormal IgA glycosylation. It has been demonstratedthat serum IgA1 display an abnormal O-glycosylation in patientswith IgA-N (35–37) or HSP (38–41) and that thisabnormal glycosylation is also found in IgA deposits on mesangialcells (42). Moreover, it has been shown that abnormal IgA glycosylationis restricted to HSP patients with nephritis while it was notobserved in HSP patients without nephritis (39). In addition,it has recently been shown that abnormally glycosylated IgAisolated from IgA-N patients significantly increased the apoptoticrate and nitric oxide synthesis activity of cultured mesangialcells, in comparison with IgA fractions isolated from controls(43). Finally, another important result arguing for a primaryevent due to abnormal IgA is the relapse of IgA deposits ontransplanted kidney in patients with IgA-N (44–47). Thesecond hypothesis is that the primary event is a fixation ofIgA on CD71 and that this fixation induces an upregulation ofCD71 expression on the cell surface either by a redistributionof cytoplasmic CD71 or by an induction of CD71 synthesis bythe cells. This upregulation of CD71 would then make the expressionof CD71 detectable by our immunoperoxidase staining; in "normal"glomeruli, this technique would not be sensitive enough to detectlow levels of CD71 expression. This upregulation of CD71 expressioncould then lead to a dramatic increase of the fixation of IgAon mesangial cells. This increase of IgA deposition could thenbe pathogenic or not, depending on the normality or abnormalityof IgA. In the setting of IgA-N, deposition of abnormally O-glycosylatedIgA would be pathogenic; in the context of other glomerulonephritis,the deposition of normally O-glycosylated IgA on mesangial cellsmay or may not be pathogenic.
We did not find a significant correlation between the intensityof IgA staining and the intensity of CD71 expression (data notshown). This is not surprising because a linear correlationbetween the extent of IgA deposits and the CD71 expression wasnot expected. Indeed, it is known that IgA deposits are composedof IgA-containing complexes of various sizes and composition(e.g., degree of O-glycosylation, anionic charge and multimerizationof IgA) (42,48–51). This introduces a number of independentparameters that can vary from patient to patient and may affectnot only the binding of IgA to CD71 and the ability of IgA complexesto activate mesangial cells, but also the staining of IgA byimmunoperoxidase or immunofluorescence.
We did not find any association between the intensity of CD71expression and clinical or biologic findings. However, thiskind of association cannot be ruled out because of the smallnumber of patients and because of the heterogeneity of CD71expression in 7 of the 16 patients. However, we found that theintensity of CD71 expression was strongly associated with thepresence of proliferative lesions. Moreover, we could show thatthis association was observed within each glomerulus taken individuallyin a same patient. Therefore, CD71 expression could be a goodmarker of proliferation in IgA-N. It has been shown that thereis a fairly good relationship between the clinical pattern atonset and the renal outcome in children with HSP nephritis (3,5,52–55),but it is well known that the best predictors of renal outcomeare the histologic lesions observed on renal biopsy (52,53,55–57).Indeed, the percentage of glomerular crescents is the majorprognosis factor; involvement of more than 50% of glomeruliis well recognized to be associated with a poor outcome in children(52,53,55–57). In adults, the proportion of crescentsis also a major prognosis factor, but the discriminating pointmay be lower than 50% (58–61). In the context of Bergerdisease, it has been shown that renal histologic lesions areof good prognostic value in children (62,63) and in adults (24,64,65).Indeed, diffuse proliferative glomerulonephritis, the extentof global glomerular sclerosis, interstitial sclerosis, andcrescents were identified as markers of poor prognosis (24,65).Therefore, it could be interesting to determine in a prospectivestudy whether the intensity of CD71 expression could be correlatedwith a poor outcome. Indeed, the fact that CD71 expression islinked to the presence of IgA deposition and that the IgA fixationon CD71 is supposed to play a role in the proliferation of mesangialcells support the hypothesis that CD71 expression could be usedas a more specific tool than other histologic lesions.
These results suggest that CD71 is probably the main receptormediating IgA deposition on mesangial cells in both types ofIgA-N, Berger disease, and HSP and that the study of the interactionof IgA and CD71 and its consequences on mesangial cells deservefurther study for a better understanding of the pathogenic mechanismsof IgA-N.
Acknowledgments
We are very grateful to Cecile Pouzet, PhD, Faculté deMédecine Xavier Bichat, Université Paris 7, fortechnical assistance in confocal microscopy.
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Received for publication July 9, 2002.
Accepted for publication October 15, 2002.
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Abstract
Introduction
RI; CD89) expressed on myeloid cells that binds IgA1 and IgA2 (16–19), the polymeric-Ig receptor (pIgR) on mucosal epithelial cells (20) that binds polymeric IgA and IgM, the Fc
2 test. All tests were two-tailed, and P < 0.05 was considered statistically significant. All statistical analyses were performed using SAS 8.0 (SAS Inc, Cary, NC) software package for PC computer. 
