1 Departments of Internal Medicine and
2 Clinical Chemistry, Tampere University Hospital and
3 Medical School, University of Tampere, Tampere, Finland
Correspondence to:
M. Pertovaara, Department of Internal Medicine, Section of Rheumatology, Tampere University Hospital, PO Box 2000, FIN-33521 Tampere, Finland.
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Abstract |
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Methods.Urinary total protein excretion from 24 h urine collection, as well as urinary excretion rates of albumin, alpha-1 microglobulin (1m) and IgG from overnight 8 h collections, were determined from 78 pSS patients (75 females, three males). Urine acidification capacity after oral ammonium chloride load was tested in 55 of these patients.
Results.Mild proteinuria (0.150.42 g/24 h) was observed in 34 patients (44%). Increased urinary excretion rates of albumin (20 µg/min),
1m (
7.0 µg/min) or IgG (
5.0 µg/min) were detected in nine (12%), nine (12%) and 11 patients (14%), respectively. Latent or overt distal renal tubular acidosis (dRTA) was observed in 18 out of 55 patients with pSS (33%). These patients had a longer duration of the disease (10±4 vs 8±4 yr; P
0.05); they also had proteinuria (67 vs 27%; P
0.025) and hypertension (44 vs 14%; P
0.05) more frequently, and significantly higher serum creatinine (92±39 vs 78±13 µmol/l; P
0.025) and serum beta-2 microglobulin (ß2m) levels (3.3±1.6 g/l vs 2.6±0.6 g/l; P
0.025) as compared to patients with normal urine acidification capacity.
Conclusions.Inadequate renal acidification capacity, as well as mild proteinuria, were frequently found in patients with pSS. Those with dRTA had longer disease duration, a higher level of serum ß2m, and they had proteinuria and hypertension more frequently than those with normal renal acidification capacity.
KEY WORDS: Renal involvement, Sjögren's syndrome, Renal tubular acidosis, Proteinuria
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Introduction |
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Somewhat contradictory figures have been reported regarding the prevalence of renal involvement in patients with pSS, the frequency varying between 2 and 67% in different studies [3, 6, 9, 10, 1418]. These studies, however, involved relatively small groups of patients, patient selection varied, or both pSS and secondary SS (sSS) patients were combined; it is thus possible that the renal lesions encountered could be associated with the underlying connective tissue disease.
We investigated the occurrence of renal involvement, especially that of dRTA and proteinuria, in a fairly large and clearly defined group of patients with pSS.
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Patients and methods |
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Methods
The clinical examination included a thoroughgoing interview of the patients covering family history, previous diseases, previous and concurrent medications, drug allergies, duration of sicca symptoms, first manifestation of the disease, existence of recurrent parotid or submandibular gland swellings, and present sicca symptoms of the eyes and mouth. Any history of upper and lower urinary tract infections and possible renal stones was recorded. The existence of hypertension was defined as previous or current use of antihypertensive drugs. Special emphasis was placed on possible extraglandular symptoms of SS (endocrine, dermatological, gastrointestinal, neurological, lymphoproliferative, musculoskeletal, renal, respiratory and vascular symptoms).
In urinalysis, specimens were screened by dipstick (Combur-10M Test, as read by Miditron Instruments, Roche, Mannheim, Germany), and in cases proving positive for erythrocytes, leucocytes, protein or nitrite, microscopy of the urinary sediment was performed according to the Finnish standardization with Sternheimer stain, where one x400 high-power field (HPF) equals 0.12 µl original urine volume [22]. The healthy upper reference limits were: for women, 1 erythrocyte/HPF, 2 leucocytes/HPF, 2 squamous epithelial cells/HPF; for men, 1 erythrocyte and 1 leucocyte/HPF, and no epithelial cells.
A 24 h urine collection was conducted and creatinine clearance (enzymatic method, Vitros, Johnson & Johnson, Clinical Diagnostics, Rochester, NY, USA), excretion of urinary total proteins (pyrogallol red method, Olli-C, Kone Instruments, Finland) and immunoglobulin light chains (Behring Nephelometric Analyser, Marburg, Germany) were determined. Proteinuria was defined as urinary total protein excretion 0.15 g/24 h. Urinary excretion of albumin, IgG and
1m was measured from samples obtained in timed (from 10 p.m. to 6 a.m.) overnight urine collection (cU) and analysed by nephelometry (Behring Nephelometric Analyser, Marburg, Germany). Urinary excretion of albumin
20 µg/min, of
1m
7.0 µg/min and of IgG
5.0 µg/min was used to discriminate increased excretion. Lysozyme was also determined from an 8 h overnight urine collection (cU) and measured using Micrococcus lysodeicticus as standard.
An ammonium chloride loading test was applied as a short-duration test [23] provided there was no known contraindication. After waking at 6.30 a.m., the patients voided, and at 7.00 a.m. they drank 200400 ml of water. At 8.00 a.m., a test meal of ammonium chloride 0.1 g/kg body weight was given in enteric-coated capsels. The test continued until 2.00 p.m. with water supplement ~100 ml/h. Urine samples were taken at the outset and at 1 h intervals up to 6 h after the test meal. Blood samples were drawn immediately, after 0.5 h and thereafter at 1 h intervals up to 5.5 h after the test meal. The voided urine volumes were recorded and samples for urine pH measurement taken in a closed syringe and pH measured with a blood gas analyser (Radiometer ABL 500 Analyser, Copenhagen, Denmark) within 15 min from sample collection. The samples for determinations of urine titratable acids and ammonium ions were stored in a refrigerator and analysed at the latest on the following day, or stored at -20°C and measured within 1 month from collection. The heparinized capillary blood samples were stored in an ice bath and analysed within 1 h (Radiometer ABL 500 Analyser, Copenhagen, Denmark). Inability to acidify the urine in maximal acidosis (latent dRTA) was defined according to the reference values related to the degree of acidosis achieved in the acid loading test, as described by Backman et al. [23].
The concentrations of serum creatinine, sodium, potassium and chloride were determined by RA-1000 analysers (Technicon/Bayer, Elkhart, IN, USA). Other standard laboratory tests included basic blood cell count, erythrocyte sedimentation rate (ESR), serum C-reactive protein (CRP), total protein, alanine aminotransferase and alkaline phosphatase. Rheumatoid factor (RF) was determined by laser nephelometry (Behring Nephelometric Analyser). Antinuclear antibodies (ANA) were determined by indirect immunofluorescence using Hep-2 cells. Antibodies to extractable nuclear antigens (ENA), including anti-ribonucleoprotein (RNP), anti-Sm, anti-SS-A, anti-SS-B and anti-Scl70 antibodies, as well as antibodies to native DNA (QUANTA Lite ds DNA, INOVA Diagnostics Inc., San Diego, CA, USA), were measured by enzyme immunoassay. Anti-salivary gland antibodies were analysed by indirect immunofluorescence (monkey salivary gland slide, INOVA Diagnostics Inc.). Serum concentrations of IgA, IgG and IgM, as well as serum complement levels (C3 and C4), were measured by laser nephelometry (Behring Nephelometric Analyser). Serum beta-2 microglobulin (ß2m) was determined by radioimmunoassay (Pharmacia beta-2-micro RIA kit, Pharmacia Diagnostics, Uppsala, Sweden), the healthy reference interval being 1.02.5 mg/l.
Renal imaging was performed with ultrasonography and with radionuclides using 99mTc-MAG (mercaptoacetyltriglycine).
The study protocol was approved by the ethical committee of Tampere University Hospital.
Statistical analysis
Statistical analysis was performed by Student's t-test, and by 2 test with Yates' correction with the Statview program. Logistic regression analysis with SPSS 6.1 microversion was applied in backward stepwise manner to define independent effects of intercorrelated variables on the occurrence of RTA and proteinuria in pSS patients.
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Results |
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Urinalysis and urinary protein excretion
In urinalysis, the dipstick for albumin was positive in three out of 78 patients (4%). The dipstick for erythrocytes was positive in 13 patients (17%); in the subsequent sediment examination, >1 erythrocyte/HPF was found in four patients. The dipstick for leucocytes was positive in 21 patients (27%); in subsequent urine microscopy, >2 leucocytes/HPF were observed in 12 patients. The culture for urine bacteria was positive in 10 (13%) cases (Table 1). Only two patients had symptoms of urinary tract infection; in other cases, the bacteriuria was asymptomatic. Since no patient had an upper urinary tract infection, eventual proteinuria encountered in these subjects is hardly explained by the asymptomatic bacteriuria.
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Renal function and imaging
The mean serum creatinine concentration in the patients was 84±15 µmol/l (mean±S.D .). In nine out of 78 patients (12%), the level of serum creatinine was increased (100 µmol/l in females and
115 µmol/l in males), three of them having levels >150 µmol/l (153, 161 and 223 µmol/l). Seven out of the nine patients with elevated serum creatinine levels were aged
65 yr. The mean value for creatinine clearance was 1.27±0.43 ml/s/1.73 m2 (the lower reference limit for healthy women being 0.94 ml/s/1.73 m2 and that for healthy men 1.32 ml/s/1.73 m2 ). Creatinine clearance was decreased in 15 (19%) of the patients, all of whom were >65 yr of age. The mean serum sodium concentration was 142±3 mmol/l, serum potassium 4.0±0.3 mmol/l and serum chloride 105±3 mmol/l.
In renal ultrasound, one patient showed unilateral hydronephrosis and, correspondingly, a severe unilateral deterioration of renal function in nephrography. Otherwise, nephropathic changes and parenchymal scars were each encountered in five patients (6%). Mild changes in renal ultrasound were encountered in 20 patients (26%) altogether. In nephrography, 26 patients out of 78 (33%) had some abnormalities; the findings were usually mild, but moderate functional changes were occasionally observed (5%) (Table 1).
Ammonium chloride loading test
Sixty-three patients with pSS agreed to participate in an ammonium chloride loading test. In nine of them, the test failed due to nausea or vomiting. The acidification capacity was abnormal in 17 of the 54 patients with diagnostic tests (Table 2), of whom one had overt dRTA, this also being encountered in one additional patient. The prevalence of dRTA in the 55 patients was thus 33% (Table 1
).
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Comparison of patients with inadequate to those with normal urine acidification ability
Primary SS patients with dRTA and those with normal urinary acidification capacity did not differ in mean age, whereas there were significant differences between the patient groups in the duration of xerostomia, the duration of the disease from diagnosis and the presence of hypertension (Table 3). There were no statistically significant differences between them in previous or current use of non-steroidal anti-inflammatory drugs, corticosteroids or disease-modifying anti-rheumatic drugs. The mean levels of blood haemoglobin, serum leucocytes, serum thrombocytes, serum CRP or blood ESR did not differ between the groups. There was a significant difference between them in the mean level of serum creatinine, but not in mean creatinine clearance (Table 3
). Mild proteinuria (0.150.42 g/24 h) was detected more often in the patients with dRTA than in those with normal acidification capacity. However, there were no differences in the occurrence of increased urinary excretion rates of albumin,
1m or IgG between the patient groups. No patients had lysozyme in the urine. No statistically significant differences were seen in the occurrence of RF, ANA, SS-A or SS-B antibodies, nor in the concentrations of serum immunoglobulins IgA, IgG and IgM (Table 3
). Anti-DNA antibodies were detected in only one patient in each group. The levels of serum ß2m were significantly higher in the patients with compromised acidification compared to those with normal acidification capacity (Table 3
).
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Comparison of patients with proteinuria to those with normal urinary protein excretion
The duration of xerostomia was significantly longer in pSS patients with mild proteinuria (0.150.42 g/24 h) than in those with normal urine protein excretion, but there were no statistically significant differences in the duration of sicca symptoms of the eyes nor in the duration of the disease from the year of diagnosis (Table 4). The frequency of hypertension did not differ between the groups, but both systolic and diastolic blood pressures measured at the study examination were significantly higher in those with proteinuria (Table 4
). The acid loading test results were more often abnormal in those with mild proteinuria than in those with normal urinary protein excretion (52% vs 18%; P
0.02). There were no significant differences between the groups in the levels of blood haemoglobin, serum leucocytes, serum thrombocytes or blood ESR, but serum CRP was higher in those with proteinuria as compared to those with normal urinary protein excretion (8.5±6.0 vs 6.6±3.3 mg/l; P
0.05). No differences were seen between the proteinuric and non-proteinuric patients in the occurrence of RF, ANA, SS-A or SS-B antibodies nor in the concentrations of serum immunoglobulins. Anti-DNA antibodies were detected in one proteinuric patient and in five patients with no proteinuria; no one of them fulfilled more than four of the 11 criteria for SLE [24]. Anti-Sm antibodies were detected in negligible titres in two proteinuric patients and four patients with normal urinary protein excretion. The levels of serum ß2m were significantly higher in those with proteinuria compared to those with normal urinary protein excretion (Table 4
).
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Discussion |
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In previous studies, the overall occurrence of renal involvement in pSS has varied from 2 to 67% [3, 6, 9, 10, 1418]. The patient groups investigated have been rather small, some studies have been retrospective [3 in part, 15, 17], patient selection has varied, and the earliest studies concerned both patients with pSS and sSS [6, 10, 15, 16], although in some of these latter the results of pSS and sSS patients were analysed and described separately. Furthermore, there are several case reports in the literature describing severe events of hypokalaemic periodic paralysis [2527] or even respiratory arrest in patients with dRTA due to SS [28].
In the earliest studies on this issue, Bloch et al. [15] investigated SS patients retrospectively as regards renal abnormalities, and found hypostenuria either transiently or persistently in 10 out of 62 patients (16%). According to the current classification, only 23 of the 62 patients were pSS patients and all four patients with persistent hypostenuria belonged to this subgroup, giving a frequency of persistent hypostenuria in pSS of 17%.
In the original work by Talal et al. [6], impairment in urinary acidification was found in 50% of a study group comprising 10 primary and two sSS patients. In a large retrospective SS study (n=171) [16] also involving both primary and sSS patients, renal manifestations were, however, investigated only in 15 patients. Ammonium chloride loading test results were abnormal in a total of six patients; an inadequate acidification capacity was observed in one of the four pSS patients (25%) [16]. In a retrospective study of 47 patients with pSS, signs of renal abnormalities (repeatedly high urine pH, low serum bicarbonate or elevated serum creatinine concentration) were found during the initial evaluation in 12 patients (26%) and at follow-up in 38% [17]. Ten renal biopsies had been performed, of which six showed tubulointerstitial nephritis, three focal glomerulonephritis and one both of these lesions [17].
Our results on the frequency of defects in acidification capacity in pSS are in fair accordance with most previous findings. In a study by Shiozawa et al. [10], abnormalities in the ammonium chloride loading test were found in 35%, but the patient group constituted only 17 patients; moreover, there were both pSS and sSS patients. In another study [11], 36 patients with pSS were investigated and abnormalities in renal tubular acidification capacity were found in 33% of the 15 patients tested. Pokorny et al. [9] studied 65 patients with pSS and found an overt kidney manifestation (either complete RTA or gross proteinuria) in 12 of them (18%). Ammonium chloride loading test was applied in 25 cases, 12 (48%) yielding abnormal results [9]. In a more recent study, Siamopoulos et al. [12] found abnormal acidification test results in 33% of the 21 patients studied.
Somewhat different figures for the prevalence of renal involvement were obtained in a retrospective survey by Vitali et al. [3] in which signs of renal involvement investigated retrospectively by blood urea nitrogen and serum creatinine, creatinine clearance and urinalysis were found in only 2% of 104 pSS patients. The authors also conducted a prospective study on 20 randomly selected patients with pSS and found abnormal acidification capacity in 15%. Also, in a recent prospective study, abnormalities in ammonium chloride loading test were found in only 12% of the 27 pSS patients studied [18].
The highest prevalence figures for acidification defects are those from Eriksson et al. [14]: they found abnormal results in the ammonium chloride loading test in 67% of 27 patients with pSS. However, their patient group was not randomly selected, involving also patients already known to have either dRTA or urolithiasis.
Here, the duration of the disease, as well as the duration of xerostomia, were found to be associated with the occurrence of overt or latent renal acidification defects. This is in accord with the results of Shiozawa et al. [10] who found the duration of the disease in SS patients with renal tubular acidification defect to be longer than in those with normal acidification capacity. On the other hand, Pokorny et al. [9] found that disease duration in the group with renal involvement was shorter than in those with normal renal findings. In both of these studies [9, 10], the patients with renal acidification defects were younger than those with normal tubular acidification, whereas in the present study there were no differences in age between the patient groups.
In our study, serum ß2m was found to be significantly higher in patients with latent or overt dRTA than in those with normal acidification capacity. This high level of serum ß2m might indicate more extensive lymphocyte infiltration, and thus reflect the severity of SS. A high level of serum ß2m has also previously been associated with renal findings in SS [29]. Contrasting results were reported by Viergever et al. [18], who found no significant correlation of serum ß2m with abnormal tubular tests.
Hypergammaglobulinaemia has previously been associated with dRTA both in SS and in other conditions [1, 6, 30, 31]. However, in a study where urinary acidification was tested in rheumatoid arthritis patients with and without hypergammaglobulinaemia, it was shown that hypergammaglobulinaemia per se does not cause impairment of renal acidification [32]. In several subsequent studies concerning renal involvement in pSS, no association has been found between serum gammaglobulin concentration and compromised renal acidification [3, 9, 12, 18]. In the present case, no significant differences in serum IgG concentrations were found between those with dRTA and those with normal acidification capacity, although there was a tendency of serum IgG levels to be higher in the dRTA group. However, in the group with dRTA, systemic corticosteroids had been used either previously or concurrently more frequently than in those with normal acidification capacity (67% vs 46%). Although this difference was not statistically significant, the use of corticosteroids may conceivably have influenced the serum IgG concentrations.
Anti-SSA and anti-SSB antibodies have been associated with systemic manifestations of SS [33]. In the present study, no differences in the occurrence of anti-SSA or anti-SSB antibodies between patients with normal and compromised renal acidification capacity were found; similar findings regarding their association with renal abnormalities have been reported by others [9, 12]. In our patients, the overall occurrence of both anti-SSA and anti-SSB antibodies was higher than in other series [3335]. This is probably due to a sensitive ELISA method; on the other hand, as anti-SSB antibodies have been found to be specific and predictive of SS [3536], their frequent occurrence also reflects the fact that patients with sSS have been excluded from our study.
Previously, proteinuria with excretion rates of >0.5 g/day has been found in <3% of SS patients [13]. In our study, mild proteinuria was detected often (44%), but in keeping with previous results, none had proteinuria 0.5 g/24 h, although two (3%) had previously suffered from proteinuria >1 g/24 h due to either vasculitis or interstitial nephritis requiring treatment with either corticosteroids or cyclophosphamide. We found that the duration of xerostomia was longer, the levels of systolic and diastolic blood pressure were higher, abnormalities in urine acidification occurred more frequently and the levels of serum ß2m were higher in pSS patients with proteinuria compared to those without. To our knowledge, such comparisons in respect of proteinuria in pSS have not previously been presented. Furthermore, it was noteworthy that in the subgroup of pSS patients whose acidification capacity was evaluated, the presence of proteinuria was significantly associated with RTA in a logistic regression model. Thus, in patients with pSS, mild proteinuria might indirectly imply the possibility of a defect in renal acidification capacity.
We found signs of tubular proteinuria, detected by urinary 1m, relatively rarely in pSS patients (12%), which conflicts somewhat with the fact that abnormalities in renal acidification were much more frequent. However, increased urinary
1m levels have been related particularly to proximal tubular disorders rather than to distal tubular disorders [37], whereas abnormalities in renal acidification indicate distal tubular dysfunction. Higher figures for tubular proteinuria have recently been reported in a study where increased urinary ß2m excretion (an indicator of proximal tubular dysfunction) was present in 26% of patients, and altogether various signs of tubular dysfunction were found in 16 (59%) of the 27 pSS patients investigated by acidification and thirsting test [18]. In another recent study, increased excretion of urinary
1m was observed in up to 46% of the 24 pSS patients [14]; however, as previously noted, the patients were not randomly selected. Shiozawa et al. [10] found increased urinary ß2m excretion in four out of 11 patients (36%).
In conclusion, latent dRTA and mild proteinuria were frequent findings in pSS patients in this study. Patients with renal acidification defects had longer disease duration and more frequent occurrence of hypertension and proteinuria than those with normal acidification capacity. Also, in a logistic regression model, particularly hypertension and proteinuria, as well as the duration of xerostomia, were shown to be associated with RTA. The pSS patients with either RTA or proteinuria had higher serum ß2m concentrations than those with normal acidification capacity and normal urinary protein excretion, which would imply that the renal manifestation is a consequence of prolonged active lymphoproliferation. All the patients with clinically significant renal disease related to SS had been identified prior to this study; the significance of the frequent subclinical renal findings can only be judged by further follow-up of these patients.
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Acknowledgments |
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References |
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