Membranous
THEODORE
Nephropathy following Volatile Hydrocarbons’
EHRENREICH,
L. YUSIS,
STUART
Received
April
Exposure
.-\ND
JACOB
to
CHVRG?
14, 1976
Excessive exposure to solvents has long been known to cause renal tubular disease in man t 1, 2). Recently, the occurrence of glomerular disease such as Goodpasture’s syndrome with proliferative and extracapillary glomerulonephritis has been related to hydrocarbon exposure (3-5). This report concerns four patients with membranous nephropathy who had a history of prolonged exposure to a number of volatile hydrocarbons. Membranous nephropathy is a chronic renal disease involving glomeruli and occurring principally in adults (61. Its clinical onset is insidious and is manifested by proteinuria or edema. Proteinuria is the hallmark of the disease and may be present for many years without symptoms. It is often a slowly progressive disease. About one-fourth of the patients improve clinically and lose their proteinuria, while up to one-fourth develop renal failure. The glomerular capillary wall lesions are distinctive, contain deposits of immunoglobulin and complement considered to represent immune-complexes, and show a morphological progression from early Stage I to late Stage IV (6). While in most cases there is no known etiology, in some patients the disease has been deemed to be secondary to specific diseases or agents such as infections, neoplasms, or chemicals (7, 8). The four cases to be described fall into this last category. METHODS
AND PATIENTS
In the course of investigating a series of patients with membranous nephropathy an occupational exposure to solvents was recorded in three cases. In another case solvent exposure was suspected because of the patient’s occupation (plastics worker). Subsequently, a detailed occupational history was obtained in all four cases. The diagnosis of membranous nephropathy in all patients was made on the basis of an initial kidney biopsy as well as the clinical history. Subsequently, additional kidney biopsies were performed. In Case 1 there was one additional biopsy and nephrectomy, and two additional biopsies in the fourth case. One patient died and an autopsy was performed. ’ Supported in part by USPHS Research Grant AM-00918 from the National Institute of Arthritis and Metabolic Diseases. 2 Reprint requests: Jacob Churg. M.D.. Renal Pathology Division, Department of Pathology. Mt. Sinai School of Medicine of CUNY. 5th Avenue and 100th Street, New York. New York 10079.
36
EHRENREICH,
YUNIS,
AND
CHURG
For light microscopy, the kidney biopsies and autopsy kidney tissue were fixed in 10% formalin with 2% sodium acetate by weight resulting in a pH close to neutral. Tissues were embedded in paraffm and 2-3 pm sections were cut. The following stains were performed: hematoxylin and eosin (H & E), periodic acid Schiff(PAS), periodic acid silver methenamine (PASM), chromotrope aniline blue (CAB), (9) and chromotrope silver methenamine (CSM) (10). For electron microscopy tissue was fixed in 1% osmium tetraoxide in phosphate buffer and embedded in epon. Grids were prepared, double stained with uranyl acetate and lead hydroxide, and examined in an RCA 1llG electron microscope at 50 or 100 kv. REPORT
OF CASES
Case 1 In 1968 proteinuria was discovered on routine physical examination of this 23-year-old male. From 1966 to 1968 he had worked in a photographic film research laboratory and was exposed to organic solvents. At the time of this initial employment, and before being exposed to solvents, his urine was found to be free of protein. He continued to be employed in the same film laboratory and 1 year later was transferred to another film making process in which he was not exposed to organic solvents. Proteinuria persisted and he was admitted to the hospital (1969). The physical examination was essentially negative. The blood pressure was 120/70, the blood urea nitrogen (BUN) and the intravenous pyelogram were normal. Urine showed 2.5 g of protein in 24 hours. Renal biopsy revealed early membranous nephropathy. Subsequently the patient developed edema and was treated with prednisone and furosemide which were ineffective and were discontinued. Another kidney biopsy was performed and showed more advanced membranous nephropathy (1971). In 1972 he developed renal failure with hypertension and was dialyzed. This was followed by bilateral nephrectomy for hypertension and later a transplant. E,rposure Histoq The patient worked in a photographic film research laboratory and his occupation involved a heat process of film making on open equipment using a variety of organic solvents which the patient reported as including acetone, acetonitrile, butyl acetate, butyl alcohol, carbon tetrachloride, chloroform, cyclohexane, dichloroethane, dichloromethane, dimethyloformamide, isopropyl alcohol, methyl alcohol, methyl ethylketone, toluene, tetrahydrofuran, trichloroethylene, and xylene. Some of these solvents were used for the cleanup of equipment and of the patient’s hands. Case 2 In 1964 at the age of 45 the patient was discovered to have 4+ proteinuria in the course of a routine physical examination. At that time there was no peripheral edema, the BUN, serum albumin, serum globulin, and serum cholesterol was normal. A kidney biopsy was performed and showed advanced membranous nephropathy. He continued to work without treatment other than restriction of fluids. The proteinuria persisted without edema. In February 1973, he was found to have elevated BUN, hypertensiona, and persistent proteinuria. The blood plsessure was
MEMBRANOUS
NEPHROPATHY
37
1801124, the fundi showed blurring and narrowing without hemorrhage. He was subsequently treated with methyldopa and chlorothiazide. At this time the urine showed 1.9 g of protein a day, BUN was 53 to 70 mg/lOO ml, serum creatinine 5.2 to 6.0 mg/ 100 ml, creatinine clearance 9 cm3 per minute. He was in renal failure, but it was considered to be stable and no further treatment was undertaken. In February 1974, he had a myocardial infarct without complications, from which he recovered. One year later he was placed on chronic hemodialysis. Exposure
History
This patient worked in the plastics industry from 1946 to 1969 with a variety of materials used in the heated condition for moulding. According to the patient, he worked first with Plexiglas moulded into brush handles and also with polyvinyl acetate in toy moulding. This period covered 2 years. A further 5 to 7 years were spent heat-moulding polyvinyl chloride. From 1964 to 1969 the patient was employed in the heat-moulding of polyvinyl chloride pipe. Since the patient reported being exposed to fumes of formaldehyde it is likely that phenoplasts were manipulated at some stage in his career as these plastics break down on heating and give off formaldehyde. It is a necessary step in moulded plastics production to clean moulding equipment with organic solvents and the patient confirmed that he used a number of organic solvents for cleanup at the end of production shifts. Apart from the foregoing it may be noted that phenoplasts may decompose into phenol, benzene, cresol, and other aromatic hydrocarbons (11). Polyesters disintegrate into acids and alcohols. Polyvinyl chlorides may give off some unreacted vinyl chloride monomer in heating and decompose to give substantial amounts of hydrogen chloride ( 12). One may conclude that the patient had skin and respiratory exposure to solvents; in addition respiratory exposure to at least formaldehyde and hydrogen chloride. Case 3 A male patient was well until 1956 when at the age of 37 he developed nephrotic syndrome which was treated with steroids and diuretics for 1 year with complete remission. At this time all drugs were stopped. In 1966 at the age of 47 there was recurrence of the nephrotic syndrome. In 1967 a renal biopsy was performed and revealed advanced membranous nephropathy. He was treated with steroids with improvement of the nephrotic syndrome. For a number of years he has been on a salt-free diet with diuretics. His present renal status is that of mild proteinuria (1-2 g/day), slightly elevated blood pressure, and a normal BUN. E-x-posure
History
He worked from 1936 in a gyroscope assembly plant. For about 15 years his job involved cleaning parts in an open tank of perchlorethylene. Around 1952 the patient was placed on a precision-type assembly and this involved cementing parts together with an epoxy preparation. First the parts were cleaned with Freon-l 12. If malfunction occurred in the assembled unit, it was taken apart and the cement softened with a “stripping” compound claimed to have a strong smell. Then the parts were cleaned with various solvents including acetone, methylethylketone, perchlorethylene, and trichlorethylene. Following the development ofsymptoms of
38
EHRENREICH,
YL’NIS,
AND
CHURG
kidney disease in 1956, the patient did not work for several months, but then resumed his previous work up to 1967. At this time his work associate, patient 4, developed the same symptoms. Since other workers showed no abnormalities on urinalysis, the symptoms were not deemed to be occupationally induced. However, at this time he was promoted to a “leader” in the same department, a position in which there was considerably less general exposure and no direct contact with cleaning solvents. In 1971 after the death of his associate, patient 4, he was transferred out of assembly to a job without solvent exposure. Case 4 This male patient was in good health until April 1967 when at the age of 44 he developed a typical nephrotic syndrome. Renal biopsy showed early membranous nephropathy. He was treated with steroids and diuretics; this was followed by complications, such as pulmonary embolism, for which anticoagulants were administered. A year later he developed hypertension (blood pressure 170/l 10) and azotemia. At this point he was treated with azathioprine and a second biopsy showed more advanced membranous nephropathy. In February 1971, at the age of 48, he was admitted to the hospital with uremia, grand ma1 seizures, and respiratory difficulty. He was treated with antibiotics, tracheostomy for the respiratory difficulty, received peritoneal dialysis, went into coma and died in severe acidosis. E.xposure
History
He was employed from 1956 in the same gyroscope assembly plant as patient 3. For 11 years he worked side by side with patient 3 sharing his work. He therefore had a similar exposure to various solvents such as acetone, methylethylketone, perchlorethylene, and trichlorethylene. Following the development of nephrotic syndrome in 1967 he had no further solvent exposure. RESULTS Pathological
Obser\?ntions
In Case 1 there was typical progression of membranous changes starting with early lesions (Stage I) in the initial biopsy (Fig. 1). through Stages II and III in a subsequent biopsy and terminating in end-stage kidney in the nephrectomy specimen (Fig. 7). In the first biopsy electron microscopy was not available and on light microscopy the glomeruli appeared to be normal with H & E and PAS stains. However, with CSM stain, small, rounded deposits could be discerned on the outer or subepithelial aspect of the basement membrane. The deposits stained red with chromotrope indicating their proteinaceous nature, and were somewhat more numerous than is generally seen in this early stage. There were no other glomerular, tubular or vascular changes. In the second biopsy the lesions had progressed to the full blown Stage II (as illustrated from another case in Figs. 2 and 3). The deposits were larger, more numerous and separated by thin projections (“spikes”) of the basement membrane both on light and electron microscopy. There was fusion (or loss) of foot processes. In some glomeruli the lesions had progressed to the advanced State III (as illustrated in Figs. 4 and 5). Here the projections encircled the deposits, some of which were incorporated into a thickened basement membrane. In addition there was widening of the mesangium in some glomerular tufts resulting
MEMBRANOUS
FIG. (arrows)
1. Early Stage I, initial of the thin basement
39
KEPHROPATHY
biopsy in Case I. The only change membrane, due to deposits. CSM
in this glomerulus x450.
is focal
FIG. 2. Full-blown Stage 11, Case 4. The basement membrane is thin and there thickening (arrow). Deposits form a thick layer on the outer aspect ofthe basement by thin spikes. CSM x 450.
is minimal membrane
thickening
mesangial separated
40
EHRENREICH,
YUNIS,
AND
CHURG
FIG. 3. Electron microscopy of the biopsy shown in Fig. 2. Numerous subepithelial electron-dense deposits (D) on the outer aspect of the basement membrane (BM) are separated by spikes (arrow). There is loss (fusion) of foot processes. x7290.
FIG. 4. Advanced Stage III, Case 2. The basement increase in mesangium. CSM x270.
membrane
is markedly
thickened.
There
is no
MEMRRANOUS
FIG. 5. Electron micrograph in advanced thin spikes (arrow) and are incorporated others pale. x6173.
NEPHROPATHY
Stage 111. Case 3. The deposits are completely into the basement membrane. Some deposits
41
encircled by are granular.
in partial sclerosis associated with intracapillary hyaline deposits. Focal tubular atrophy was present. In the nephrectomy specimen (Fig. 71 most of the glomeruli were obsolete or obsolescent with adhesions and fibrous crescents. The membranous change could still be discerned in some glomeruli and others were relatively unaffected showing only slight sclerosis. There was widespread tubular atrophy with cast formation and interstitial inflammation. Case 2 (Figs. 4 and 61 had one biopsy with typical advanced Stage III lesions. Case 3 (Fig. 5) also had one biopsy showing both Stage II and Stage III glomerular lesions with little or no increase in mesangium. In Case 4 (Figs. 2 and 3) there were three biopsies all showing Stage II lesions with some progression to Stage III in the last biopsy. Sclerosis, absent in the first biopsy, was present in segments of some glomeruli in subsequent biopsies. Glomerular obsolescence and focal tubular atrophy progressed in a similar manner. At autopsy the membranous change persisted with varying degrees of glomerular sclerosis, obsolescence, and collapse; there was widespread tubular atrophy with many casts and interstitial inflammation. The clinical and pathological findings in all four cases are summarized in Table I. COMMENTS
It is proposed that there may be a causal relationship between solvent exposure and membranous nephropathy in the cases presented. Of the four patients, three
42
EHRENREICH,
YUNIS,
AND
CHURG
micrograph in advanced sclerosis, Case 2. There is ingrowth of mesangial matrix into FIG. 6. Electron the capillary loop and the basement membrane (arrows) which shows advanced membranous change with dense. pale, and granular deposits. x6173.
had a long exposure to a number of known solvents for 2, 11, and 20 years, respectively, before developing symptoms. Two of these patients (Cases 3 and 4) had the same quality of exposure under the same working conditions for different periods. The third patient (Case 2) was exposed for 18 years to solvents generated in the course of heating plastics and two solvents used for cleaning equipment. While specific solvents could not be identified in this case, it is known that there is solvent exposure in the plastics industry. In none of the instances were personal protective measures taken. The morphological stage of membranous nephropathy has been related to the duration of the disease which is believed to begin with the early Stage I (6). In the cases presented, the biopsy following the shortest exposure, 2 years in Case 1, showed early lesions, whereas the lesions were more advanced in the other cases with exposures of 11, 18, and 20 years, respectively. It is not known whether the progression of the glomerular lesions is related to the duration of the exposure or of the disease. Of the four patients, three developed renal failure (two were treated). The fourth, who was treated, has had good renal function with persistent proteinuria 7 years after the onset of symptoms. This may be considered a good response to treatment since he has lost his nephrotic syndrome and the disease is apparently arrested, although not cured. For comparison, in a large series of cases of treated and
MEMBRANOUS
43
NEPHROPATHY
FIG. 7. End stage kidney disease in Case I. Marked glomerular sclerosis atrophy. In oneglomerulus without sclerosis (arrow) membranous nephropathy thick-walled capillaries. PAS x 108.
and widespread tubular is present in the patent.
nontreated idiopathic membranous nephropathy the occurrence of renal failure was 25% (7) If this proportion of renal failure (75%) will be found in larger groups of exposed subjects, it would suggest that solvent exposure results in a more severe form of membranous nephropathy. The pathogenesis of membranous nephropathy following toxic chemicals such as solvents is uncertain. In Goodpasture’s syndrome related to hydrocarbon exposure it was postulated that the inhaled toxic chemicals affected the pulmonary alveoli
Exposure onset
before
Symptoms
at
Initial
Patient
symptoms
1
7 years
onset Proteinuria
Age
Exposure
24
3 years
7
18 years
Proteinuria
4.5
IX years
3
20 years
Nephrotic syndrome
47
30 years
Nephrotic syndrome
44
4
biopsy
of
11 years
I I years
Outcome
Stage of M.N. Early Stage I and
II
Advanced Stage III Full-blown advanced and III Full-blown II
and Stage Stage
II
Renal failure 3 years after biopsy Renal failure 9 years after biopsy Improved with persistent proteinuria Renal failure I year after biopsy: died
Treatment Prednisone
Diuretics
Prednisone
Prednisone Azathioprine
44
EHRENREICH,
YUNIS,
AND
CHURG
giving rise to antibodies which then damaged the glomeruli (3). In the cases of solvent-induced membranous nephropathy there was no history of pulmonary disease, nor were any lung changes found at autopsy. Another mechanism has been described which may be pertinent to these cases. In idiopathic membranous nephropathy autologous renal tubular epithelial antigen as well as immunoglobulins and complement were found in the glomerular capillaries (13). It has been suggested and shown experimentally (14) that the glomerular lesions are caused by endogenous tubular antigens complexed with antibodies. This pathogenetic mechanism would appear to be appropriate in solvent-induced membranous nephropathy inasmuch as solvents are known to cause tubular damage which may be the initial event culminating in glomerular lesions. It is of interest that in gold nephropathy (15) (membranous nephropathy following gold therapy) x-ray microanalysis confirmed the presence of gold within the tubular epithelium and not in the glomeruli in which the lesions were present (16). Due to its insidious nature, solvent-induced membranous nephropathy may not be detected until it is far advanced. At first, it may be manifested only by proteinuria, and edema or nephrotic syndrome may not appear for many years. Screening for proteinuria in exposed individuals provides a simple method of detection. SUMMARY
Membranous nephropathy occurred in four male patients following exposure of 2, 11, 18, and 20 years, respectively, to various organic solvents. There was absorption through lungs and skin. It is impossible to identify the causative agent among the substantial number of solvents to which the individuals were exposed. There may be a causal relationship between membranous nephropathy and solvent exposure. Although the mechanism of production of membranous nephropathy in these cases is not known, an indirect immunological process mediated by endogenous tubular antigen is suggested. Exposed individuals may have proteinuria for long periods before onset of overt manifestations. Examination for proteinuria is a simple method of screening in these cases. ACKNOWLEDGMENTS We thank Dr. Kingsley Kay who obtained and evaluated the occupational who provided biopsy material and clinical information in Case 1: the Medical Bronx Veterans Administration Hospital and Norman Katz for photography, technical assistance.
histories: Dr. John Abbott Illustration Service of the and Ms. Gloria Francis for
Note added in proof: After the manuscript was submitted we learned that Lagrue et al. (17) compared exposure to organic solvents and other inhaled toxic substances in 108 patients with biopsy proven. nonsystemic glomerulonephritis with that of a matched control population. Exposure was more frequent (62% vs 28%) and of a greater degree (6.25 vs 3.90,~ < 0.005) in the patient than in the control group. Of the 108 patients, 17 had membranous nephropathy (18).
REFERENCES 1. Allen, A. C. (1962). “The Kidney: Medical and Surgical Diseases.” 2nd ed. p. 324, Grune & Stratton, New York. 2. Heptinstall. R. H. (1974). “Pathology of the Kidney.” 2nd ed. Little, Brown, Boston. 3. Beirne, G. .I., and Brennan, .I. T. (1972). Glomerulonephritis associated with exposure to hydrocarbons: mediated by antibodies to glomerular basement membrane. Arch. Environ. Health 25, 365-369.
MEMBRANOUS
NEPHROPATHY
45
4. Zimmerman, S. W., Groehler. K., and Beirne. G. J. (1975). Hydrocarbon exposure and chronic glomerulonephritis. Lance? 2, 199-201. 5. Beirne, G. J., Wagnild, J. P., Zimmerman. S. W., Macken, P.. and Burkholder, P. M. (1975). Idiopathic crescentic glomerulonephritis (ICGN)-An analysis of 40 patients. (Abstract). Kid. Int. 8, 406. 6. Ehrenreich, T., and Churg, J. (1968). Membranous nephropathy. In “Pathology Annual.” S. C. Sommers, p 145, Appleton-Century-Crofts, New York. 7. Ehrenreich, T.. Porush. J. G.. and Churg. J., rt al. (1974). Idiopathic and secondary membranous nephropathy. (Abstract). Kid. fnt. A. 6, 38A. 8. Row, P. G.. Cameron, J. S., Turner, D. R., Evans. D. J.. White. R.H.R.. Ogg, C. S., Chantler, C., and Brown, C. B. (1975). Membranous nephropathy: Long-term follow-up and association with neoplasia. Quart. J. Med. 44, 207-239. 9. Churg, J.. and Prado. A. (1956). A rapid Mallory trichrome stain (chromotrope-aniline blue). Arch. Pathol. 62, N-506. 10. Ehrenreich, T.. and Espinosa, T. (1971). Chromotrope silver methenamine stain of glomerular lesions. Amer. J. Clin. Pathol. 56, 448-451. I I. Malten. K. E.. and Zielhuis, R. L. (1964). “Industrial Toxicology and Dermatology in the Produc. tion and Processing of Plastics.” p 12. Elsevier. New York. 12. Malten. K. E., and Zielhuis, R. L. (1964). “Industrial Toxicology and Dermatology in the Production and Processing of Plastics.” p. 149. Elsevier, New York. 13. Naruse. T., Miyakawa. Y.. Kitamura, K., and Shibata. S. (1974). Membranous glomerulonephritis mediated by renal tubular epithelial antigen-antibody complex. J. Aller,~ C/in. Zmmunol. 54, 311-318. 14. Naruse, T., Fukasawa, T.. and Miyakawa, Y. (197.5). Laboratory model of membranous giomerulonephritis in rats induced by pronase-digested homologous renal tubular epithelial antigen. Lab. Invest. 33, 141-146. 15. Tornroth. T.. and Skrifvars. B. (1974). Gold nephropathy prototype of membranous glomerulone. phritis. Amer. J. Pathol. 75, 573-590. 16. Watanabe. 1.. Cuppage, F. E.. Moore, J.. and Whittier, Jr., F. C., (1974). Gold nephropathy: Ultrastructural, fluorescent and microanalytic study of two human cases. (Abstract), Lob. Invest. 30, 392. J. (1976). Nonsystemic 17. Lagrue, G.. Kamalodine. J., Guerrero. J.. Zhepova, F.. Bernaudin. glomerular diseases and inhalation of organic solvents. Kid. Int. (Abstract) 10, 337. 18. Lagrue. G. Personal communication.
THEODORE
Nephropathy following Volatile Hydrocarbons’
EHRENREICH,
L. YUSIS,
STUART
Received
April
Exposure
.-\ND
JACOB
to
CHVRG?
14, 1976
Excessive exposure to solvents has long been known to cause renal tubular disease in man t 1, 2). Recently, the occurrence of glomerular disease such as Goodpasture’s syndrome with proliferative and extracapillary glomerulonephritis has been related to hydrocarbon exposure (3-5). This report concerns four patients with membranous nephropathy who had a history of prolonged exposure to a number of volatile hydrocarbons. Membranous nephropathy is a chronic renal disease involving glomeruli and occurring principally in adults (61. Its clinical onset is insidious and is manifested by proteinuria or edema. Proteinuria is the hallmark of the disease and may be present for many years without symptoms. It is often a slowly progressive disease. About one-fourth of the patients improve clinically and lose their proteinuria, while up to one-fourth develop renal failure. The glomerular capillary wall lesions are distinctive, contain deposits of immunoglobulin and complement considered to represent immune-complexes, and show a morphological progression from early Stage I to late Stage IV (6). While in most cases there is no known etiology, in some patients the disease has been deemed to be secondary to specific diseases or agents such as infections, neoplasms, or chemicals (7, 8). The four cases to be described fall into this last category. METHODS
AND PATIENTS
In the course of investigating a series of patients with membranous nephropathy an occupational exposure to solvents was recorded in three cases. In another case solvent exposure was suspected because of the patient’s occupation (plastics worker). Subsequently, a detailed occupational history was obtained in all four cases. The diagnosis of membranous nephropathy in all patients was made on the basis of an initial kidney biopsy as well as the clinical history. Subsequently, additional kidney biopsies were performed. In Case 1 there was one additional biopsy and nephrectomy, and two additional biopsies in the fourth case. One patient died and an autopsy was performed. ’ Supported in part by USPHS Research Grant AM-00918 from the National Institute of Arthritis and Metabolic Diseases. 2 Reprint requests: Jacob Churg. M.D.. Renal Pathology Division, Department of Pathology. Mt. Sinai School of Medicine of CUNY. 5th Avenue and 100th Street, New York. New York 10079.
36
EHRENREICH,
YUNIS,
AND
CHURG
For light microscopy, the kidney biopsies and autopsy kidney tissue were fixed in 10% formalin with 2% sodium acetate by weight resulting in a pH close to neutral. Tissues were embedded in paraffm and 2-3 pm sections were cut. The following stains were performed: hematoxylin and eosin (H & E), periodic acid Schiff(PAS), periodic acid silver methenamine (PASM), chromotrope aniline blue (CAB), (9) and chromotrope silver methenamine (CSM) (10). For electron microscopy tissue was fixed in 1% osmium tetraoxide in phosphate buffer and embedded in epon. Grids were prepared, double stained with uranyl acetate and lead hydroxide, and examined in an RCA 1llG electron microscope at 50 or 100 kv. REPORT
OF CASES
Case 1 In 1968 proteinuria was discovered on routine physical examination of this 23-year-old male. From 1966 to 1968 he had worked in a photographic film research laboratory and was exposed to organic solvents. At the time of this initial employment, and before being exposed to solvents, his urine was found to be free of protein. He continued to be employed in the same film laboratory and 1 year later was transferred to another film making process in which he was not exposed to organic solvents. Proteinuria persisted and he was admitted to the hospital (1969). The physical examination was essentially negative. The blood pressure was 120/70, the blood urea nitrogen (BUN) and the intravenous pyelogram were normal. Urine showed 2.5 g of protein in 24 hours. Renal biopsy revealed early membranous nephropathy. Subsequently the patient developed edema and was treated with prednisone and furosemide which were ineffective and were discontinued. Another kidney biopsy was performed and showed more advanced membranous nephropathy (1971). In 1972 he developed renal failure with hypertension and was dialyzed. This was followed by bilateral nephrectomy for hypertension and later a transplant. E,rposure Histoq The patient worked in a photographic film research laboratory and his occupation involved a heat process of film making on open equipment using a variety of organic solvents which the patient reported as including acetone, acetonitrile, butyl acetate, butyl alcohol, carbon tetrachloride, chloroform, cyclohexane, dichloroethane, dichloromethane, dimethyloformamide, isopropyl alcohol, methyl alcohol, methyl ethylketone, toluene, tetrahydrofuran, trichloroethylene, and xylene. Some of these solvents were used for the cleanup of equipment and of the patient’s hands. Case 2 In 1964 at the age of 45 the patient was discovered to have 4+ proteinuria in the course of a routine physical examination. At that time there was no peripheral edema, the BUN, serum albumin, serum globulin, and serum cholesterol was normal. A kidney biopsy was performed and showed advanced membranous nephropathy. He continued to work without treatment other than restriction of fluids. The proteinuria persisted without edema. In February 1973, he was found to have elevated BUN, hypertensiona, and persistent proteinuria. The blood plsessure was
MEMBRANOUS
NEPHROPATHY
37
1801124, the fundi showed blurring and narrowing without hemorrhage. He was subsequently treated with methyldopa and chlorothiazide. At this time the urine showed 1.9 g of protein a day, BUN was 53 to 70 mg/lOO ml, serum creatinine 5.2 to 6.0 mg/ 100 ml, creatinine clearance 9 cm3 per minute. He was in renal failure, but it was considered to be stable and no further treatment was undertaken. In February 1974, he had a myocardial infarct without complications, from which he recovered. One year later he was placed on chronic hemodialysis. Exposure
History
This patient worked in the plastics industry from 1946 to 1969 with a variety of materials used in the heated condition for moulding. According to the patient, he worked first with Plexiglas moulded into brush handles and also with polyvinyl acetate in toy moulding. This period covered 2 years. A further 5 to 7 years were spent heat-moulding polyvinyl chloride. From 1964 to 1969 the patient was employed in the heat-moulding of polyvinyl chloride pipe. Since the patient reported being exposed to fumes of formaldehyde it is likely that phenoplasts were manipulated at some stage in his career as these plastics break down on heating and give off formaldehyde. It is a necessary step in moulded plastics production to clean moulding equipment with organic solvents and the patient confirmed that he used a number of organic solvents for cleanup at the end of production shifts. Apart from the foregoing it may be noted that phenoplasts may decompose into phenol, benzene, cresol, and other aromatic hydrocarbons (11). Polyesters disintegrate into acids and alcohols. Polyvinyl chlorides may give off some unreacted vinyl chloride monomer in heating and decompose to give substantial amounts of hydrogen chloride ( 12). One may conclude that the patient had skin and respiratory exposure to solvents; in addition respiratory exposure to at least formaldehyde and hydrogen chloride. Case 3 A male patient was well until 1956 when at the age of 37 he developed nephrotic syndrome which was treated with steroids and diuretics for 1 year with complete remission. At this time all drugs were stopped. In 1966 at the age of 47 there was recurrence of the nephrotic syndrome. In 1967 a renal biopsy was performed and revealed advanced membranous nephropathy. He was treated with steroids with improvement of the nephrotic syndrome. For a number of years he has been on a salt-free diet with diuretics. His present renal status is that of mild proteinuria (1-2 g/day), slightly elevated blood pressure, and a normal BUN. E-x-posure
History
He worked from 1936 in a gyroscope assembly plant. For about 15 years his job involved cleaning parts in an open tank of perchlorethylene. Around 1952 the patient was placed on a precision-type assembly and this involved cementing parts together with an epoxy preparation. First the parts were cleaned with Freon-l 12. If malfunction occurred in the assembled unit, it was taken apart and the cement softened with a “stripping” compound claimed to have a strong smell. Then the parts were cleaned with various solvents including acetone, methylethylketone, perchlorethylene, and trichlorethylene. Following the development ofsymptoms of
38
EHRENREICH,
YL’NIS,
AND
CHURG
kidney disease in 1956, the patient did not work for several months, but then resumed his previous work up to 1967. At this time his work associate, patient 4, developed the same symptoms. Since other workers showed no abnormalities on urinalysis, the symptoms were not deemed to be occupationally induced. However, at this time he was promoted to a “leader” in the same department, a position in which there was considerably less general exposure and no direct contact with cleaning solvents. In 1971 after the death of his associate, patient 4, he was transferred out of assembly to a job without solvent exposure. Case 4 This male patient was in good health until April 1967 when at the age of 44 he developed a typical nephrotic syndrome. Renal biopsy showed early membranous nephropathy. He was treated with steroids and diuretics; this was followed by complications, such as pulmonary embolism, for which anticoagulants were administered. A year later he developed hypertension (blood pressure 170/l 10) and azotemia. At this point he was treated with azathioprine and a second biopsy showed more advanced membranous nephropathy. In February 1971, at the age of 48, he was admitted to the hospital with uremia, grand ma1 seizures, and respiratory difficulty. He was treated with antibiotics, tracheostomy for the respiratory difficulty, received peritoneal dialysis, went into coma and died in severe acidosis. E.xposure
History
He was employed from 1956 in the same gyroscope assembly plant as patient 3. For 11 years he worked side by side with patient 3 sharing his work. He therefore had a similar exposure to various solvents such as acetone, methylethylketone, perchlorethylene, and trichlorethylene. Following the development of nephrotic syndrome in 1967 he had no further solvent exposure. RESULTS Pathological
Obser\?ntions
In Case 1 there was typical progression of membranous changes starting with early lesions (Stage I) in the initial biopsy (Fig. 1). through Stages II and III in a subsequent biopsy and terminating in end-stage kidney in the nephrectomy specimen (Fig. 7). In the first biopsy electron microscopy was not available and on light microscopy the glomeruli appeared to be normal with H & E and PAS stains. However, with CSM stain, small, rounded deposits could be discerned on the outer or subepithelial aspect of the basement membrane. The deposits stained red with chromotrope indicating their proteinaceous nature, and were somewhat more numerous than is generally seen in this early stage. There were no other glomerular, tubular or vascular changes. In the second biopsy the lesions had progressed to the full blown Stage II (as illustrated from another case in Figs. 2 and 3). The deposits were larger, more numerous and separated by thin projections (“spikes”) of the basement membrane both on light and electron microscopy. There was fusion (or loss) of foot processes. In some glomeruli the lesions had progressed to the advanced State III (as illustrated in Figs. 4 and 5). Here the projections encircled the deposits, some of which were incorporated into a thickened basement membrane. In addition there was widening of the mesangium in some glomerular tufts resulting
MEMBRANOUS
FIG. (arrows)
1. Early Stage I, initial of the thin basement
39
KEPHROPATHY
biopsy in Case I. The only change membrane, due to deposits. CSM
in this glomerulus x450.
is focal
FIG. 2. Full-blown Stage 11, Case 4. The basement membrane is thin and there thickening (arrow). Deposits form a thick layer on the outer aspect ofthe basement by thin spikes. CSM x 450.
is minimal membrane
thickening
mesangial separated
40
EHRENREICH,
YUNIS,
AND
CHURG
FIG. 3. Electron microscopy of the biopsy shown in Fig. 2. Numerous subepithelial electron-dense deposits (D) on the outer aspect of the basement membrane (BM) are separated by spikes (arrow). There is loss (fusion) of foot processes. x7290.
FIG. 4. Advanced Stage III, Case 2. The basement increase in mesangium. CSM x270.
membrane
is markedly
thickened.
There
is no
MEMRRANOUS
FIG. 5. Electron micrograph in advanced thin spikes (arrow) and are incorporated others pale. x6173.
NEPHROPATHY
Stage 111. Case 3. The deposits are completely into the basement membrane. Some deposits
41
encircled by are granular.
in partial sclerosis associated with intracapillary hyaline deposits. Focal tubular atrophy was present. In the nephrectomy specimen (Fig. 71 most of the glomeruli were obsolete or obsolescent with adhesions and fibrous crescents. The membranous change could still be discerned in some glomeruli and others were relatively unaffected showing only slight sclerosis. There was widespread tubular atrophy with cast formation and interstitial inflammation. Case 2 (Figs. 4 and 61 had one biopsy with typical advanced Stage III lesions. Case 3 (Fig. 5) also had one biopsy showing both Stage II and Stage III glomerular lesions with little or no increase in mesangium. In Case 4 (Figs. 2 and 3) there were three biopsies all showing Stage II lesions with some progression to Stage III in the last biopsy. Sclerosis, absent in the first biopsy, was present in segments of some glomeruli in subsequent biopsies. Glomerular obsolescence and focal tubular atrophy progressed in a similar manner. At autopsy the membranous change persisted with varying degrees of glomerular sclerosis, obsolescence, and collapse; there was widespread tubular atrophy with many casts and interstitial inflammation. The clinical and pathological findings in all four cases are summarized in Table I. COMMENTS
It is proposed that there may be a causal relationship between solvent exposure and membranous nephropathy in the cases presented. Of the four patients, three
42
EHRENREICH,
YUNIS,
AND
CHURG
micrograph in advanced sclerosis, Case 2. There is ingrowth of mesangial matrix into FIG. 6. Electron the capillary loop and the basement membrane (arrows) which shows advanced membranous change with dense. pale, and granular deposits. x6173.
had a long exposure to a number of known solvents for 2, 11, and 20 years, respectively, before developing symptoms. Two of these patients (Cases 3 and 4) had the same quality of exposure under the same working conditions for different periods. The third patient (Case 2) was exposed for 18 years to solvents generated in the course of heating plastics and two solvents used for cleaning equipment. While specific solvents could not be identified in this case, it is known that there is solvent exposure in the plastics industry. In none of the instances were personal protective measures taken. The morphological stage of membranous nephropathy has been related to the duration of the disease which is believed to begin with the early Stage I (6). In the cases presented, the biopsy following the shortest exposure, 2 years in Case 1, showed early lesions, whereas the lesions were more advanced in the other cases with exposures of 11, 18, and 20 years, respectively. It is not known whether the progression of the glomerular lesions is related to the duration of the exposure or of the disease. Of the four patients, three developed renal failure (two were treated). The fourth, who was treated, has had good renal function with persistent proteinuria 7 years after the onset of symptoms. This may be considered a good response to treatment since he has lost his nephrotic syndrome and the disease is apparently arrested, although not cured. For comparison, in a large series of cases of treated and
MEMBRANOUS
43
NEPHROPATHY
FIG. 7. End stage kidney disease in Case I. Marked glomerular sclerosis atrophy. In oneglomerulus without sclerosis (arrow) membranous nephropathy thick-walled capillaries. PAS x 108.
and widespread tubular is present in the patent.
nontreated idiopathic membranous nephropathy the occurrence of renal failure was 25% (7) If this proportion of renal failure (75%) will be found in larger groups of exposed subjects, it would suggest that solvent exposure results in a more severe form of membranous nephropathy. The pathogenesis of membranous nephropathy following toxic chemicals such as solvents is uncertain. In Goodpasture’s syndrome related to hydrocarbon exposure it was postulated that the inhaled toxic chemicals affected the pulmonary alveoli
Exposure onset
before
Symptoms
at
Initial
Patient
symptoms
1
7 years
onset Proteinuria
Age
Exposure
24
3 years
7
18 years
Proteinuria
4.5
IX years
3
20 years
Nephrotic syndrome
47
30 years
Nephrotic syndrome
44
4
biopsy
of
11 years
I I years
Outcome
Stage of M.N. Early Stage I and
II
Advanced Stage III Full-blown advanced and III Full-blown II
and Stage Stage
II
Renal failure 3 years after biopsy Renal failure 9 years after biopsy Improved with persistent proteinuria Renal failure I year after biopsy: died
Treatment Prednisone
Diuretics
Prednisone
Prednisone Azathioprine
44
EHRENREICH,
YUNIS,
AND
CHURG
giving rise to antibodies which then damaged the glomeruli (3). In the cases of solvent-induced membranous nephropathy there was no history of pulmonary disease, nor were any lung changes found at autopsy. Another mechanism has been described which may be pertinent to these cases. In idiopathic membranous nephropathy autologous renal tubular epithelial antigen as well as immunoglobulins and complement were found in the glomerular capillaries (13). It has been suggested and shown experimentally (14) that the glomerular lesions are caused by endogenous tubular antigens complexed with antibodies. This pathogenetic mechanism would appear to be appropriate in solvent-induced membranous nephropathy inasmuch as solvents are known to cause tubular damage which may be the initial event culminating in glomerular lesions. It is of interest that in gold nephropathy (15) (membranous nephropathy following gold therapy) x-ray microanalysis confirmed the presence of gold within the tubular epithelium and not in the glomeruli in which the lesions were present (16). Due to its insidious nature, solvent-induced membranous nephropathy may not be detected until it is far advanced. At first, it may be manifested only by proteinuria, and edema or nephrotic syndrome may not appear for many years. Screening for proteinuria in exposed individuals provides a simple method of detection. SUMMARY
Membranous nephropathy occurred in four male patients following exposure of 2, 11, 18, and 20 years, respectively, to various organic solvents. There was absorption through lungs and skin. It is impossible to identify the causative agent among the substantial number of solvents to which the individuals were exposed. There may be a causal relationship between membranous nephropathy and solvent exposure. Although the mechanism of production of membranous nephropathy in these cases is not known, an indirect immunological process mediated by endogenous tubular antigen is suggested. Exposed individuals may have proteinuria for long periods before onset of overt manifestations. Examination for proteinuria is a simple method of screening in these cases. ACKNOWLEDGMENTS We thank Dr. Kingsley Kay who obtained and evaluated the occupational who provided biopsy material and clinical information in Case 1: the Medical Bronx Veterans Administration Hospital and Norman Katz for photography, technical assistance.
histories: Dr. John Abbott Illustration Service of the and Ms. Gloria Francis for
Note added in proof: After the manuscript was submitted we learned that Lagrue et al. (17) compared exposure to organic solvents and other inhaled toxic substances in 108 patients with biopsy proven. nonsystemic glomerulonephritis with that of a matched control population. Exposure was more frequent (62% vs 28%) and of a greater degree (6.25 vs 3.90,~ < 0.005) in the patient than in the control group. Of the 108 patients, 17 had membranous nephropathy (18).
REFERENCES 1. Allen, A. C. (1962). “The Kidney: Medical and Surgical Diseases.” 2nd ed. p. 324, Grune & Stratton, New York. 2. Heptinstall. R. H. (1974). “Pathology of the Kidney.” 2nd ed. Little, Brown, Boston. 3. Beirne, G. .I., and Brennan, .I. T. (1972). Glomerulonephritis associated with exposure to hydrocarbons: mediated by antibodies to glomerular basement membrane. Arch. Environ. Health 25, 365-369.
MEMBRANOUS
NEPHROPATHY
45
4. Zimmerman, S. W., Groehler. K., and Beirne. G. J. (1975). Hydrocarbon exposure and chronic glomerulonephritis. Lance? 2, 199-201. 5. Beirne, G. J., Wagnild, J. P., Zimmerman. S. W., Macken, P.. and Burkholder, P. M. (1975). Idiopathic crescentic glomerulonephritis (ICGN)-An analysis of 40 patients. (Abstract). Kid. Int. 8, 406. 6. Ehrenreich, T., and Churg, J. (1968). Membranous nephropathy. In “Pathology Annual.” S. C. Sommers, p 145, Appleton-Century-Crofts, New York. 7. Ehrenreich, T.. Porush. J. G.. and Churg. J., rt al. (1974). Idiopathic and secondary membranous nephropathy. (Abstract). Kid. fnt. A. 6, 38A. 8. Row, P. G.. Cameron, J. S., Turner, D. R., Evans. D. J.. White. R.H.R.. Ogg, C. S., Chantler, C., and Brown, C. B. (1975). Membranous nephropathy: Long-term follow-up and association with neoplasia. Quart. J. Med. 44, 207-239. 9. Churg, J.. and Prado. A. (1956). A rapid Mallory trichrome stain (chromotrope-aniline blue). Arch. Pathol. 62, N-506. 10. Ehrenreich, T.. and Espinosa, T. (1971). Chromotrope silver methenamine stain of glomerular lesions. Amer. J. Clin. Pathol. 56, 448-451. I I. Malten. K. E.. and Zielhuis, R. L. (1964). “Industrial Toxicology and Dermatology in the Produc. tion and Processing of Plastics.” p 12. Elsevier. New York. 12. Malten. K. E., and Zielhuis, R. L. (1964). “Industrial Toxicology and Dermatology in the Production and Processing of Plastics.” p. 149. Elsevier, New York. 13. Naruse. T., Miyakawa. Y.. Kitamura, K., and Shibata. S. (1974). Membranous glomerulonephritis mediated by renal tubular epithelial antigen-antibody complex. J. Aller,~ C/in. Zmmunol. 54, 311-318. 14. Naruse, T., Fukasawa, T.. and Miyakawa, Y. (197.5). Laboratory model of membranous giomerulonephritis in rats induced by pronase-digested homologous renal tubular epithelial antigen. Lab. Invest. 33, 141-146. 15. Tornroth. T.. and Skrifvars. B. (1974). Gold nephropathy prototype of membranous glomerulone. phritis. Amer. J. Pathol. 75, 573-590. 16. Watanabe. 1.. Cuppage, F. E.. Moore, J.. and Whittier, Jr., F. C., (1974). Gold nephropathy: Ultrastructural, fluorescent and microanalytic study of two human cases. (Abstract), Lob. Invest. 30, 392. J. (1976). Nonsystemic 17. Lagrue, G.. Kamalodine. J., Guerrero. J.. Zhepova, F.. Bernaudin. glomerular diseases and inhalation of organic solvents. Kid. Int. (Abstract) 10, 337. 18. Lagrue. G. Personal communication.
