Both Celiac Disease (Nontropical Sprue) and Tropical Sprue Are Char- Acterized by Diarrhea, Weight Loss, Steatorrhea and Malabsorption of Various Foodstuffs
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MORPHOLOGIC CHARACTERISTICS OF JEJUNAL BIOPSY IN CELIAC DISEASE AND TROPICAL SPRUE ERIc A. SCHENK, M.D.; I. MICHAEL SAMLOFF, M.D., AND FREDERICK A. KLIPSTEIN, M.D. From the Departments of Pathology and Medicine, University of Rochester School of Medicine and Dentistry, Rochester, N.Y., and College of Physicians and Surgeons of Columbia University, New York, N.Y. Both celiac disease (nontropical sprue) and tropical sprue are char- acterized by diarrhea, weight loss, steatorrhea and malabsorption of various foodstuffs. Malabsorption of folic acid and vitamin B12 com- monly results in a megaloblastic anemia. Celiac disease may be dis- tinguished clinically from tropical sprue by its hereditary nature, its occurrence in temperate as well as tropical climes, and its relationship to dietary gluten. Tropical sprue is found exclusively in tropical and subtropical climes, and occurs in endemic, and occasionally epidemic form. Tropical sprue is not influenced by a gluten free diet, but therapy with folic acid, vitamin B12 or oral antibiotic agents commonly result in a clinical remission. Histologically definable abnormalities of the jejunal mucosa in celiac disease and in tropical sprue have thus far not allowed differentiation of these conditions. Peroral biopsy specimens in both show partial or complete absence of villi, an abnormal surface epithelium and an in- flammatory cell infiltrate in the lamina propria. It has been proposed that this pattern of change in the jejunal mucosa is an entirely non- specific reaction to injury regardless of the inciting cause.' Since the epidemiologic features and response to therapy indicate that celiac disease and tropical sprue are distinct and separate disease processes, the present study was undertaken to determine whatever morphologic similarities and differences might be definable in these conditions by the use of more elaborate cytochemical and histochemical techniques than have heretofore been utilized. This work was supported in part by grants HE o6089 and CA-02332 from the United States Public Health Service, an Advanced Research Fellowship from the American Heart Association (Dr. Schenk), and an Advanced Clinical Fellowship from the American Cancer Society (Dr. Samloff). Presented in part at the Annual Meeting of the International Academy of Pathology, Philadelphia, Pa., March, x965. Accepted for publication, June 22, I965. 765 766 SCHENK, SAMLOFF AND KLIPSTEIN Vol. 47, No. 5 rz4 a) 0 >- 0 0% . + + coan Wo0 V) + U 10 M. It + +T an O %O N 0z 04 el 0 N-. an Ol) H O .t_ 4 + U) V0 0 a1) rx4 0 "? It + N CO 00 e + VdU)En 00 i = + + an 00 '4+ 2 in 0 U 0 a) 00 H an °° + '4 00 () + + coUX Ul) I" 0 N) + 14 + + 0 U) in~ 0 0 . 1 1 a) 1 %q 00 I an IO O4 H C# N T T r. U %00 %0 \0 ci c- a) > "' O? %O It CAn N O't + 0 c' m + + Ud a) cn 16a4 I444 00 Cd u 0~~~~~~~~~= 0.~~~ ~~~~ 54~ ~~~~~~~~~~~~~~~~~~~~c 4 - a0a,0 0 8~~~~~ a .) anco9 cis ~~~~~~~~~~~ .~~~~~~~~~~~~~gC o8 0 a tko 0 40fr p p-C~~ UC), P4 Nov., z965 JEJUNAL BIOPSY 767 MATERIAL AND METHODS Peroral jejunal biopsy tissues were obtained from 7 patients with untreated celiac disease and from 4 patients with untreated tropical sprue. Pertinent clinical features and laboratory data are outlined in Table I. Of the 7 patients with celiac disease, 4 (Cases 2, 4, S and 7) had had recurrent episodes of diarrhea and weight loss since childhood, and 3 (Cases I, 3 and 6) first developed these symptoms in adult life. Two of the patients (Cases I and 2) were relatively asymptomatic at the time of biopsy. Of the patients with tropical sprue, symptoms began, in case i,S months be- fore admission while she was still a resident of Puerto Rico. Case 2, a native of Puerto Rico but a resident of New York City for 14 years, had symptoms for 4 months. Case 3 developed symptoms I year after leaving Puerto Rico and was ad- mitted i month later. Case 4 had symptoms for 6 years while a resident in the Domin- ican Republic and subsequently for i year as a resident of New York City. All biopsies were made under fluoroscopic control in the region of the ligament of Treitz in the morning after a 12- to 14-hour fast. Usually 2, and at times 3 speci- mens were obtained during each intubation without moving the tube by turning the patients into supine, prone and lateral positions. Each biopsy specimen was placed on a no. i Onksponge (Histomed, Inc., Paterson, N. J.) and oriented so that its submucosal surface rested on the sponge. One of the specimens from each case was placed on a cryostat chuck, quick-frozen in dry ice, and sectioned at io p with a Lipshaw cryostat at-28' C. Serial sections were taken up on slides and immediately stored at-85o C in a cascade type deep freeze unit. The following histochemical stains were performed on the frozen sections: acid phosphatase by the method of Burstone2 at pH 5.2, using naphthol AS-TR as substrate; alkaline phosphatase by the method of Burstone2 at pH 8.2, using naphthol AS-MX as substrate; succinic dehydrogenase by the method of Nachlas and asso- ciates,3 using tetranitroblue tetrazolium, but no phenazine methosulfate; lactic de- hydrogenase by the method of Hess, Scarpelli and Pearse,4 using tetranitroblue tetrazolium and DPN but no polyvinyl pyrrolidone; adenosinetriphosphatase (ATPase) by the lead nitrate method of Wachstein and Meisel,5 and also by the calcium-cobalt method of Padykula and Herman;6 cytochrome oxidase by the method of Burstone 7; and diphosphopyridine nucleotide diaphorase (DPN) and triphospho- pyridine nucleotide diaphorase (TPN) by the nitroblue tetrazolium method of Scarpelli, Hess and Pearse.8 Appropriate controls for these histochemical procedures consisted of (a) the use of specific inhibitors, as discussed by the authors of the various methods given above; (b) the incubation of duplicate slides in media with and without substrate; (c) the use of jejunal biopsy tissues obtained from control patients in each incubation media; and (d) the simultaneous incubation of specimens obtained at different times and from different patients during this study. Fresh frozen sections, fixed in io per cent neutral buffered formalin, were stained forlipid with oil red 0. The other specimens were fixed in io per cent neutral buffered formalin, embedded in paraffin, sectioned at 6 g and stained with hematoxylin and easin and with the Gomoritrichrome (for connective tissue), the periodic acid-Schiff (PAS), Alcian blue and mucicarmine (for mucins and mucopolysaccharides) methods. Control patients comprised 13 native inhabitants of the continental United States, 6 males and 7 females, ranging in age from 27 to 74. There were also 2 normal control Puerto Ricans. None of these patients had celiac disease or tropical sprue. Two of the controls were normal male volunteers.Jejunal tissues were obtained and the specimens were processed as described above. 768 SCIIENK, SAMLOFF AND KLIPSTEIN Vol. 47, No. 5 RESULTS Histologic Features of Jejunal Mucosa Normal. With the dissecting microscope the normal villi appeared as tall, delicate, round, finger-like projections. Occasional tongue or leaf shaped forms were present. The average height of the mucosa, measured from the base of the crypts to the tips of the villi, was o.85 mm with a range of 0.78 to 0.9I mm. Microscopic sections showed tall and slender villi. The surface epithelium consisted of regular columnar cells. De- generating cells were frequently seen at the tips of villi. Occasionally, sloughing of single or clumps of the degenerating cells from the tips of the villi was apparent. The lamina propria contained a sparse infiltrate of lymphocytes, plasma cells, histiocytes, and rarely, eosinophils. In- flammatory cells were extremely uncommon between surface or crypt epithelial cells. Endothelial lined vascular and lymphatic channels were always distinct and adjacent to a delicate basement membrane which separated the surface epithelium from the lamina propria (Fig. io). Celiac Disease. Examination of tissue from 5 of the 7 patients with the dissecting microscope showed a flat mucosa with an occasional mosaic pattern. Histologically, the jejunal mucosa was flat because of an absence of villi in all patients. The average mucosal height was 0.72 mm, with a range from o.6i to o.88 mm. The superficial epithelial cells were pseudostratified and non-columnar, with considerable variation in both size and shape. There were nuclear pyknosis and focal cyto- plasmic vacuolization. An inflammatory cell infiltrate composed of lymphocytes and plasma cells was present within the surface epithelium and occasionally within that of the crypts. The basement membrane separating the surface eipthelium from the lamina propria was thick- ened in many areas and stained as collagen. An increase in the amount of connective tissue staining as collagen was present in the lamina propria which contained a dense inflammatory cell infiltrate consisting of lymphocytes, plasma cells, histiocytes and eosinophils. Lymphatic and vascular channels here, especially in the region beneath the base- ment membrane, were difficult to define and either absent or obscured by the dense inflammatory cell infiltrate (Fig. ii). Alcian blue and PAS stains showed a decreased number of goblet cells in the surface epithelium and an increased number in the crypt epithelium. The local- ization and staining reaction of the acid mucopolysaccharides of goblet cells stained with Alcian blue, and the neutral mucopolysaccharides of the brush border stained with PAS, were similar to those seen in control biopsy tissues (Figs. I3 and 14). Tropical Sprue. There was considerable variation in the degree of Nov., z965f JEJUNAL BIOPSY 769 mucosal abnormality. Examination with the dissecting microscope re- vealed a normal villous pattern in case 2, prominent villous ridging in cases i and 4, and a flat convoluted pattern in case 3.