Digestive Diseases and Sciences, Vol. 36, No. 3 (March 1991), pp. 347-352

Usefulness of Fecal al-Antitrypsin Clearance and Fecal Concentration as Early Indicator of Postoperative Asymptomatic Recurrence in Crohn's Disease MONICA BOIRIVANT, MD, FRANCESCO PALLONE, MD, ANTONIO CIACO, MD, MARINA LEONI, MD, STEFANO FAIS, MD, and ALDO TORSOLI, MD

The aim o f this study was to evaluate in Crohn's disease the possible usefulness o f al-antitrypsin clearance and fecal concentration in the early detection of postoperative asymptomatic recurrence. Eleven adult patients with small bowel Crohn's disease undergoing elective resection were enrolled in the study and prospectively followed for one year. Three, six, and 12 months after surgery the al-antitrypsin clearance and fecal concentration were measured, and the disease activity was assessed. All patients were free of active symptoms throughout the study. One year after surgery small bowel radiology was perJbrmed in all patients. Radiographic evidence of recurrent macroscopic disease was found in five of the 11 patients. Three months after surgery both arantitrypsin clearance and fecal concentration were significantly lower (P < 0.01) than before surgery. There was no difference at this time between patients with recurrence and those with no recurrence. In patients with recurrence both al-antitrypsin clearance and fecal concentration significantly increased at six months in comparison with the values at three months (P < 0.02). Both measurements were significantly higher at six and 12 months in this group of patients than in those with no recurrence and in normal controls (P < 0.01). At six and 12 months al-antitrypsin clearance was above the upper normal lim# in all patients with recurrence. We conclude that fecal al-antitrypsin clearance is a noninvasive, inexpensive, sens#ive marker o f asymptomatic recurrence in CD patients who are under regular supervision after surgery. KEY WORDS: Crohn's disease; postoperative recurrence; cta-antitrypsin clearance; fecal al-antitrypsin.

Recurrence after intestinal resection and anastomosis is almost ineluctable in Crohn's disease (CD) (1, 2). There is evidence that gross abnormalities of the Manuscript received March 2, 1990; revised manuscript received June 12, 1990; accepted June 14, 1990. From the Cattedra di Gastroenterologia, Universit~ "La Sapienza," Rome; and Dipartimento di Medicina Sperimentale e Clinica, Universit~ di R. Calabria, Catanzaro, Italy. Work supported in part by grant 280/02.12.01.54, 1986, from the University of Rome " L a Sapienza." Address for reprint requests: Dr. F. Pallone, c/o Cattedra di Gastroenterologia, Clinica Medica 2, Policlinico Umberto I, 00161 Rome, Italy.

intestine proximal to the anastomotic site are detectable in two-thirds of the patients as early as one year after resection and that these changes occur without overt clinical manifestations (asymptomatic recurrence) (3), Although no available treatment has been proven to be of value in symptom-free patients, the early diagnosis of asymptomatic recurrence may be important in order to schedule a proper follow up and to correctly interpret symptoms that may be related to the surgical procedure itself.

Digestive Diseases and Sciences, Vol. 36, No. 3 (March 1991)

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BOIRIVANT ET A L TABLE l. VALUES OF FECAL ~I-ANTITRYPSIN CLEARANCEAND CONCENTRATION, DAILY STOOL WEIGHT, AND SERUM O.~-ANTITRYPSIN IN PATIENTS BEFORE SURGERY AND DURING POSTOPERATIVE FOLLOW-UP

Fecal oQ-AT concentration (mg/lO0 g)

Fecal al-AT clearance (ml/day)

Months after surgery

Months after surgery Patient

Group A 1 2 3 4 5 M --- SEM Group B 1 2 3 4 5 6 M --- SEM Controls M --- SEM

Before surgery

12

Before surgery

3

6

3

6

12

302 352 92 186 85 203 --+ 54

99 88 44 31 24 57 -- 15

289 65 96 86 294 166 -----51

213 144 73 123 246 150 -----31

108 235 50 164 146 141 ----_30

41 55 40 70 22 46 -----8

65 44 70 92 115 77 --+ 12

83 90 30 90 223 103 -----32

229 70 108 81 250 202 157 -----32

48 59 26 31 30 28 37 -----5

61 38 36 22 35 56 41 -- 6

61 43 61 52 26 19 44 --+ 7

291 70 260 49 200 1000 312 --+ 143

21 33 25 24 24 29 26 --+ 2

34 24 14 27 29 54 30 -----5

14 12 20 30 12 25 19 --+ 3

14 -- 1

T h e f e c a l e x c r e t i o n o f a l - a n t i t r y p s i n ( a l - A T ) is a r e l i a b l e m a r k e r o f i n t e s t i n a l p r o t e i n l o s s a n d is i n c r e a s e d in p a t i e n t s w i t h a c t i v e C D ( 4 - 6 ) . B o t h Otl-AT c l e a r a n c e a n d a l - A T f e c a l c o n c e n t r a t i o n h a v e b e e n s h o w n to b e a b n o r m a l in C D (5-7), a n d it has been suggested that both measurements may r e f l e c t in C D t h e o c c u r r e n c e o f a c t i v e i n f l a m m a t i o n in t h e gut (7, 8). R a i s e d v a l u e s o f a r A T c l e a r a n c e h a v e b e e n r e p o r t e d in C D p a t i e n t s w i t h r e c u r r e n c e a f t e r b o w e l r e s e c t i o n (6, 9). T h e a i m o f the p r e s e n t s t u d y w a s to e v a l u a t e the possible usefulness of al-AT clearance and arAT fecal c o n c e n t r a t i o n in the e a r l y d e t e c t i o n o f a s y m p t o matic r e c u r r e n c e in r e s e c t e d C D patients. This pros p e c t i v e longitudinal s t u d y in r e s e c t e d patients ena b l e d us to further e x a m i n e w h e t h e r the raised a l - A T fecal e x c r e t i o n in C D is r e l a t e d to the p r e s e n c e o f intestinal d i s e a s e r a t h e r t h a n clinical s y m p t o m s .

MATERIALS AND METHODS Patients. Eleven patients with small bowel CD undergoing elective surgery were included in the study. The disease was confined to the distal ileum in all patients. The patients' age ranged between 20 and 51 years. Indications for surgery were recurrent subobstruction (5), fistulae (2), failure of medical treatment (2), and intraabdominal abscess (2). The surgical procedure was ileocecal resection with side-to-side ileocolonic anastomosis in all patients. In no patient did histology show inflammation at the resection margins. According to a protocol of regular supervision, patients were evaluated three and six

348

21 -----6

months after surgery and every six months thereafter. At each attendance the disease activity was assessed by the Simple index (10) supplemented by laboratory measurements including ESR, C-reactive protein, and serum orosomucoids (11, 12). One year after resection, the reoccurrence of disease was assessed by radiology (barium meal follow through) in all patients. Informed consent was obtained from all patients enrolled. Recurrence was detected in five of the 11 patients at the anastomotic site (group A), while in six patients no evidence of recurrence was found (group B). Recurrence at the anastomotic site was defined by the presence of narrowing the preanastomotic loop with ulcerations, nodularity, and/or cobblestone appearance. No patient in either group had overt clinical symptoms throughout the study period. The simple index ranged between 0 and 2 (median 1). In those patients with Simple index >0, the number of liquid stools per day accounted for the final score. There was no difference in terms of Simple index between the two groups. Similarly, no significant difference between the two groups was observed at each time interval in terms of ESR, C-reactive protein, and serum orosomucoids. ESR (millimeters per hour) readings were 7 -+ 1 in group A and 10 --+ 7 in group B at three months, 18 - 13 and 11 -+ 10 at six months, and 20 - 15 and 9 - 4 at 12 months. C-reactive protein (milligrams per deciliter) was 0.7 -+ 0.2 in group A and 0.6 --- 0.1 in group B at three months, 0.7 +-- 0.2 and 0.6 --- 0.1 at six months, and 1.0 - 0.5 and 0.7 -+ 0.1 at 12 months. Serum orosomucoids levels (milligram per deciliter) were 90 --- 22 in group A and 76 --- 18 in group B at three months, 93 --- 20 and 90 - 16 at six months, and 106 +-- 36 and 100 - 24 at 12 months. Fecal oq-AT Measurements. The fecal a l - A T excretion was measured three, six, and 12 months after surgery in all patients. Serum and fecal a l - A T concentration was Digestive Diseases and Sciences, Vol. 36, No. 3 (March 1991)

FECAL al-ANTITRYPSIN IN RECURRENT CROHN'S DISEASE

TABLE l. CONTINUED

Daily stool weight (g/day)

Serum al-A T (mg/dl)

Months after surgery Before surgery

3

6

859 470 307 340 140 423 • 120

434 360 438 150 340 344 • 48

350 180 140 496 205 100 245 • 61

400 557 200 300 280 225 327 • 50

Months after surgery 12

Before surgery

3

6

12

800 280 440 250 320 418 • 101

500 400 560 300 320 416 • 45

307 314 166 300 240 265 • 29

180 225 400 338 305 289 • 35

180 190 320 268 125 217 • 31

195 250 230 220 290 237 • 14

340 500 600 300 210 250 367 • 56

800 655 600 415 350 225 507 • 79

445 179 336 299 164 495 320 • 55

175 310 194 232 224 229 227 • 18

189 316 230 360 175 240 252 • 27

184 182 195 240 160 293 209 • 19

191 • 43

quantitated by radial immunodiffusion using commercially available plates (NOR and LC-Partigen, Behring, F.R.G.). Serum and fecal samples were kept at -20 ~ C until tested. Serum cq-AT was measured in undiluted samples and expressed as milligrams per deciliter. A 2-g aliquot of the homogenized 24-h stools was diluted 1 to 4 in saline and spun at 1500-3000 g for 40 min. Fecal al-AT concentration was measured using 20 ixl of the supernatant and expressed as mg/100 g stool weight, al-AT clearance was calculated (cq-AT -- 1 day stool weight x fecal al-AT/serum ~I-AT) and expressed as milliliters per day. Normal values were obtained from 10 healthy controls (laboratory staff). For the statistical analysis, the Student's t test for paired and unpaired samples were used as appropriate.

RESULTS In healthy controls c~I-AT clearance was 14 -+ 5 ml/day, and fecal oq-AY concentration was 21 -+ 6 rag/100 g (mean -+ SEM). Values higher than the mean plus 3 so of the normal control values (ie, ~I-AT clearance 62 ml/day, fecal oq-AT 80 mg/100 g) were considered as abnormal. Serum ~I-AT, oq-AT clearance, a~-AT fecal concentration, and daily stool weight measured in each patient before surgery and in the postoperative follow up are shown in Table 1. Before surgery ~ - A T excretion was abnormal in all patients, and there was no significant difference before surgery between patients in group A and group B in terms of either CXl-ATclearance and fecal ~ - A T concentration. There was no significant difDigestive Diseases and Sciences, Vol. 36, No. 3 (March 1991)

240 + 16

ference between group A and B at each time interval postoperatively in terms of either serum ~l-AT and stool daily output. In both groups stool weight was higher than normal. Three months after surgery mean al-AT clearance and fecal concentration were significantly lower (P < 0.01) than the preoperative values in both groups. There was no significant difference between groups A and B at three months in terms of etl-AT clearance (57 + 15 and 37 + 5 ml/day, respectively). Mean values of fecal a r A T concentration tended to be slightly higher in group A than group B (46 -+ 8 and 26 --- 2 mg/100 g, respectively) although no individual value in either group was higher than the upper normal limit (Table 1). Figure 1 and 2 show that in group A both Otl-AT clearance and fecal concentration significantly increased at six months in comparison with the values at three months (P < 0.02). Both measurements were significantly higher at six months in group A (166 + 51 and 77 -+ 12, respectively) than in group B (41 -+ 6 and 30 -+ 5, respectively) (P < 0.01). In group A, values of etl-AT clearance at 12 months were similar to that observed at six months while fecal ~I-AT further increased at 12 months in comparison to the six-month values (P < 0.05). Both al-AT clearance and fecal concentration at 12 months were significantly higher in group A (160 + 31 and 103 -+ 31, respectively) than in group B (44 + 7 and 19 -+ 3, respectively) (P < 0.01). 349

BOIRIVANT ET AL

20r E

1541

L.

10(1

o I--< I

-~. . . . .ooo.oo~

. . . . ~o~ . . . . . . . . . . . . . . . . . . .

I

I

~ months a f t e r

,~ surgery

Fig 1. Fecal al-antitrypsin clearance in patients with Crohn's disease three, six, and 12 months after bowel resection. Points in the curves are mean values -+ SEM (vertical bars) of five patients with asymptomatic recurrence (solid line) and of six patients with no recurrence (dotted line). Differences between the two groups were significant (P < 0.01) at six and 12 months.

All patients in group A had abnormal ~I-AT clearance at six and 12 months. Fecal al-AT was abnormal in two patients at six months and in four at 12 months. No abnormal value of either al-AT clearance and fecal concentration was found in group B patients at three, six, and 12 months. Thus, positive and negative predictive values at six months were 100% for a~-AT clearance. In addition, two patients in group A were measured for a~-AT clearance at 18 and 24 months and exhibited persistently abnormal values (ranging between 94 and 232 ml/day). Two patients in group B showed a marked increase in the a~-AT clearance (>200 ml/day) six months after the end of the study period (ie, 18 months after surgery) and in both

"~ 120

g v

r

7

T....................'~,-....................~

v u

months a f t e r

surgery

Fig 2. Fecal c~,-antitrypsin concentration in patients with Crohn's disease three, six, and 12 months after bowel resection. Points in the curves are the mean -+ SEM (vertical bar) of five patients with asymptomatic recurrence (solid line) and of six patients with no recurrence (dotted line). Differences between the two groups were significant at six and 12 months (P < 0.01).

350

patients a second x-ray examination showed definite evidence of recurrent CD at the anastomosis site. All patients enrolled in the study have been followed for a minimum of 36 months after surgery. Three patients in group B (patients 1, 2, and 6) developed radiographic recurrence in the period between six and 24 months after the end of the study. Three patients in group A (patients 2, 4, 5) and two in group B (patients I and 2) have developed symptomatic recurrence. In all these five patients colonoscopy was done when symptoms developed and endoscopic evidence of recurrent disease at the anastomotic level was found. DISCUSSION The measurements of either fecal a1-AT clearance and/or concentration have potential usefulness in the assessment of patients with CD (6-8). Our prospective, longitudinal study demonstrated that in patients with CD who had a bowel resection, an increase of a]-AT fecal excretion over the early postoperative period is significantly associated with asymptomatic recurrence. The measurement of the fecal oq-AT clearance proved to be a sensitive and specific marker of recurrent macroscopic disease. Our data showed that a rapid increase of the fecal al-AT clearance during the first year postoperatively is a specific indicator of asymptomatic recurrence. Moreover, in all patients with recurrence, al-AT clearance highly increased as early as six months after surgery, suggesting that this test is reliable in discriminating those patients who are developing a recurrence. The fecal al-AT concentration increased progressively in the postoperative period in patients with recurrence. In this study the fecal al-AT concentration appeared to be less sensitive than oq-AT clearance, confirming data of previous reports (4, 6). However, our data at 12 months showing that fecal at-AT concentration was i n creased when recurrent macroscopic disease was detected suggest that this measurement may well be of value in the diagnosis of established asymptomatic recurrence in resected CD patients. In this study we used nonlyophilized stools. This might have been the cause of the apparent reduced sensitivity of fecal al-AT concentration to predict radiographic recurrence. In fact, the increased stool weight observed in all our patients and, consequently, the increased stool water might have affected the measurements of the fecal eq-AT. MeaDigestive Diseases and Sciences, Vol. 36, No. 3 (March 1991)

FECAL etl-ANTITRYPSIN IN RECURRENT CROHN'S DISEASE s u r e m e n t s of fecal e t l - A T c o n c e n t r a t i o n in lyophilized stool have b e e n reported to correlate well with those in nonlyophilized stools (13), while in other studies this correlation was not found (14). In this study recurrence was defined by radiology. Since our aim was to investigate the possible usefulness of fecal eta-AT in detecting macroscopic recurrence, we decided to use small bowel radiology because it is by far the most used and reliable method for the detection of macroscopic CD and is the only practical noninvasive method for the diagnosis of small bowel CD (15). Furthermore, small bowel radiology is relatively inexpensive, well-tolerated by patients and, therefore, appropriate for a study of asymptomatic patients. Endoscopy proved to be of value in examining the ileocolonic anastomosis and the preanastomotic loop in resected CD patients (3). It is likely that by using radiology we may have missed minute alterations of the mucosal surface such as aphthous ulcers, which are a relatively specific endoscopic finding in CD (16). However, the significance of these tiny ulcers is not clear, and it remains to be determined whether aphthous ulcers at the anastomotic site are an indicator of established recurrence (3, 17). Previous reports based on correlation studies between fecal eta-AT and clinical variables have shown that in CD measurements of the fecal excretion of oh-AT provide a reliable measure of the disease activity and have indicated that fecal eta-AT excretion may reflect the ongoing intestinal inflammatory activity in these patients (6-8, 18). Data of the present study provide further evidence that in CD the fecal excretion of eta-AT as measured by the eta-AT clearance is an indicator of active intestinal disease. In a study using lyophilized stools a correlation between etl-AT clearance and random etl-AT fecal concentration was evidenced, indicating that the ett-AT fecal excretion can be measured reliably by simple and inexpensive methods (19). Whether the intestinal protein loss assessed by measuring the fecal eti-AT excretion entirely depends on the active intestinal inflammation is not clear (9). The role of other factors affecting the structure and function of the intestinal mucosa in CD (20, 21) remains to be established. Whatever the significance of fecal etl-AT in CD, this study showed that fecal eta-AT clearance is a noninvasive, inexpensive, sensitive, and specific marker of asymptoDigestive Diseases and Sciences, Vol. 36, No. 3 (March 1991)

matic recurrence in CD patients who are under regular supervision after surgery. ACKNOWLEDGMENTS The authors are indebted to Professor V. Speranza and Dr. S. Minervini (2nd Department of Surgery, University of Rome) who have operated on all patients included in this study and to Dr. R. Cruciani, ITOR Hospital, Rome, who performed small bowel radiology. The authors also thank Miss Alessandra Piconi, and Mr. Mario Termine for their assistance. REFERENCES 1. De Dombal FT, Burton I, Goligher JC: Recurrence of Crohn's disease after primary excisional surgery. Gut 12:519-527, 1971 2. Greenstein AJ, Sachar DB, Pasternack BS, Janowitz HD: Reoperation and recurrence in Crohn's colitis and ileocolitis. N Engl J Med 293:685-690, 1975 3. Rutgeerts P, Geoboes K, Vantrappen G, Kerremans R, Coenegrachts JL, Coremans G: Natural history of recurrent Crohn's disease at the ileocolonic anastomosis after curative surgery. Gut 25:665-672, 1984 4. Florent C, L'Hirondel C, Desmazures C, Aymes C, Bernier JJ: Intestinal clearance of alpha-l-antitrypsin. Gastroenterology 81:777-780, 1981 5. Thomas DW, Sinatra FR, Merrit RJ: Random fecal alpha-1antitrypsin concentration in children with gastrointestinal disease. Gastroenterology 80:776-782, 1981 6. Karbach U, Ewe K, Bodenstein H: Alphal-antitrypsin , a reliable endogenous marker for intestinal protein loss and its application in patients with Crohn's disease. Gut 24:718723, 1983 7. Meyers S, Wolke A, Field SP, Feuer E J, Johnson JW, Janowitz HD: Fecal a-l-antitrypsin measurement: An indicator of Crohn's disease activity. Gastroenterology 89:1318, 1985 8. Karbach U, Ewe K, Dehos H: Antiinflammatory treatment and intestinal cq-antitrypsin clearance in active Crohn's disease. Dig Dis Sci 30:229-235, 1985 9. Fischbach W, Becker W, Mossner J, Koch W, Reiners C: Faecal alpha-l-antitrypsin and excretion of 111 indium granulocytes in assessment of disease activity in chronic inflammatory bowel diseases. Gut 28:386-393, 1987 10. Harvey RF, Bradshaw JM: A simple index of Crohn's disease activity. Lancet 1:514, 1980 11. Torsoli A, Pallone F, Boirivant M: A contribution to decision making in Crohn's disease. Ital J Gastroenterol 15:138139, 1983 12. Boirivant M, Leoni M, Tariciotti D, Fais S, Squarcia O, Pallone F: The clinical significance of serum C reactive protein levels in Crohn's disease. J Clin Gastroenterol 10:401-405, 1988 13. Catassi C, Cardinali E, D'Angelo G, Coppa V, Giorgi L: Reliability of random fecal a-l-antitrypsin determination on nondried stools. J Pediatr 109:500-502, 1986 14. Wilson CM, McGillan K, Thomas DW: Determination of fecal a-l-antitrypsin concentration by radial immuno-

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BOIRIVANT ET AL diffusion: Two systems compared. Clin Chem 34:372-376, 1988 15. Goldberg HI, Caruthers SB, Nelson JA, Singleton JW: Radiographic fihdings of the National Cooperative Crohn's Disease Study. Gastroenterology 77:925-937, 1979 16. Pera A, Bellando P, Caldera D, Ponti V, Astegiano M, Barletti C, David A, Arrigoni A, Rocca G, Verme G: Colonoscopy in inflammatory bowel disease. Diagnostic accuracy and proposal of an endoscopic score. Gastroenterology 92:181-185, 1987 17. Ni Xin-Yu, Goldberg HI: Aphthoid ulcers in Crohn's disease: Radiographic course and relationship to bowel appearance. Radiology 158:589-596, 1986

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18. Meyers S, Lichtiger S, Feuer EJ, Lahman EA, Janowitz HD: Fecal a-l-antitrypsin as a measure of Crohn's disease activity. The effect of therapy and anatomical extent of disease. J Clin Gastroenterol 10:491-497, 1988 19. Thomas DW, Sinatra FR, Merritt R: Fecal a-l-antitrypsin excretion in young people with Crohn's disease. J Pediatr Gastroenterol Nutr 2:491-496, 1983 20. D'Agostino L, Daniele B, Pallone F; Pignata S, Leoni M, Mazzacca G: Postheparin plasma diamine oxidase in patients with small bowel Crohn's disease. Gastroenterology 95:1503-1509, 1988 21. Carr ND, PuUan BR, Schofield PF: Microvascular studies in non-specific inflammatory bowel disease. Gut 27:542-549, 1986

Digestive Diseases and Sciences, Vol. 36, No. 3 (March 1991)