A324 AGA ABSTRACTS GASTROENTEROLOGY, Vol. 108, No. 4

SPHEROID-L/KE VESICLES: A NOVEL METHOD FOR PREPARATION OF I-HGHLY POLARIZED PRIMARY EPITHELIAL CELLS FROM NORMAL HUMAN GASTROINTESTINUM AS A TOOL FOR INVESTIGATIONS. M.J. Sassier, H.-J. Boxberger, T.F. Meyer, H.-D. Becker. Dept. of General Surgery, University Tuebingen and Dept. of Infektionsbiologie, Max-Planek-Institut, Tuebingen, F.R.G.

Cell cultures of the human digestive tract usually lack histophysiological characteristics including the high degree of polarization, the inner and outer configuration and other features observed in normal tissue. Alternatively tumor cell lines or conventional two-dimansional (2-D) cell cultures were often used for studying physiological mechanisms. Because of poor results in propagating gastrointestinal epithelia, the purpose of this study was to set up an in vitro model of normal human gastrointestinal calls in order to avoid the drawbacks of the 2-D cultures for the study of normal and pathological features such as the infection with Helicobacter pylori and its genetically modified types. For observation electron microscopy was performed.

Human epithelial ceils from stomach, small intestine and colon were propagated in order to grow as spheroid-fike multieallular vesicles. In contrast to the cells grown on artificial substrate the 3-D cells represent a prismatic form, wall developed junctional complexes, desmosomes, dictyosomes and a brush border with glycocalyx and thigbtly packed microvilli. As seen by using the EP4 antibody, the cell membrane is expressing recognizable epitopes. Because gastric cells are still alive under pH 3.5 conditions, the steps of infections with Helieobacter pylori of different modifications (fin-, without urease activity) could be observed by electron microscopy under nearly normal conditions.

In summary our results indicate that normal gastrointestinal cells can be cultured as muhicell vesicles with the consequence of building more structures necessary for epithelial functions. In our opinion, therefore, the results of our studies demonstrate that 3-D call cultures represent a suitable modal system for in vitro research purposes which approximate normal conditions.

CYTOCHROME P450 3A ENZYMES IN INTESTINAL AND COLON CELL LINES. K.-Fr. Sewing, A. Lampen, I. Haokbarth, A. Bader, U. Christians

Intestinal cell lines are well established models for studying transport for xenobiotics, nutrients and ions in the gut mucosa. Drug metabolism in combination with the mucosal barrier and the immune system plays an important role in protecting the organism from absorption of toxic xenobiotica. On the other hand, intestinal metabolism of xenobiotics might be involved in activation of carcinogens and therefore in the development of intestinal cancer. Intestinal cell lines might be a useful tool for further investigating the role of intestinal drug metabolism. In particular enzymes of the P450 3A subfamily are the major P450 component in the small intestine. Activity of P450 3A enzymes was characterized by using the specific P450 3A substrates ciclosporin and tacrolimus. The metabolite patterns generated were quantified by HPLC and HPLC/electrospray/mass spectrometnj. The metabolism of ciclosporin and tacrolimus was inhibited by the specific P450 3A inhibitor troleandomycin and induced by the P450 3A inducer dexamethasone. P450 3A proteins were detected by Western- blot using a specific polyclonal antibody and the respective mRNAs by Northern blotting. The following call lines were examined for P450 3A activities: IEC-6 (rat duodenum), IEC-18 (rat ileum), FHS 74 (fetal human intestine), HUTU 80 (human duodenum), HCT8 (human ileum), CaCo2 (human colon). Of these cell lines only CaCo2 cells metabolized ciclosporin and tacrolimus. The metaboUte patterns were similar to those produced by humansmall intestinal and liver microsomes. Addition of 500 #mol/l troleandomycin inhibited 13-O-demethyl tacrolimus formation by 71%. Western blot analysis showed that P450 3A and P450 1A enzymes are expressed while P450 2E1, 2C and 2136 could not be detected. Pre-treatment of CaCo2 cells with the specific P450 3A inducer dexamethasone failed to induce metabolism of ci¢losporin and tacrolimus. No difference in P450 3A expression could be detected in comparison with untreated cells. Additionally, human P450 3A4 eDNA failed to detect mRNA even under low stringency conditions. Since the P450 3A substrates ciclosporin and tacrolimus are metabolized and metabolism was inhibited by a specific P450 3A inhibitor, CaCo2 cells represent a model for P450 3A-dependent intestinal metabolism. Since P450 3A4 cDNA failed to detect the respective mRNA and dexamathasone did not induce metabolism and P450 3A formation in these calls it is concluded that a P4503A enzyme is expressed that differs from P450 3A3,4 or 5 which requires further identification. Supported by DFG grant SFB280, project A8~

PREGNANCY AND INTESTINAL LYMPHANGIECTASIA: A CASE REPORT HA Shah and RE Sockolow. Division of Pediatric Gastroenterology and Nulrition. Montefiore Medical Center, Albert Einstein College of Medicine. Bronx, New York.

Primary intestinal lymphangiectasla (PIL) is an uncommon condition, leading to a protein losing enteropathy accompanied by hypoproteinemia, lymphopenla, and fat malabsorption. Though there are numerous case reports o f intestinal lymphangieetasia in the English literature, only two of them are about pregnancy and intestinal lymphangiectasia (Hoffbrand 1966 and Liu 1980). These case reports describe uneventful pregnancies with gestationsl remission of the disease, followed by a postpartum relapse.

We report a pregnancy in an 18 year old female with all the cIassical features of PIL. During her pregnancy she was supplemented with parenteral nutrition and an enteral formula high in MCT. Compliance with her diet as well as parenteral nutrition was poor. She had close monitoring of her laboratory values and had frequent ultrasound examinations for fetal development, Her course was marked with multiple episodes of severe hypoalbuminemia ( 0.9 g/dl), hypocalcemia (6.7 mg/dl) and hypomagnesemia (1.0 mEq/L), requiring hospitalizations for corrections of the same. Towards the end of her pregnancy, her central line was removed because of a fungal infection. Despite this, she went on to successfully complete her pregnancy, and delivered a healthy 6 lb 14 oz male by cesarean section. The newborn was noted to have normal serum albumin and "calcium levels. In view of the hypoalbuminemia, there were concerns about the likelihood of proper healing of the cesarean incision, but it healed well Our patient seemed to have temporary remission of her disease with normalization of serum albumin for one month postpartum. She currendy continues to be periodicaUy symptomatic, requiring intervention to correct the serologic abnormalities.

Our case differs from the previously described cases in that her eventful gestation was marked with exacerbations of the disease. This unusual case suggests that a successful pregnancy is feasible in PIL, even when severe hypoalbominemia and hypocalcemia are seen. R highlights the importance of close follow up and frequent monitoring of laboratory studies with prompt intervention as indicated.

EVALUATION OF LATINO EXPATRIATES FOR EVIDENCE OF TROPICAL SPRUE. L.M. Siegel. P.H.R. Green, J. Lindenbanm, C.J. Lightdale, P.D. Stevens, RJ. Garcia-Carmsquillo, S. Goodman, H. Rotterdam. Depamnents of Medicine and Pathology, Columbia University College of Physicians and Surgeons, New York, N.Y.

Over the past 20 years, the number of Latino expatriates in New York City has increased 4field to over 2 million, while the number of patients diagnosed with Tropical Sprue (TS) at our hospital has not changed, averaging 2.5/yr (range: 0-4/yr). Methods: Over a 12 month period, all Latino patients tested for vitamin B12 and folate levels were reviewed. Patients with Confirmed vitamin deficiency who did not have a definite non- sprue etiology (e.g. pernicious anemia, AIDS enteropathy, pregnancy)on non-invasive evaluation underwent push enteroscopy (using the Olympus SIF-100 Enterosc0pe) with biopsy of the jejunum and duodenum. ]~esults: Out of a total of 5755 samples tested, 1571 patients (27%) were of Latino origin by surname. Of this total, 144 (9.1%) had subnormal B12 (<200pg/mi) and/or folate (<2.1ng/ml) levels. 25 undelnvent enteroscopy after initial studies failed to yield a diagnosis. Endoscopically, 2 patients had mucosal scalloping, two had small bowel erythema, and one had hookworms. A diagnosis was established on pathology in 16 patients, including TS in 10, chronic atrophic gastritis in 3, hookworm in one, sarcoidosis in one, and Crohn's disease in one. The two patients with scalloping endoscopically had the greatest degree of microscopic villous atrophy (subtotal atrophy). The 10 with TS ranged in age from 29 to i8, and had been in the US for 1 to 17 years. Eight had abdominal bloating; only 5 had dian'hea. Two were asymptomatic. Low serum B12 was present in 9/10; 5 had low/borderline folate levels. The MCV was elevated in 8 patients (range 99-131fl), although a macrocytic anemia was only present in six. Inflammation and villous blunting were present in both the jejunum and the duodenum in all 10. With therapy, symptoms resolved in all patients. Conclusions: (1) TS is underdiagnosed in the Latin0 communitY of NYC, party due to a lack of classic symptoms. (2) The endoscopic findings of scalloped mucosa and loss of duodenal folds were only seen in patients with severe villous atrophy. (3) Duodenal biopsy is highly sensitive for the diagnosis of TS;