MUC1 and MUC5AC conserved domains expressed by epithelial tissue in mammalian species.

E LACUNZA; J BARA; A SEGAL EIRAS; MV CROCE

Robinson College, Cambridge, UK

Workshop; 9th International Workshop on Carcinoma-associated Mucins. Mucins in Health and Disease; 2007

  MUC5AC N-terminus cysteine rich regions and the cytoplasmic tail of MUC1 are conserved among different mammalian species. Objetive: to compare the expression of MUC1 and MUC5AC mucins in epithelia from different mammalian species. Materials: Tissue samples from trachea , lung, esophagus, stomach, small and large intestine, liver, pancreas, mammary and salivary glands and bladder were obtained from rat, rabbit, cat, pig, cow and humans. Two anti/human antibodies were employed: CT33Ab, against MUC1CT and 45M1 MAb reactive with MUC5AC. Methods: Tissues were studied by standard immunohistochemistry .Also, tissues were homogeneized, centrifuged and analyzed by SDS-PAGE, Western blot (WB) and Dot blot. Muc5ac mucin from rat stomach homogenates was immunoprecipitated by incubation with 45M1 MAb , immune complexes obtained were bound to protein A Sepharosa CL-4B, eluted and analyzed by SDS-PAGE and WB. Finally, the effect of neuraminidase treatment on Muc5ac-anti-MUC5AC binding was studied. Results: By immunohistochemistry, CT33 Ab was reactive in most tissues while 45M1 MAb showed a staining restricted to gastric surface epithelium and goblet cells from trachea and lung. By WB, with CT33 Ab, a band of approximately 35KDa was detected; reaction with the 45M1 MAb showed bands at > 180KDa  in the stomach samples and lung secretions from rat, cat, pig and cow. Since rabbit lung and stomach samples did not show any reaction, Dot blot assay was employed and a strong reaction was detected in samples treated and untreated with neuraminidase. When Muc5ac was isolated by immunoprecipitation, a band at 200 KDa was obtained. Conclusions:1-By IHC, Muc1 showed a wide expression while Muc5ac reaction was restricted to gastric surface epithelium and goblet cells from trachea and lung.2- By SDS Page and WB, the molecular weight of Muc1CT and Muc5ac was 35KDa and 180-200KDa.3- Methodology employed was useful to isolate Muc5ac.4- Neuraminidase trretment enhanced 45M1 epitope recognition.