Biology of the Cell. Volume 63. 1988

ULTRASTRUCTURE OF ACCESSORY ADRENOCORTICAL NODULES IN WlSTAR RATS. Anna S. BELLONI ( 1 ) , Francesco MUSAJO (2) , Giuseppina MAZZOCCHI (1) and Gastone G. NUSSDORFER ( 1 ) . (1) Department o f Anatomy and (2) 2nd S u r g i c a l C l i n i c , U n i v e r s i t y o f Padua, I t a l y .

More than 70% o f b i l a t e r a l l y a d r e n a l e c t o m i z e d Wis t a r r a t s s u r v i v e and w i t h i n t h r e e months deve lop one o r two consp icuous a d r e n o c o r t i c a l nodules (2-3 mm in d i a m e t e r ) , which, thouEh d i s p l a y i n g an obvious histoloEical zonation, are not associated with chromaffin tissue. Middle and inner cells of the nodules

are similar to zona fasciculata (ZF) and zona reticularis elements of the adult rat adrenal Eland, res

pectively. Middle cells show numerous round mitochondria with vesicular cristae, while inner cells con

tain ovoid mitochondria with tubule-convolute cristae. Both cell types possess abundant smooth endopla

smic reticulum and few lipid droplets, as well as a very elaborated microvillous system. Conversely,

outer (subcapsular) cells of the accessory nodules do not resemble lena Elomerulosa (ZG) elements,

since they display ovoid mitochondria with vesicular cristae instead of elonEated orEanelles with tubu

lo-laminar cristae. Moreover, morphometry shows that their volume is about two-fold that of the true

ZG cells (1400 um3 versus 700 um3). These morpholoEical data accord well with the fact that basal plasm8

concentration of aldosterone, at variance with that of corticosterone, remains very low three months

after bilateral adrenalectomy. The lack of differentiated ZG cells in the accessory adrenocortical nod

ules could be explained by the very elevated blood level of ACTH (about 5-times hiEher than in sham-

operated rats), in fact, chronic ACTH hypersecretion is known to transform ZG cells into ZF elements

(i). Some lines of evidence suEEest the existence of a paracrine control of adrenal zona corticalis by

zona medullaris (i). We think that accessory adrenocortical nodules, lackinE zona medullaris, could be

a Eood experimental model to Eain insiEht into this problem.

(i) Nussdorfer G.G., Int. Rev. Cytol. 98, 1-405 (1986).

TRANSFERT TRANSEPITHELIAL DE PROTEINES AU NIVEAU DE L'INTESTIN POSTERIEUR CHEZ SALMO GAIRD- NERI. Marie-France SIRE(1), Pierre-Yves LE BAIL (2), Uranie GEORGOPOULOU(1) et Jean-Marie VERNIER(I). (lJ Laboratoi~e de ~ p h y s ~ o l o g i z d~ la N , ~ o n d~ Poissons, UA 646 CNRS, Bat 44~, U~ iv~ i t~ Pari~ Sad, 91405 Or~ay C~dzx, Franc~; (2) La~or~oir~ d~ Physio~ogie d~s Poisson~, INRA, CoJ~u~ d~ B~au~i~, ~5042 R~nn~s C~d~x, Franc~.

Chez la Truite arc-en-ciel, les diffdrenciations structurales observdes, au niveau des cellules dpith~liales de l'intestin postdrieur, conf~rent ~ celui-ci une fonction permanente d'absorption des protdines sous forme macromoldculaire. Les protdines absorbdes sent transfdr~es dans un syst~me vacuo- laire supranucldaire o~ elles sent ddgraddes.

Nous nous sommes demandds si des protdines ne pouvaient pas dchapper ~ la ddgradation intracellulai. re , traverser l'dplthdlium et ~tre a l'origine d'une rdponse immunltaire.

Nous visualisons le transfert partiel a l'espace intercellulaire de la peroxydase de raifort (HRP, PM 40 000 - cytochimie) et de l'hormone de croissance bovine (b-GH, PM 20 000 - anti GH-protdine A-or colloldal). Ce transfert s'effeetue via un syst~me vdslculaire tandis que l'essentlel des protdines est retrouvd dans les vacuoles du hyaloplasme aplcal. Depuls l'espace intercellulalre, les protdlnes antlgdniques gagnent l'espace interstitiel de la lamina propria o~ elles peuvent entrer en contact avec les nombreuses cellules infiltrdes, macrophages en particulier qui los internalisent. Les deux protdi- nes sent retrouvdes dans les grains des "granule cells" souvent assocides aux dldmonts vasculaires sous dplthdllaux de la lamina proprla. Cos cellules,carao~drlstlques du tube digesti£ des Salmonidds, appartlendralent au syst~me lympho~de de l'animal.

La prdsence de ces protdines dans le plasma a pu ~tre ddmontrde et quantIFi4el - pour I'HRP en combinant ELISA et chet,iluminescsnce~

- pour la b-GH par dosages radioimmunologlque (RIA) et par radlordcepteurs hdpatlques (RRA).

HOLECULAR IHHUNOELECTRON HICROSCOPY AND IHAGE PROCESSINGz AN APPROACH TO EPITOPE HAPPING ON HIGH MOLECULAR WEIGHT PROTEINS

J.LANY (1), P.BILLIALD ( I ) j J.C.TAVEAU ( l t l N.BOISSET ( l ) j O.HOTTA (2), J.N.LAIIY ( I t (1) Laboratoire de Biochimie, Facult6 de Pharmacie, 2bis Bd TonnellG, 37042 TOURS CEDEX. (2) Centre Harcel DelGpine, CNRS, Avenue de la Recherche Scientifiques 45045 ORLEANS CEOEX

Localizing epitopes on high-Hr proteins can be approached by various methods including correlations between immunological a f f i n i t i es and various structural properties (aminoacid sequencew hydrophi l ic i ty , cr istal lographic temperature factors e t c . . ) . Among these methodsp X-ray cristal lography and molecular immunoelectron microscopy (HIEH) lead to the direct local izat ion of epitopes on antigens. Of course, the resolution of HIEH is worse than that of cristal lography but this disadvantage is balanced by the fact that MIEH requires no cr is ta l and can be used with any high-Hr antigen.

Original ly, our laboratory used MIEH to solve the quaternary structure of hemocyanin, a 24-merit copper containing respiratory pigment of the scorpion Androctonus austra l is (L~u~y et al. 1981, 2__0 IB49-1856). The method consisted of an incubation of the native oligomer with subunit-specific polyclonal Fab fragments, followed by an observation in the electron microscope of the pur i f ied immunocomplexes.

Recently, three improvements were added to the method: 1/ Polyclonal antibodies were replaced by monoclonal antibodies. 2/ Mult ivariate s ta t i s t i ca l methods (correspondence analysis hierarchical c lass i f icat ion) allowed the discrimination of images with subtle differences enabling a precise local izat ion of the epitope-protope contact area with respect to the contour line of the antigen. 3/ Overlaps between neighboring epitopes were systematically investigated by immunological (ELISA), biochemical (PAGEt and E.H. methods (MIEH using pur i f ied subunitst.

The result ing procedure is now being used with various antigens including scorpion hemocyanin and human alpha2-macroglobulin.

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