Expression of cytokine genes in rat iris-ciliary body

Thursday, Sep 24, 1992 La Falma/C

X ICER Abstracts 566

4

MOLECULAR RECEPTOR

EXPRESSION BODY.

BIOLOGICAL IDENTIFICATION OF MUSCARINIC SUBTYPE IN HUMAN NONFKfMBNFED BFFPHELLAL

ampmseutbttbetmusfonnedhumau what compooetus of seem nonpigmented epithelial Mttsariuic acetylcboliue receptors mediate secretion in tissues such as stomach aud salivary glauds. Muscarinic

Ciliiry body is the site of aqueous humor pmdur%n. Recent studii show that the ciliary body not only secre&s water and electrolytes but also produces immu substance(s) that is most likety tran factor-6 (TGF-6). To investigate possible production and secretion of

pho
TRANSPORT BY PROTEIN KINASE CILIARY EPITHELIUN (ODNt)

and University KY USA

Eye Research Institute, Medicine, Louisville,

Visual Sciences, of Louisville

Kentucky School

were sacrifii and the irts-ciliary body (ICB) were collected. Poly A+ RNA was recovered from the !CB bySusi . . Minr&ast Track, and CONA was reverse-tmns&ed & degenerate primers for rat interteukin-1 (Ii-l), 11-2, IL-6, TGF61, tumor necrosis factor-a (TNF-a), and y-interferon (INF-y) were synthesized and PCR was performed. Subckxting of the PCR products to plasmids followed nucleo&te sequendng and Southern blot analysis of the PCbF; products confirmed the expression of the penes for TGF-fJ1, TNF-a, INF-y in rat ICB. Thisstudyshowsthatsomf,cytokine~areexpressedinrat ICB, and suggest the cytokmes may p ay rmportant roles under the normal and/or disease states.

C IN Lions

of

We examined whether second messengers modulate ciliary epfthelium Na,K-ATPase activity. Several signal transduction pathways operate in ciliary epfthelfum. Initially we showed that cANP, acting via protein kinare A, may directly inhibit the Wa,K-ATPase (Invest Ophthalnol Vfs Scf 31:2164, 1990). Non e report that protein kfnase C may stimulate Na,K-ATPase. In &(b flux studies with the ODM2 cell line derived frqbuman non-pigmented cilfary epithelium, the ouabafn-sensitive Rb uptake rate was significantly increased by 1,2-dfoctanoylglycerol (DOG) or phorbol 12,13-dfbutyrate (PDDu), protein kinase C activators. In the presence of H-7, a protefn kfnaoe C inhibitor, the size of When protefn klnase C was the PDDu effect was reduced by 60x. down-regulated, the response to PDDu was almost abolished. Amfloride &so fnbtblted the PDDu-induced increase of ouabainsensitive Rb. This observation suggests that a large part of the effect of PDDu upon the Na,K-ATPase is mediated by the activation of the Na-H exchanger which leads to an increased cytoplasmic sodium level. PDDu did not alter the pattern of &Rb efflux, suggesting that potassium channels are not activated. DDG did abolish the ouabaln-insensitive, However, both PDBu a bumetanide-sensitive %tb uptake component suggesting that protein kfnase C actfvatlon also inhibits the Na/K/2Cl cotransporter. Supported

by NE1 Grant

No.

EY06915

566

6 CONTROL OF TRANSPORT WECHANISHS IN THE EPITHELSJM. EPlTHEL!IJM. Mittag, T.W., Wori, H., Rr#%ayashi,

of Ophthalmology and Pharmacology Medicine, New York, NY 10029. The main onuyiaes that

drive

the secretory

, Mount

CILIARY PROCESS Ii. Departments

Sinai

process

School

forming

of

the

agueous humorarethe Wa+/K+ ATPase and carbonic anhydrase. We have t%r&ore attax&ed to eeMalish a connection between these two enzymes and the cAUP signal transdu&ion system. Frob&ns relabadto DARP-32, a phosphoprotein originally found in neural tissue, are also present in ciliary epithelium. In kidney cells and in brain slices the nhos~horvlated form of this p&?in, DARPP-32 aZfeo'& the a&& of iiai~K+ ATPase. ThePh=phorYlationofS pIU@ins (DARP-32 in neural tissue and DARplOO in ciliary m) is dependent on CAMP.

Thus, cAMP m

of dlbry

xediatedby DARP-relatedproteins. COtdin another DARP-related

secreted

into

the

aqueous

epitzldial

cation

pumps

Rovineciliaryproceeses protein of 38 kDa.

humor.

HOWeVer,

onli

may be which

is

mRNA

for

DARP-32 was found--the increased size was caused bv wsttins.laJional glytx%aylation. Ciliary processes also contained a HCO sensitive adanylylcyclase which was independent of for&&in or~&%ulated CAMP formation. Intracellular levels of lico3-, dependent on carbonic anhydraso, may autoregulate transcellular Clflux by controlling Clentry (via HCO -/Clexchange) and Clexit (via CAMP activation of dchannels). These results indicate a fundamental xole 5zrcAMPandite &&en&al connection to the

major

enzymes

responsible

for

aqueous

humor

OF CYTOKINE GENES IN RAT IRIS-CILIARY imdogy, Faculty of Medicine, Kyoto Japan.

ciliary epithclium is importaet for idae+q potential Weatmcnts for Glaucoma. We am utiliig p&muse &am maetiou @CR) to detamiue

REGULATION OF SODIUN-PDTASSIUN CULTURED HUMAN NDN-PIGNENTED

569

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formation.

S.169