INTRODUCTION. Ji-Hye Lee 1, Seon-Mi Yu 1, Eun-Kyung Yoon, Won-Kil Lee, Jae-Chang Jung*, Song-Ja Kim

Transcription

1 J Koren Me Si 27; 22: 89-7 ISSN Copyright The Koren emy of Meil Sienes 2,4 5-Deoxy- -ProstglninJ2 Regultes Deifferentition through Peroxisome Prolifertor-tivte Reeptor- -Depenent Pthwy ut Not Expression in rtiulr Chonroytes Peroxisome prolifertor-tivte reeptors- (PPR- ) is ritil for phenotype etermintion t erly ifferentition stges of mesenhyml ells, wheres its physiologil role is unler. Therefore, we investigte the role of 5-eoxy- -pro- 2,4 stglninj2 (), the nturl reeptor lign for PPR-, on eifferentition n inflmmtory responses, suh s expression n PGE2 proution, in rtiulr honroytes. Our t inite tht the use loss of ifferentite honroyte phenotype s emonstrte y inhiition of type II ollgen n proteoglyn synthesis. lso inue expression n PGE2 proution. The -inue eifferentition effet seems to e epenent on PPR- tivtion, s the PPRE luiferse tivity inrese n PPR- ntgonist, DGE, olishe type II ollgen expression. However, DGE i not lok -inue expression. Colletively, our finings suggest tht PPR- -epenent n -inepenent mehnisms of -inue eifferentition n inflmmtory responses in rtiulr honroytes, respetively. itionlly, these t suggest tht trgete moultion of the PPR- pthwy my offer novel pproh for therpeuti inhiition of joint tissue egrtion. Key Wors : Cell Differentition; Cylooxygense 2; PPR Gmm; Chonroytes Ji-Hye Lee, Seon-Mi Yu, Eun-Kyung Yoon, Won-Kil Lee, Je-Chng Jung*, Song-J Kim Deprtment of iologil Sienes, College of Nturl Sienes, Kongju Ntionl University, Gongju; Deprtment of iology*, College of Nturl Sienes, Kyungpook Ntionl University, Degu, Kore These uthors ontriute eqully to this work. Reeive : 6 Deemer 26 epte : 24 Jnury 27 ress for orresponene Song-J Kim, Ph.D. Deprtment of iologil Sienes, Kongju Ntionl University, 82 Shinkwn-ong, Gongju, Kore Tel : , Fx : E-mil : ksj85@kongju..kr *This work ws supporte y Kore Reserh Fountion Grnt fune y the Koren Government (MOEHRD, si Reserh Promotion Fun) (KRF ), Rheumtoi Reserh Center Progrm (KOSEF Grnt R ). INTRODUCTION Peroxisome prolifertor-tivte reeptors (PPRs) re memers of the lign-tivte nuler hormone reeptor superfmily (). Three PPR isoforms (, n ) iffer in their tissue istriution n lign speifiity (). PPRis present in hert, kiney, n liver n ppers to primrily regulte genes involve in lipi n lipoprotein metolism (, 2), wheres PPR- expression is uiquitous (3, 4) n its physiologil role is unler (5). PPR- is expresse preominntly in ipose tissue, with lower levels in musle n liver (6, 7). lthough the physiologil role of PPR- hs not een lerly etermine, it ppers to e n importnt trnsriptionl regultor of genes involve in gluose n lipi metolism. It ws originlly hrterize s regultor of ipoyte ifferentition n lipi metolism. PPRws lso shown to e expresse in other ell types, inluing T lympoytes n honroytes (8-). PPR- n heteroimerize with t lest one other memer of the steroi reeptor superfmily, retinoi i reeptor (RXR) (). Lign-tivtion of the PPR- :RXR heteroimer hs een shown to moulte the trnsription of genes involve in lipi metolism, inflmmtion, n ell ifferentition (2). Reent eviene lso shows tht the nturl reeptor lign for PPR-, (3), n syntheti ntiieti thizoliineione rugs (e.g. RL49653, rosiglitzone [Rtz] n iglitizone), inhiit mrophge n monoyte tivtion (4) n suppress tumor ell growth (5). Speifi ligns for the PPR- n RXR hve een shown to t synergistilly to inue terminl ifferentition of humn liposrom ells (6) in vitro n to enhne insulin sensitivity in ieti nimls (7). To mintin rtilge homeostsis, rtiulr honroytes synthesize rtilge-speifi extrellulr mtrix moleules, inluing type II ollgen n sulfte proteoglyns (8). This homeostsis is lost in rtilge iseses suh s rheumtoi rthritis (R) n osteorthritis (O), eventully leing to rtilge estrution. Crtilge estrution involves loss of ifferentite honroyte phenotypes (i.e. eifferentition), whih is hrterize y the esstion of type II ollgen expression n onset of firolsti type I n type III ollgen expression (9). Interleukin- (IL- ) n tumor nerosis ftor (TNF ) hve een shown to inhiit the synthesis of rtilge omponents n to promote their egrtion y tivting metlloproteinses (2). Conurrently to these effets, they inue the proution y honroytes 89

2 892 J.-H. Lee, S.-M. Yu, E.-K. Yoon, et l. of inflmmtory meitors suh s prostglnins, nitri oxie (NO), n other retive oxygen speies (2). To te most of the rugs mrkete s nonsteroil nti-inflmmtory rugs or ortiosterois re unle to prevent rtilge mge (22). Thus, intense investigtions re rrie out to preise the trnsution pthwys tht impir rtilge homeostsis in orer to fin new strtegies tht woul e helpful to prevent rtilge estrution. In this stuy, we investigte whether, PPRtivtor, my moulte the ifferentition n inflmmtion in rit rtiulr honroytes. We report here tht PPR- tivtion inues eifferentition ut not inflmmtion in rtiulr honroytes. These results suggest tht PPR- gonists my provie new therpeuti pproh for R n O. MTERILS ND METHODS Monolyer ulture of rit rtiulr honroytes n experimentl ulture onition Rit rtiulr honroytes were relese from rtilge slies of 2-week-ol New Zeln white rits y enzymti igestion. riefly, rtilge slies were septilly issete n then issoite enzymtilly for 6 hr in.2% ollgense type II (38 U/mg soli, Sigm Louis, MO, U.S..) in phosphte-uffere sline, n then iniviul ells were otine y olleting the superntnt fter rief entrifugtion. The ells were resuspene in Duleo s moifie Egle s meium (DMEM, Gio-RL, Githersurg, MD, U.S..) supplemente with % (v/v) fetl ovine-lf serum, 5 g/ml streptomyin, n 5 units/ml peniillin, fter whih they were plte on ulture ishes t ensity of 5 4 ells/m 2. The meium ws hnge every 2 ys fter seeing, n ells rehe onfluene in pproximtely 5 ys. Differentition sttus of rtiulr honroytes ws etermine y exmining the umultion of sulfte glyosminoglyn with lin lue stining or expression of type II ollgen (Chemion, Temeul, C, U.S..) y immunolot nlysis s esrie previously (23). Immunolot nlysis Whole ell lystes were prepre y extrting proteins using uffer ontining 5 mm Tris-HCl, ph 7.4, 5 mm NCl, % Noniet P-4, n.% soium oeylsulfte, supplemente with protese inhiitors ( g/ml leupeptin, g/ml pepsttin, g/ml protinin n mm of 4-[2-minoethyl] enzenesulfonyl fluorie) n phosphtse inhiitors ( mm NF n mm N3VO4). The proteins were size-frtionte y SDS-polyrylmie gel eletrophoresis n trnsferre to nitroellulose memrne. The nitroellulose sheet ws then loke with 3% non-ft ry milk in Tris-uffere sline. ws etete using ntioy purhse from Cymn Chemil (nn ror, MI, U.S..)., ERK-2, n PPR- were etete using ntioies purhse from Snt Cruz ioteh (Snt Cruz, C, U.S..). The ns were visulize using peroxise-onjugte seonry ntioies n hemiluminesene. Immunohistohemistry Rit joint rtilge explnts or rthriti rtilge were fixe in 4% prformlehye in PS for 24 hr t 4, wshe with PS, ehyrte with gre ethnol, emee in prffin, n setione t 4- m thikness. The setions were stine y stnr proeures using lin lue or ntioy ginst type II ollgen or n visulize y eveloping with kit purhse from DKO (Crpinteri, C, U.S..), following the proeure reommene y the mnufturer. PGE2 ssy PGE2 proution ws etermine y mesuring the levels of ellulr n serete PGE2 using n ssy kit (mershm Phrmi ioteh, NJ, U.K.). riefly, honroytes were seee in stnr 96-well mirotiter pltes t 2 4 ells/ well. Following ition of the inite phrmologil regents, superntnt ws use to quntify the mount of PGE2, oring to the mnufturer s protool. PGE2 levels were lulte ginst stnr urve of PGE2 n normlize ginst the mount of genomi DN. Trnsfetion n reporter ssys The reporter onstrut (3xPPRE-TK-lu/pGL3) use in the trnsfetions ontine three opies of the peroxisome prolifertor response element (PPRE) site from the P2 enhner (RE7) inserte upstrem of miniml thymiine kinse (TK) promoter in the pgl3 si luiferse vetor n RXR DN ( gift from Dr. J, Kim t Seoul Ntionl University). Trnsfetion of the reporter vetor ws performe s esrie previously (23). Chonroytes ells were grown in % fetl lf serum/dmem n o-trnsfete with vrious reporter plsmis ( g) n pcmv -gltosise plsmi (.5 g) (Clonteh, Plo lto, C, U.S..) using Lipofetmine (Life Tehnologies, In., Grn isln, NY, U.S..), following the proeure reommene y the mnufturer. The trnsfete ells were ulture in omplete meium for 24 hr n use for further ssy. Cells were trete with inresing onentrtions (-2 M) or times (-24 hr) of. Luiferse n -gltosise tivities were etermine in ll extrts using Luiferse ssy (Promeg, Mison, WI, U.S..) n Glto-light system (Tropix, In., efor, M, U.S..). The tivities were orrete for trnsfetion effiieny with -gl tivity n expresse s fol-inution reltive to the level

3 Cuses Deifferentition in rtiulr Chonroytes 893 in untrete ells. Eh ssy ws performe in triplite. Dt re the men +/- SD of three seprte experiments performe with ells from ifferent nimls (**p. versus non trete ells). Reverse trnsriptse polymerse hin retion (RT-PCR) Primry ulture honroytes were trete with. Totl RN ws isolte using RN STT-6 (Tel-Test, In., Frienswoo, TX, U.S..) n reverse-trnsrie with MoMLV-RT (Invitrogen, Cliforni, U.S..) s previously esrie (26). The following primers n onitions were use for PCR in rit rtiulr honroytes: for type II ollgen (37-p prout), 5 -GC CCC TG CG TC TG CG-3 (sense) n 5 -GC CGC CT TG TGG TCT CC-3 (ntisense) with n nneling temperture of 55 ; for (386-p prout), 5 -GCG CGT GC GC C G GG CC CCC GG TT C GT C-3 (sense) n 5 -CG GG TCT CG TGT TGG G TG CGT CGC TGC TC GCT C-3 (ntisense) with n nneling temperture of 62 ; for (298 p prout), 5 - TC GCC CG CG C TC CT-3 (sense) n 5 -GTG TC TGG TG TC GC CG-3 (ntisense) with n nneling temperture of 52 for glyerlehyes 3-phosphte ehyrogense (GPDH; 299-p prouts), 5 -TC CC TCT TCC GG GC G-3 (sense) n 5 -CC T GCC G GTG GTC GT-3 (ntisense) with n nneling temperture of 5 The primers for rit type II ollgen n rit were e- signe se on the sequene of humn homologs, n sequening of PCR prout showe 89% n 93% homology with orresponing humn genes. Dt nlyses n sttistis The results re expresse s the mens±s.e. vlues lulte from the speifie numer of etermintions. nlysis of vrine (NOV) ws performe to ompre iniviul tretments with their respetive ontrol vlues. The Turkey s honestly signifint ifferene metho ws employe when ( M) (hr) ollgen ollgen ( M) ollgen GPDH sorne (% of ontrol) sorne (% of ontrol) (hr) ( M) C ( M) D Fig.. uses eifferentition of rtiulr honroytes. () Primry ulture honroytes were trete with the PPR- tivtor ( M) for vrious time perios (upper pnel, ) or with the inite onentrtions of for 24 hr (lower pnel, ). Expression of type II ollgen n ws etete using immunolot nlysis. () rtiulr honroytes were left untrete (on) or were trete with M for 24 hr, n expression of type II ollgen,, n GPDH ws etermine y RT-PCR. G- PDH ws use s loing ontrols. (C) n (D) umultion of sulfte glyosminoglyn ws quntifie y lin lue stining. The t represent results of typil experiment ( n ) or men vlues±s.d. (C n D) from t lest four inepenent experiments. The letters on the r inite signifint ifferenes mong mens (n=5) t p=.5 using the Turkey s honestly signifint ifferene metho.

4 894 J.-H. Lee, S.-M. Yu, E.-K. Yoon, et l. mking multiple omprisons mong mens (24). RESULTS inues eifferentition in rtiulr honroytes To exmine the effets of on rtiulr rtilge honroyte ifferentition, rit rtiulr honroytes in primry ulture were trete with M for 48 hr. Uner these experimentl onitions isplye no ytotoxiity, s ssesse y the onversion of tetrzolium ompoun to its formzn prout, s well s trypn lue exlusion (t not shown). inhiite type II ollgen, mrker for ifferentition of honroytes, n SOX- 9 expression, mjor trnsription ftor tht regultes type II ollgen expression, in ose- n time-epenent mnner s etermine y immunolot nlysis (Fig. ). RT- PCR lso showe tht expression of type II ollgen n SOX- 9 ws reue (Fig. ). Similrly, tretment of primry ulture ells loke the umultion of sulfte proteoglyn in ose- n time-epenent mnner (Fig. C, D). The effets of rosiglitzone, PPR- nturl lign, on eifferentition n expression in rtiulr honroytes showe the sme results (t not shown). These results inite tht inues eifferentition of rtiulr honroytes in primry ulture ells. inreses expression n PGE2 proution in rtiulr honroytes ( M) (hr) ( M) GPDH The effet of on expression n PGE2 proution ws investigte using primry ulture honroytes. In honroytes, inrese protein levels of in time- n ose-epenent mnner s etermine y immunolot nlysis (Fig. 2). expression ws inrese hr fter tretment, n the levels peke t 6 hr. tretment, however, i not ffet PPRexpression levels. lso enhne expres- PPR- PPR- PGE2 (ng/well) PGE2 (ng/well) (hr) ( M) C ( M) D Fig. 2. inues expression n PGE2 proution in rtiulr honroytes. () Primry ulture honroytes were trete with ( M) for vrious time perios (upper pnel, ) or with the inite onentrtions of for 24 hr (lower pnel, ). Expression of n PPR- ws etete using immunolot nlysis. () rtiulr honroytes were left untrete (on) or were trete with M for 24 hr, n expression of n GPDH ws etermine y RT-PCR. GPDH ws use s loing ontrols. (C) n (D) PGE2 proution ws mesure using n PGE2 ssy kit n normlize y etermining the mount of totl genomi DN. The t represent results of typil experiment or men vlues±s.d. from t lest four inepenent experiments. The letters on the r inite signifint ifferenes mong mens (n=5) t p=.5 using the Turkey s honestly signifint ifferene metho.

5 Cuses Deifferentition in rtiulr Chonroytes 895 sion s etermine y RT-PCR (Fig. 2). Consistent with the inution of expression, stimulte PGE2 proution in time- n ose-epenent mnner (Fig. 2C, D). The proution of PGE2 ws etermine y using PGE2 ssy kit. PGE2 levels were lulte ginst stnr urve of PGE2. These t inite tht not only uses eifferentition of rtiulr honroytes ut lso stimultes expression n PGE2 proution. uses eifferentition n inution of expression in rtilge explnts To exmine the effets of on rtiulr rtilge honroyte eifferentition n expression, rtilge explnt ultures were trete with M for 24 hr, n rtilge-speifi mtrix moleules were etermine. use rmti loss of type II ollgen Control lin lue ollgen Fig. 3. uses eifferentition n expression in rtiulr honroytes. Crtilge explnts were untrete or trete with M for 24 hr. ollgen,, n proteoglyn were etete y immunohistohemil stining n lin lue stining, respetively. The t represent results of typil experiment onute t lest four times. (Fig. 3, mile pnel) n sulfte proteoglyns (Fig. 3, left pnel) s etermine y immunohistohemil stining n lin lue stining, respetively. However, stimulte expression (Fig. 3, right pnel) s etermine y immunohistohemil stining. These t inite 5- PGJ2 lso inues eifferentition n expression in rtilge explnts. stimultes PPR- -tivtion in rtiulr honroytes To etermine whether the PPR- expresse in the rit rtiulr honroytes were trnsriptionlly tive, PPRE lone upstrem of luiferse (3xPPRE-TK-Lu/pGL3) ws trnsiently trnsfete into rtiulr honroyte ells, n the ells were then trete with. s expete, expression of luiferse tivity ws signifintly inue fter tretment in time- n ose-epenent mnner (Fig. 4). The trnsfetion effiieny ws ove 4% effiieny. These results inite tht inrese PPRtrnsription tivity in rtiulr honroytes. regultes on eifferentition vi PPR- - epenent pthwy ut not expression euse in rtiulr honroytes uses oth eifferentition n expression/pge2 proution, we next exmine whether PPR- tivtion is ssoite with eifferentition n/or expression n PGE2 proution. -inue inhiition of type II ollgen n expression ws prevente y the ition of M DGE (isphenol iglyiyl ether, PPR- ntgonist) (Fig. 5). The -inue inhiition of sulfte proteoglyn on primry ulture honroytes ws PPRE luiferse tivity (Reltive light) PPRE luiferse tivity (Reltive light) (hr) ( M) ( M) (24 hr) Fig. 4. stimultes PPR- trnsriptionl tivity. Primry ulture honroytes ells were o-trnsfete with reporter plsmis (3xPPRE-TK-Lu/pGL3) n RXR expression plsmis (RXR /psg5). pcmv -gltosise plsmi ws use s n internl ontrol for the trnsfetion effiieny. Trnsfete ells were trete with ( M) for vrious time perios () or with the inite onentrtions of for 24 hr (). The ell extrts ws ssye for luiferse tivity n -gltosise tivity. The t represent results of typil experiment or men vlues±s.d. from t lest four inepenent experiments. The letters on the r inite signifint ifferenes mong mens (n=5) t p=.5 using the Turkey s honestly signifint ifferene metho.

6 896 J.-H. Lee, S.-M. Yu, E.-K. Yoon, et l. +DGE ollgen ERK-2 sorne (% of ontrol) DGE Fig. 5. Deifferentition n expression meite y regultes through PPR- -epenent n -inepenent mehnism, respetively. Chonroytes were trete with M DGE (PPR- ntgonist) n then with M for 24 hr. ollgen,,, n ERK-2 were etete y immunolot nlysis (). ERK-2 ws use s loing ontrols. umultion of sulfte glyosminoglyn ws quntifie y lin lue stining (). PGE2 ws mesure using n ssy kit (C). (D) Primry ulture honroytes were o-trnsfete with PPRE-ontining reporter plsmi (3xPPRE-TK-Lu/pGL3) n RXR expression plsmis (RXR / psg5) n pcmv -gl plsmi, n the ells were trete with or without M DGE for 3 min prior to M for 24 hr. The ell extrts were ssye for luiferse tivity n -gltosise tivity. The t represent results of typil experiment () or men vlues±s.d. (, C n D) from t lest four inepenent experiments. The letters on the r inite signifint ifferenes mong mens (n=5) t p=.5 using the Turkey s honestly signifint ifferene metho. PGE2 (ng/well) DGE C PPRE luiferse tivity (Reltive light) DGE D ompletely reovere y DGE pretretment (Fig. 5). - DGE i not ffet -stimulte expression (Fig. 5) or PGE2 proution (Fig. 5C). To eluite the eifferentition mehnism, we exmine PPR- -response element (PPRE)-luiferse tivity. s shown in Fig. 5D, -enhne PPR- tivity ws loke y DGE pretretment. On the sis of these results, we suggest tht regultes eifferentition vi PPR- -epenent pthwy n expression/pge2 proution vi PPR- -inepenent pthwy. DISCUSSION Chonroytes in norml rtiulr rtilge re unique ell type in tht their ifferentite phenotype is reversile. Chonroyte phenotype is regulte y lne of noli n toli moleulr retions tht re involve in mintining homeostsis of rtilge tissue (23). Differentite honroytes lose their phenotype n trnsform into firolst-like ells upon exposure to solule ftors IL- (25), n nitri oxie (26) or uring seril suulture in vitro (23). Suh estrution of homeostsis is elieve to e involve in the pthophysiology of rthritis (27). It ws shown previously tht hs nti-inflmmtory effets on oth humn honroytes () n rheumtoi synovil firolsts (28). However, the present stuy emonstrte the pro-inflmmtory effet of in rit rtiulr honroytes, suggesting toli role of in rtilge. Thus fr, ontroversy still remins regring the role of in inflmmtory responses (29). Inee, the pthwy hs een shown to hve nti- n pro-inflmmtory effet (29), epening on experimentl onitions n/or ell types. PPR- is onstitutively expresse in rtiulr honroytes s shown y immunolot in Fig. 2. The presene of the expression of the PPR- in honroytes my provie new insight in the unerstning of the mehnisms whih le to the loss of rtilge homeostsis. Uner inflmmtory n pthologi onitions, however, PPR- plys n importnt role. tivtion of this nuler reeptor y hs een shown to e ritil for the inution of rheumtoi rthritis n osteorthritis. PPR- thus eserves onsiertion for linil testing in egenertive rtilge iseses (3). In this stuy, we emonstrte tht n inrese in PPRtivities is require for -inue eifferentition of rit knee joint rtiulr honroytes. However, tretment PPR- tivity is not require for inflmmtory responses (i.e. expression/pge2 proution). Thus, -inue expression/pge2 proution is meite y PPR- -inepenent pthwy. In 995, 5- PGJ2 ws reporte to e high-ffinity lign for PPR-, n iverse of its effets hve een ttriute to the tivtion of this nuler reeptor (). However, there is inresing eviene tht hs iverse PPR- -inepenent effets (3). Therefore, further stuies re neessry to eluite inflmmtory responses-relte ifferent signling mehnisms. In summry, urrent knowlege inites tht PPRplys key role in the regultion of eifferentition in rtiulr honroytes. REFERENCES. Tontonoz P, Hu E, Spiegelmn M. Stimultion of ipogenesis in firolsts y PPR gmm 2, lipi-tivte trnsription ftor. Cell 994; 79: Chwl, Lzr M. Peroxisome prolifertor n retinoi signling

7 Cuses Deifferentition in rtiulr Chonroytes 897 pthwys o-regulte preipoyte phenotype n survivl. Pro Ntl Si US 994; 9: Kliewer S, Formn M, lumerg, Ong ES, orgmeyer U, Mngelsorf DJ, Umesono K, Evns RM. Differentil expression n tivtion of fmily of murine peroxisome prolifertor-tivte reeptors. Pro Ntl Si US 994; 9: mri EZ, onino F, ilhu G, umr N, Grimli P. Cloning of protein tht meites trnsriptionl effets of ftty is in preipoytes. Homology to peroxisome prolifertor-tivte reeptors. J iol Chem 995; 27: Shmit, Eno N, Rutlege SJ, Vogel R, Shinr D, Ron G. Ientifition of new memer of the steroi hormone reeptor superfmily tht is tivte y peroxisome prolifertor n ftty is. Mol Enorinol 992; 6: Chwl, Shwrz EJ, Dimulngn DD, Lzr M. Peroxisome prolifertor-tivte reeptor (PPR) gmm: ipose-preominnt expression n inution erly in ipoyte ifferentition. Enorinology 994; 35: Lin FT, Lne MD. CCT/enhner ining protein lph is suffiient to initite the 3T3-L ipoyte ifferentition progrm. Pro Ntl Si US 994; 9: Clrk R, ishop-iley D, Estr-Hernnez T, Hl T, Puington L, Pul SJ. The nuler reeptor PPR gmm n immunoregultion: PPR gmm meites inhiition of helper T ell responses. J Immunol 2; 64: orji K, Grills JP, Gouze JN, Mglou J, Shohn H, Keller JM, inhi, Du M, Netter P, Terlin. Eviene for the presene of peroxisome prolifertor-tivte reeptor (PPR) lph n gmm n retinoi Z reeptor in rtilge. PPRgmm tivtion moultes the effets of interleukin-et on rt honroytes. J iol Chem 2; 275: Fhmi H, Di ttist J, Pelletier JP, Mineu F, Rnger P, Mrtel- Pelletier J. Peroxisome prolifertor--tivte reeptor gmm tivtors inhiit interleukin-et-inue nitri oxie n mtrix metlloproteinse 3 proution in humn honroytes. J rthritis Rheum 2; 44: Kliewer S, Umesono K, Noonn DJ, Heymn R, Evns RM. Convergene of 9-is retinoi i n peroxisome prolifertor signlling pthwys through heteroimer formtion of their reeptors. Nture 992; 358: Fjs L, Deril M, uwerx J. Peroxisome prolifertors-tivte reeptor-gmm: from ipogenesis to rinogenesis. J Mol Enorinol 2; 27: Formn M, Tontonoz P, Chen J, run RP, Spiegelmn M, Evns RM. 5-Deoxy-elt 2, 4-prostglnin J2 is lign for the ipoyte etermintion ftor PPR gmm. Cell 995; 83: Riote M, Li C, Willson TM, Kelly CJ, Glss CK. The peroxisome prolifertor-tivte reeptor-gmm is negtive regultor of mrophge tivtion. Nture 998; 39: Srrf P, Mueller E, Jones D, King FJ, Dengelo DJ, Prtrige J, Holen S, Chen L, Singer S, Flether C, Spiegelmn M. Differentition n reversl of mlignnt hnges in olon ner through PPRgmm. Nt Me 998; 4: Tontonoz P, Singer S, Formn M, Srrf P, Flether J, Flether CD, run RP, Mueller E, ltiok S, Oppenheim H, Evns RM, Spiegelmn M. Terminl ifferentition of humn liposrom ells inue y ligns for peroxisome prolifertor-tivte reeptor gmm n the retinoi X reeptor. Pro Ntl Si US 997; 94: Mukherjee R, Dvies PJ, Cromie DJ, ishoff ED, Cesrio RM, Jow L, Hmnn LG, oehm MF, Monon CE, Nzn M, Pterniti JR Jr, Heymn R. Sensitiztion of ieti n oese mie to insulin y retinoi X reeptor gonists. Nture 997; 386: DeLise M, Fisher L, Tun RS. Cellulr intertions n signling in rtilge evelopment. Osteorthritis Crtilge 2; 8: Snell LJ, igner T. rtiulr rtilge n hnges in rthritis. n introution: ell iology of osteorthritis. rthritis Res 2; 3: ren WP, Dyer JM. Inhiition of the proution n effets of interleukin- n tumor nerosis ftor lph in rheumtoi rthritis. rthritis Rheum 995; 38: Lotz M, lno FJ, von Kempis J, Duler J, Mier R, Villiger PM, Geng Y. Cytokine regultion of honroyte funtions. J Rheumtol Suppl 995; 43: Dingle JT. Crtilge mintenne in osteorthritis: intertion of ytokines, NSID n prostglnins in rtiulr rtilge mge n repir. J Rheumtol Suppl 99; 28: Yoon YM, Kim SJ, Oh CD, Ju JW, Song WK, Yoo YJ, Huh TL, Chun JS. Mintenne of ifferentite phenotype of rtiulr honroytes y protein kinse C n extrellulr signl-regulte protein kinse. J iol Chem 22; 277: SS Institute 989. SS/STT User s Guie, Version 6. SS Institute, Cry, NC, US. 25. Golring M, irkhe J, Snell LJ, Kimur T, Krne SM. Interleukin suppresses expression of rtilge-speifi types II n IX ollgens n inreses types I n III ollgens in humn honroytes. J Clin Invest 988; 82: Kim SJ, Ju JW, Oh CD, Yoon YM, Song WK, Kim JH, Yoo YJ, ng OS, Kng SS, Chun JS. ERK-/2 n p38 kinse oppositely regulte nitri oxie-inue poptosis of honroytes in ssoition with p53, spse-3, n ifferentition sttus. J iol Chem 22; 277: Gy S, Gy RE, Koopmn WJ. Moleulr n ellulr mehnisms of joint estrution in rheumtoi rthritis: two ellulr mehnisms explin joint estrution?. nn Rheum Dis 993; 52 (Suppl ): Tsuouhi Y, Kwhito Y, Kohno M, Inoue K, Hl T, Sno H. Feek ontrol of the rhionte se in rheumtoi synovioytes y. iohem iophys Res Commun 2; 283: Shit T, Kono M, Osw T, Shit N, Koyshi M, Uhi K. 5-eoxy-elt 2,4-prostglnin J2. prostglnin D2 metolite generte uring inflmmtory proesses. J iol Chem 22; 277: Shn ZZ, Msuko-Hongo K, Di SM, Nkmur H, Kto T, Nishiok K. potentil role of 5-eoxy-elt-postglnin J2 for inution of humn rtiulr honroyte poptosis in rthritis. J iol Chem 24; 279: Glss CK, Rosenfel MG. The oregultor exhnge in trnsriptionl funtions of nuler reeptors. Genes Dev 2; 4: 2-4.