Modified Suture Technique in a Mouse Heart Transplant Model

Transcription

1 Originl Artil Moifi Sutur Thniqu in Mous Hrt Trnsplnt Mol Song Su, 1,2 Tois R. Türk, 1,3 Shngli Wu, 4 Hu Fn, 1,5 Jin Fu, 1 Kun Wu, 1 Ulrih Flögl, 6 Zhoping Ding, 6 Anrs Krin 1 n Olivr Witzk, 1 1 Dprtmnt of Nphrology, Univrsity Hospitl Essn, Univrsity of Duisurg-Essn, Grmny, 2 Dprtmnt of Hptoiliry Surgry, Th Affilit Hospitl of Lu zhou Mil Collg, Chin, 3 Dprtmnt of Nphrology n Hyprtnsion, Fririh-Alxnr Univrsity, Erlngn-Nurmrg, Grmny, 4 Institut for Cllulr Thrputis, Univrsity of Louisvill, USA, 5 Dprtmnt of Hptoiliry n Pnrtosplni Surgry, Bijing Choyng Hospitl Affilit to Cpitl Mil Univrsity, Chin, 6 Institut for Hrt n Cirultory Physiology, Hinrih Hin Univrsity Düsslorf, Grmny. BACKGROUND: Th mous ominl hrt trnsplnttion mol is si n importnt immunologil rsrh mol. W vlop thniqu for pling ntir vrting suturs inst of hlf invrting n hlf vrting suturs for nstomosis twn onor n ripints vl vins. Th purpos of this stuy ws to vlut this moifi mtho. METHODS: Eh thniqu ws us in 25 mi sujt to isogni ominl hrt trnsplnttion. Ripint oprtion tim, grft wrm ishmi tim, tim of vl nstomosis, n r-ting tim wr ror. Aftr trnsplnttion, th hrtt ws plpt through th ominl wll on y for 100 ys. RESULTS: Ripint oprtion tim (40.7 ± 2.5 min vs ± 2.3 min, p < 0.01), v-vl nstomosis tim (8.4 ± 1.3 min vs ± 1.2 min, p < 0.01), n wrm ishmi tim wr signifintly shortr (23.4 ± 1.7 min vs ± 1.6 min, p < 0.01) with th moifi thniqu. R-ting tim ws 1.2 ± 0.4 minuts with th moifi thniqu vs. 1.5 ± 0.5 minuts (p = 0.04). Thr ws tnny for lss surgil omplitions in th moifi group, ut thr wr no iffrns in survivl rts. CONCLUSION: Th nw suturing thniqu for mous ri trnsplnttion filitts sir nstomosis of th outflow trt, thry ruing oprtion, wrm ishmi, n r-ting tims. [Asin J Surg 2011;34(2):86 91] Ky Wors: hrt trnsplnttion mol, htrotopi trnsplnt, mous Introution With th vlopmnt of mirosurgry n molulr iology, th mous mol for orgn trnsplnts hs om inrsingly populr. Th mous hrt trnsplnttion mol is frquntly us us of its onvnin for th xmintion of grft funtion y plption of ri ontrtion. Sin th first htrotopi hrt trnsplnttion in th mous rport y Corry t l, 1 mny susqunt moifitions hv ontriut to th vlopmnt of this niml mol. 2 4 W prviously vlop novl thniqu in th mous hrt trnsplnttion mol tht opt th onor s thori infrior vn v inst of th pulmonry rtry s th outflow trt. 5 Compr with tritionl mthos, this novl thniqu filitts Arss orrsponn n rprint rqusts to Profssor Olivr Witzk, Univrsity Hospitl Essn, Dprtmnt of Nphrology, Univrsity of Duisurg-Essn, Huflnstrß 55, Essn, Grmny. E-mil: olivr.witzk@uk-ssn. Riv: Ot 20, 2009 Rvis: Apr 27, 2010 Apt: My 14, Elsvir. All rights rsrv. 86 ASIAN JOURNAL OF SURGERY VOL 34 NO 2 APRIL 2011

2 MODIFIED TECHNIQUE OF MOUSE HEART TRANSPLANT MODEL vssl nstomosis n thrfor shortns th grft wrm ishmi tim. In this ppr, w rport furthr moifition of our thniqu n first rsults using this moifi thniqu. W pl ntir vrting suturs inst of hlf invrting n hlf vrting suturs for th nstomosis twn onor n ripints vl vins to rs th thnil iffiulty, shortn th oprtion tim, n rs postoprtiv omplitions. Mtrils n mthos Animls Ml C57BL/6J (H-2 ) inr mi (g 8 12 wks n wighing g) wr us for isogni trnsplnttion. Mi wr otin from th Animl Cntr of Essn Univrsity Hospitl (Essn, Grmny). All of thm wr hous unr stnr onitions t onstnt room tmprtur n humiity n sujt to rgulr 12- hour light/rk yls. Foo n wtr wr suppli liitum. All nimls riv r in omplin with th Prinipls of Lortory Animl Cr, n th protool ws pprov y th lol Animl Cr n Rsrh Committ. Instrumnts A si st of mirosurgil instrumnts from Asulp (Tuttlingn, Grmny) ws us, inluing on stright fin-tip forps (Ctlogu numr: BD331R), on stright spring-typ mirosissors (OC498R), on urv springtyp mirosissors (OC499R), Brrqur miro nl holr (FD284R), n two 5-mm mirovsulr ullogs. A mirosurgil mirosop (OPMI 1 FR, Crl Ziss, Grmny) ws us with 10, 15, n 25 in-prour intrhngl mgnifitions. Th suturs for nstomosis of vssls wr m of nonsorl nylon (10-0) n pl with nl (RESORBA, Nürnrg, Grmny). Silk (7-0) ws us for ligtion in onor prourmnt. 5 Ansthsi Mi wr nsthtis with mix solution of 2% xylzin hyrohlori (Rompun, Byr Animl Hlth, Grmny) n 10% ktmin hyrohlori (Dopln, Agrirns Brsil, Lt.) ilut to 2 mg/ml n 10 mg/ml, rsptivly, in 0.9% slin solution. Th solution ws injt intrpritonlly t osg of 0.1 ml for h 10 g oy wight. Stuy protool Fifty mi wr ivi into two groups. Twnty-fiv unrwnt th moifi nstomosis thniqu (moifi mtho group), n th othr 25 unrwnt th stnr mtho tht w prviously introu (stnr mtho group). 5 Surgry All oprtions wr prform unr ln, ut not stril, onitions. Th surgil n othr in vivo prours sri in this ppr wr prform y singl surgon. Mi wr shv n lns with 70% lohol for surgry. All surgil instrumnts n sutur mtrils wr th sm s thos prviously sri. 5 Donor oprtion Th onor oprtion ws prform s prviously sri. 4,5 Brifly, iltrl thorotomy ws prform unr p nsthsi. Th mous ws thn hprinis with 0.1-mL injtion of slin solution ontining 50 IU hprin intrvnously vi th infrior vn v. Th hrt ws thn flush with 2 ml i-ol prsrvtion solution (Custoiol, Köhlr GmH, Hmurg, Grmny) vi th sning ort until omplt rioplgi. Th sning ort ws isst from th surrouning tissu n trnst proximl to th innomint rtry. Th thori infrior vn v ws ut s los s possil to th iphrgm to mximis th imtr of th vssl. Nxt, th hrt ws lift up n 7-0 silk ligtur ws pl roun th hrt to ligt ll othr vssls. Th grft hrt ws susquntly rst n stor in prsrvtion solution t 4 C. Ripint oprtion Unr p nsthsi, th ripint ws pl in th supin position on th oprting or with th ul prt of its oy towr th surgon. Th following prours from vssl prprtion to ort nstomosis wr prform s prviously sri. 5 Brifly, min trnsvrs ominl inision ws m. Th intstins wr push towr th lft si of th omn to xpos th ominl ort n infrior vn v. Th two vssls wr isst from th surrouning tissus. Nxt, n pproximtly 4-mm stion of th two vssls ws prpr for th ltr nstomosis. Two mirovsulr ullogs wr pl to lok th loo flow in oth vssls. Nxt, two optiml opnings wr m in th ASIAN JOURNAL OF SURGERY VOL 34 NO 2 APRIL

3 SU t l ort n vl vin, rsptivly. Th opning in th vl vin ws pproximtly 2 mm phli to th opning in th ort. Th hrt grft ws thn rmov from th prsrvtion solution, n th hrt grft s ort ws nstomos n-to-si with th ripint s ort s prviously rport. 1,2,4,5 Aftr nstomosis of th ort, th ripint ws position with th ul prt of its oy towr th oprtor s right si, n th hrt grft ws pl t th lft si of th ominl vity. At this tim, grt ttntion ws pi to onfirm th orrt irtion of th onor vn v to voi twisting. A simpl n onvnint mtho ws us to omplish this prour: flushing th onor s vn v with ol prsrvtion solution to ngorg n istn it. Thus, th opning of th onor s vl vin ws quit lrly istinguish. Nxt, rnil ornr stith ws insrt through th opning of th onor s n ripint s vn v n ti. A running vrting sutur (out/in-in/out stith pttrn) ws opt on th ntrior si of th vins from rnil n to ul n (usully 5 6 stiths). Th hrt grft ws susquntly turn to th right si of th omn, n th mous ws turn 180 lokwis to rstor th sm position s tht t th ginning of th grft ort-ripint ort nstomosis prour. Th sutur nl ws thn pss through th tringl-lik fil nlos y th onor s ort, onor s vn v, n ripint s vssls (Figur 1A n 1B). Anothr running vrting sutur (out/in-in/out stith pttrn) ws ontinu from th ul ornr to th rnil ornr. With th lst stith, knot ws m with th first sty stith. Aftr ompltion of th ronstrution of th grft s inflow n outflow trts, th two mirovsulr lmps wr opn to r-prfus th hrt trnsplnt. Th grft wrm ishmi tim (tim twn th n of ol storg to r-prfusion) n th r-ting tim (tim to rstortion of hrtt) wr ror. Th intstins wr rstor to thir suitl pl. Th omn ws los in two lyrs with ontinuous 5-0 silk sutur. Th ripint oprtionl tim ws ror s wll. Finlly, th niml ws pl on hting p n monitor visully until it ws ompltly wk. It ws lft to rovr t room tmprtur with immit n unrstrit ss to foo n wtr. A B1 B2 f Crnil si f f Crnil si Cul si Cul si C Crnil si Cul si Figur 1. Th iffrnt nstomosis mthos of th vl vin. (A n B) Moifi mtho. Entir vrting sutur: () hrt, () onor vn v, () onor ort, () ripint vn v, () ripint ort, n (f) tringllik fil; (A n B1). Th sutur squn t th ntrior wll ws from rnil n to ul n, whih is running vrting sutur (out/in-in/out stith pttrn). At th ul n of th vnotomy, th nl ws pss through th tringl-lik fil n th mous ws turn 180 lokwis; (B2) Aftr pssing th nl through, th sutur ws ontinu t th postrior wll from ul n to rnil n, gin s running vrting sutur. Th whol nstomosis ws n vrting sutur without hnging th sutur squn; (C) Stnr mtho. Hlf invrting n hlf vrting sutur t th vl nstomoti stom: () hrt, () onor vn v, () onor ort, () ripint vn v, n () ripint ort. Th sutur squn is from postrior wll (invrting sutur) to ntrior wll (vrting sutur). 88 ASIAN JOURNAL OF SURGERY VOL 34 NO 2 APRIL 2011

4 MODIFIED TECHNIQUE OF MOUSE HEART TRANSPLANT MODEL Furthr osrvtions n msurmnts Th hrtt ws plpt through th ominl wll on y until 100 ys ftr surgry. Th funtion of th onor hrt ws ssss using sujtiv sor of 0 3 (0 for no ting, 0.5 for vry wk ting, 1 for wk ting, 2 for mort ting, n 3 for full ting). 6 In ition, n ltroriogrm (Bio Amp, ADInstrumnts, Austrli) ws mploy to trmin th grft funtion in th moifi mtho group t 30 ys postoprtion. t th vn v s nstomoti stom (survivl rt: 88%). In th moifi mtho group, only on mous i of ling n nothr i of ilus without ny vsulr omplition (survivl rt: 92%). No sttistil iffrn ws foun twn th two groups. Th omprison of th funtion of onor hrts vlut with th plption sor lso show no iffrns twn th two groups. Disussion Sttistil nlysis Th rsults r prsnt s mn ± stnr vition. Sttistil nlysis ws prform y Stunt s t tst. A vlu of p < 0.05 ws onsir signifint. For sttistil nlysis, SPSS 13.0 (SPSS, In., Chigo, IL, USA) ws us. Rsults Th v-v nstomosis tim ws signifintly shortr with th moifi thniqu (8.4 ± 1.3 minuts) thn with th stnr thniqu (12.1 ± 1.2 minuts, p < 0.01). Ripint oprtion tim ws signifintly shortr with th moifi thniqu thn with th stnr mtho (40.7 ± 2.5 minuts vs ± 2.3 minuts, p < 0.01). Similrly, wrm ishmi tim ws signifintly shortr with th moifi thniqu (23.4 ± 1.7 minuts vs ± 1.6 minuts, p < 0.01). R-ting tim ws 1.2 ± 0.4 minuts with th moifi thniqu n 1.5 ± 0.5 minuts with th stnr thniqu (p = 0.04; Tl 1). On month postoprtion, grfts of th moifi mtho group show rgulr hrt funtion in th ltroriogrm. Complitions In th stnr mtho group, on mous i of ling n two grfts stopp ting us of vnous stnosis Tl 1. Comprison twn moifi mtho n stnr mtho (n = 25 pr group) Moifi mtho Stnr mtho Ripint oprtion (min) 40.7 ± ± 2.3 < Wrm ishmi tim (min) 23.4 ± ± 1.6 < Tim for v-vl 8.4 ± ± 1.2 < nstomosis (min) R-ting tim (min) 1.2 ± ± p Th ominl-htrotopi hrt trnsplnt in mi is si n importnt mol for trnsplnt rsrh, ut it is iffiult to mstr within short prio us it involvs n vn thniqu. Tritionlly, n nstomosis twn th onor s pulmonry rtry n th ripint s ominl vn v is sutur to ronstrut th outflow trt of th grft. 1 Th pulmonry rtry of th onor hrt is jnt to n shortr thn th sning ort. Th sning ort is opt to ronstrut th inflow trt of th grft. Consquntly, if w omplt th nstomosis twn th onor s n th ripint s ort, th onor s ort will prtly oupy th surgil fil for onor s pulmonry rtry-ripint s vn v nstomosis, spilly th postrior si of th vins. This inrss th thnil iffiulty of th oprtion n prolongs th ripint oprtion n grft wrm ishmi tim. Morovr, in th tritionl ominl trnsplnttion mol, two nstomoti stoms r jnt to h othr n th vill sgmnt lngth of th onor pulmonry rtry is shortr thn tht of th ort; thus, th grft hrt is pron to rott roun th vssl-form xl, whih sily ls to loking of th outflow trt n ongstion of th grft. In gnrl, plmnt of intrrupt suturs is ommon mtho for vssl nstomosis 7 ; it rsults in lss striturs/stnoss of th nstomosis thn o running suturs. Howvr, intrrupt suturs usully rquir mor tim for knotting. Th itionl tim rquir uring trnsplnttion ggrvts th ishmi injury of th grft. Prhuting nstomosis is postrior-wllfirst ontinuous suturing thniqu for nstomosis of vssls. This thniqu is ommonly prform in stting with limit sp whn th postrior wll of th vssl is iffiult to xpos. Som groups my us it in th mous mol. Howvr, it is iffiult for nw mirosurgil trins to mstr within short tim us of its mning suturing thniqu in th postrior wll, ASIAN JOURNAL OF SURGERY VOL 34 NO 2 APRIL

5 SU t l spilly if th postrior wll is iffiult to visulis. 8,9 Furthrmor, th rtion of vl t th onor s vsulr n is ommon wy to nlrg n nstomoti stom. This thniqu is us in murin trnsplnttion,.g. whn th imtr of th onor s vin is it smllr thn tht of th ripint. Th iffiulty of this mtho is in ontrol of th ngl whil utting th onor s vssl us th rsultnt imtr of th onor s n pns on th utting ngl. If th ut is not t xtly th sm ngl, th orifi will not nily roun/ovl, ut ngulr. In our mol, th onor s vin ws ut s los s possil to th iphrgm to nlrg th nstomoti stom n to hiv similr imtr t th outflow trt in ll oprtions. W prviously mploy th thori vl vin of th onor hrt for th nstomosis with th ripint vl vin to ronstrut th outflow trt of th grft hrt. 5 This shortn oth th oprtion tim n th wrm ishmi tim, n it ru th inin of omplitions suh s vssl twisting or grft hrt ongstion. Howvr, in our stnr mtho mol, th sutur pttrn of th nstomoti stom on th v vin i not hng; w still us th tritionl sutur pttrn prviously sri (prforming th nstomosis of th postrior wll with insi to outsi stiths) (Figur 1C). 1,8 With this mtho, th sutur t th postrior wll follows n in/out-out/in stith pttrn, whih tully is th invrting sutur n oul rsult in th involvmnt of th outr wll s wll s outr tissus of vins into th nstomoti stom. Thrfor, w still xprin th omplition of vnous stnosis with th stnr mtho. An ntir vrting sutur without ny outr loo vssl wlls involv in th nstomoti stom n furthr ru th possiility of vsulr nothlium injury 6 n involvmnt of outsi tissus t th nstomoti stom. This is th il pttrn for vsulr nstomosis. W sign this nw nstomoti pttrn in our moifi mol on th sis of our stnr mtho mol. Our mol prsrvs th vntg of th stnr mtho in tht sgmnt of th onor s vn v of qut lngth n suppli for nstomosis, mking ompltly running-vrting sutur for outflow trt vill. On th othr hn, omplting th nstomosis of th postrior si of th vins ws thnilly th most iffiult prt of th ripint oprtion us th postrior sutur lin must within viw ut ws lwys vry iffiult to xpos. 8 In ition, th sutur squn h to hng t th ul ornr (from n in-out/out-in stith pttrn on th postrior si to n out-in/in-out on th ntrior si) in th tritionl nstomosis, whih rsult in itionl thnil iffiulty. Using our moifi nstomosis pttrn, xposing th postrior si of th vin n hnging th nl sutur squn o not rquir long prios of tim us only on kin of sutur pttrn ns to prform. This simplifis th prformn of th surgry. Nw surgons my mor quikly mstr this sir mtho. Th omprison twn th outoms of th stnr n moifi mthos show tht shortr oprtion n wrm ishmi tims wr hiv in th moifi group n wr ssoit with shortn grft r-ting tims ftr r-prfusion. Howvr, thr wr no signifint iffrns in th omprisons of longtrm survivl rt n ri funtion. This is proly u to th vry short ol ishmi tim; th onor hrt i not suffr svr ishmi injury. In rsrh involving xprimnts on prsrvtion or rioplgi solutions, th onor is usully sujt to prolong ol storg or prfusion, whih ls to mor svr mg to th onor hrt. In this s, th moifi thniqu oul th ttr hoi for ths xprimnts us of th mor stnris wrm ishmi tims. In ition, no vnous stnosis ourr in our moifi mtho group. W ttriut this fining to th omplt vrting sutur pttrn, whih ru th potntil for vsulr nothlium injury n involvmnt of th outr wlls of loo vssls t th nstomoti stom. In onlusion, on th sis of our novl hrt trnsplnt mol in th mous, th moifi mtho furthr filitts th nstomosis of th outflow trt in ripint oprtions. This moifi mtho n mor sily lrn n mstr y novi mirovsulr surgons. Rfrns 1. Corry RJ, Winn HJ, Russll PS. Primrily vsulriz llogrfts of hrts in mi. Th rol of H-2D, H-2K, n non-h-2 ntigns in rjtion. Trnsplnttion 1973;16: Wng K, Zhng N, Li H. Improv thniqu of mous htrotopi hrt grft rtrnsplnttion. Mirosurgry 2006;26: Wng M, Bi J, Bumnn M, t l. Nw mol for simultnous hrt n kiny trnsplnttion in mi. Mirosurgry 2003; 23: ASIAN JOURNAL OF SURGERY VOL 34 NO 2 APRIL 2011

6 MODIFIED TECHNIQUE OF MOUSE HEART TRANSPLANT MODEL 4. Li Q, Wng Y, Yi T, t l. Thniqu for rtrnsplnting htrotopi hrt grfts in mi. Mirosurgry 2004;24: Wu K, Zhng J, Fu J, t l. Novl thniqu for loo iruit ronstrution in mous hrt trnsplnttion mol. Mirosurgry 2006;26: Mrtins PN. Assssmnt of grft funtion in ront mols of hrt trnsplnttion. Mirosurgry 2008;28: Wtn H, U K, Ohkouhi M, t l. Postrior-wll-first ontinuous suturing omin with onvntionl intrrupt suturing for mirovsulr nstomosis. J Ronstr Mirosurg 2006;22: Dj A, Dll Olmo L, Croi R, t l. Htrotopi ri xnotrnsplnttion in ronts: rport of rfin thniqu in hmstr-to-rt mol. Mirosurgry 2005;25: Singh PB, Goyl NK, Kumr A, t l. Rnl trnsplnttion using liv onors with vsulr nomlis: slvgl surgil hllng. Sui J Kiny Dis Trnspl 2008;19: ASIAN JOURNAL OF SURGERY VOL 34 NO 2 APRIL