D-4F Does Not Mitigate Key Parameters in Rat Model of Severe Pulmonary Hypertension
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1 Marquette University Biomedial Engineering Faulty Researh and Publiations Biomedial Engineering, Department of D-4F Does Not Mitigate Key Parameters in Rat Model of Severe Pulmonary Hypertension Robert C. Molthen Marquette University, Shelley Baumgardt Medial College of Wisonsin K. A. Prithard Jr Medial College of Wisonsin Published version. Published as part of the proeedings of the onferene, Proeedings of the 8th World Congress for Miroirulation, Milwaukee, Wisonsin, 2007: Publisher Link Medimond Editore, International Proeedings Division. Used with permission. Robert Molthen was affiliated with the Medial College of Wisonsin at the time of publiation.
2 D-4F Does Not Mitigate Key Parameters in Rat Model of Severe Pulmonary Hypertension R. Molthn, S. Baumgardt, and K.A. Prithard Jr. Departmelll of Veterans Affairs, 'ltlbloki VAMC and Medial College of Wisonsin, Milwaukee. USA Summary We e mployed a newly developed rat model of severe pulmonary hypertension (PH) to study if treatment with apolipoprotein (apo)-mimeti peptide, D- 4F, mitigates PH and re lated path ology. Sprague-Dawley rats underwent surgery to partiall y li gate their left pulmonary artery, after 3-days th ey were injeted with monorotaline (MeT) and then randoml y seleted to reeive a daily injeti on of D-4F or saline. After 2 1-days of treatment the ani mals were examined with respet to isolated lung hemodynami measurement s and X ray mirofoal angiography. T here was no signifiant differene in ri ght-toleft heart weight, hange in body weight, pulmonary vasular resistane (PVR), or pulmonary miro-vasul ar density between D-4F and saline treated rals. Introdution Pulmonary hypertension results in remodeling of large and small pulmonary vessels, a disturbane in the deliate balane of venti lation/perfusion, and inreased load on the right heart. In late slages of the disease, severe PH is aompanied by plexiform and neoi ntimal les ions whih signifi antl y inrease pulmonary vasular resistane I. '. A we ll aepted rat mode l of PH in volves a si ngle subutaneous injetion of MeT, however, thi s model does not resu It in neointimal lesions. T herefore, we have developed a rat model of severe PH that ombines partial li gati on of the left pulmonary art ery w ilh MeT injeti on resulting in the formation of neointimal lesions wh ih more losely mimis severe PH in human" The model is inherently useful for 2007 by MEDIMOND S.r.I. H815C
3 210 8th World Congress for Miroirulation investigating possible anti-remodeling treatments aimed at reduing ellul ar proliferation and, in turn, lowering pulmonary vasu lar resistane. Apolipoprotein (a po) A-I and apoa-i-mimeti peptides hold great promise for treatment of atheroslerosis" '. D-4F has been shown to signifiantly redue rapidly evolving atherosleroti lesions, oxidation and innammation; and, there is evidene that it has the ability to improve endothelial funtion"' '. For these reasons, D-4F was thought to be a good andidate for treatment of vasular remodeling in this proliferative model, in whih histology also onfirmed marked innammatory infiltrates'. Materials and Methods All the studies were done under approval of the Zabloki VA Medial Center IACUC review board and in ompliane with the National Researh Counil's Guide for the Care and Use of Laboratory Animals. Fifteen (15) male Sprague-Dawley rats ( gm) underwent a reovery surgery in whih the rat is anesthetized, intubated, then through thoraotomy the left pulmonary artery is isolated, looped with 4-0 suture and redued suh that the outer vessel diameter was 0.61 mm. The animals were allowed to reover for 3-days, then injeted with MCT (60 mg/kg, sub-q), and randomly seleted to reeive an injetion of either D-4F (I mg/ kg/day, i.p.) (N=8), or an equal volume of saline (ontrol C, N=7). Twenty-one (21) days after injetions. the animals were anesthetized, a ardia blood sample drawn, the main pulmonary artery and trahea annulated, and the heart and lungs exised. The right ventrile was disseted from the heart and both segments weighed separately as a measure of right ventriular hypertrophy. The isolated intat lung was prepared as previously desribed"" Brieny, the lung was ventilated, rinse of blood and perfused with a physiologi saline solution. Then the airway pressure held onstant at 6 mmhg, the pulmonary vein pressure (Ppv) kept ambient. and pulmonary arterial (I' PAl pressure measured over a range of now rates (Q, 0-40 ml/min). Finally, the perfusate was replaed with a vasular X-ray ontrast medium and mirofoal angiography pelformed. All statistial omparisons were pejformed usi ng at-test (Pd"0.02) SigmaStat 3.0 (S PSS, Chiago, IL). Results There was no signifiant differene in key struture/funtion parameters of the PH model between the D-4F treated and the ontrol rats. No signifiant hange was deteted in the right ventriular hypertrophy, as measured by the ratio of the weight of the ri g ht ventrile ompared to the left ventrile plus septum, see Figure I. There was no signifiant differene in the gain in body weight over the 24-day experiment, see Figure 2. Treatment with D-4F did not result in a signifiant derease in pulmonary vasular resistane, (PPA - P,,)/Q,()(), where Q,oo indiates now rate was normalized to body weight at 100 ml/min/kg, see Figure 3. Mirofoal X-ray angiography did not detet any differene in the pulmonary mirovasular density, whih is signifiantly redued
4 Milwaukee, Wisonsin, USA, AI/gusT 15-19, E 06 D> ';; J t: i 0.-4 ~ Z ~ 02 0: 0.0 '---- Figure I. Ratio of the weight of th e righl ventrile ompared (a {he left ventrile plus seplllm. as a measure 0/ right ventriular hypertrophy. in MeT models of PH', as shown in Figure 4. Interestingly, 2 of the rats that were reeiving OAF treatment di ed approximately 17 days into the experiment (none o f the ontrol rats died) and 0-4F treated rats appeared to have more labored breathing and their lungs appeared worse ompared to nontreated PH rats on gross assessment. Conlusion Treatment with OAF was not shown to mitigate measures of strutural or E.2' ~ J ~ GO ~ D,5; '" '0. U, Figure 2. Gain in body weight over the 24-days of exper;melll.
5 212 8th World COl/gress for Miroirlliatiol/ 030 D.DD ' ' FiKure 3. Pulmonary vasular resistane at a flow rate oj 100 mllminlkg. D-4F - Figure 4. Example high~magnifialion X-ray 011g;08'-01ll,\' of lower /ullg lobe in ontrol Gild D-4F Ireared rats. Inrravasular pres,mre is sel to 21 mmhg and airway pressure 6 mml-ig. Sale 011 left image 1001tlll. funtional injury indued by a model of severe PH. For some animals it may have exaerbated this experimentall y-indued illness and/or the progression of th e disease model. 0-4F has been shown to be highly effetive at improving endothelial ell funtion, whih may playa role in reduing rapid evolving vasular lesions and oxidative injury in experimentally-indued atheroslerosis. MeT is known to be an endothelial ell toxin, however, knowledge
6 Milwaukee, Wisonsin, USA, August 15-J9, of the role and amount o f oxidative stress in the M et m odel is limited 'O One possibility is that D -4F m ay aelerate vasular olusi ve lesions in this model of severe PH by proteting endothelial ells from apoptoti ell death. D -4F may resue endothelial ells that sloughed off of the vessel wall, allowing them to remain in irulati on lo nger than normal and providing them greater opportunity to beo m e trapped in the lung and subsequently ompliate and aelerate vasulopathy in the mirovasulature of this model of PH. Supported by Department of Veterans Affairs. Referenes I. JEFFERY, T. K. and WANSTALL, J.., Pulmonary vas ul ar remodeling: a target for t.herapeuti intervention in pu lmonary hypertension, Pha rmaol Ther, I, 1-20, TUDER, R. M., et ai., The pat.hobiology of pulmonary hypertension. EndotheliUl11, Clin Chest Med, 3, , 200 I. 3. MOLTHEN, R., et ai., Blood flow modu lation indues differenes in models of pultnonary vasul ar remodeling in the rat, Proeed in gs of the Amerian Thorai Soiety (PATS), A687, L1, X., et ai., Differential effets of apol ipoprotein A -I-mimeti peptide on evol v ing and established atheroslerosis in apolipoprotein E-null mie, Cirulation, 12, , NA VAB, M., et ai., Human apolipoprotein A-I and A-I mimeti peptides: potenti al for atheroslerosis reversa l, Curr Opin Lipidol, 6, 645-9, SHAH, P. K., Aute HDLlApo A I Infusion Therapy for Remodeling/Regression of A theroserolsis: A promising New Treatment Paradigm, Inadian Heart J, , FORRESTER, J. S. and SHAH, P. K., Emerging strategies for inreasing highde nsity lipoprotei n, Am j Card iol, I I, , OU, Z., et ai., L-4F, an apolipoprolein A-I mi'l1eti, restores nitri oxide and suproxide anion balane in low-density lipoprotein-lreated endothelial ells, Cirulation, I I, MOLTHEN, R.., t ai., Quantitative models of the rat pulmonary arterial tree morphomet.ry appl ied to hypoxia-indued arteri al remodeling, j Appl Physiol, 6, ; disussion 2354, AZIZ, S. M., et ai., Oxidative stres mediates monorotaline-indued alterations in tenas in expression in pulmonary artery endothelial ells, Int j Biohem Cell Bioi, 5, , 1997.
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