Nitro-Oxidative Stress Biomarkers in Active and Inactive Men

Transcription

1 University of Msschusetts Amherst From the SelectedWorks of Srh Witkowski 212 Nitro-Oxidtive Stress Biomrkers in nd Inctive Men L. Bjork N. T. Jenkins, University of Mrylnd Srh Witkowski, University of Msschusetts - Amherst J. M. Hgerg, University of Mrylnd Aville t:

2 Physiology & Biochemistry 279 Nitro-Oxidtive Stress Biomrkers in nd Inctive Men Authors L. Bjork 2, N. T. Jenkins 2, S. Witkowski 1, J. M. Hgerg 2 Affilitions 1 University of Msschusetts, Kinesiology, Amherst, United Sttes 2 University of Mrylnd, Deprtment of Kinesiology, College Prk, United Sttes Key words exercise trining nitric oxide rective oxygen species ging ccepted fter revision Jnury 5, 212 Biliogrphy DOI /s Pulished online: Ferury 29, 212 Int J Sports Med 212; 33: Georg Thieme Verlg KG Stuttgrt New York ISSN Correspondence Dr. Jmes Hgerg University of Mrylnd Deprtment of Kinesiology 255 Vlley Drive College Prk United Sttes Tel.: +1/31/ Fx: +1/31/ hgerg@umd.edu Astrct Oxidtive stress mrkers re novel fctors shown to e relted to crdiovsculr (CVD) risk. We exmined the effects of long-term exercise, ge, nd their interction on plsm oxidized LDL (ox- LDL), nitrotyrosine, nd myeloperoxidse (MPO) levels, ll iomrkers of oxidtive stress, nd determined their ssocition with plsm nitric oxide (NOx) levels s n index of NO iovilility. Older (62 ± 2 yr) ctive men (n = 12) who hd exercised for > 3 yers nd young (25 ± 4 yr) ctive men (n = 7) who hd exercised for > 3 yers were ge- nd BMI-mtched to older (n = 11) nd young (n = 8) inctive men. Young sujects hd Introduction Crdiovsculr disese (CVD) is the leding cuse of deth in the developed world, nd conventionl risk fctors such s hypertension nd dyslipidemi hve long een used to estimte n individul s CVD risk. However, in the lte 199 s, growing evidence suggested tht these conventionl risk fctors might only explin hlf of CVD cses [ 1 ]. In ddition, only out 6 % of the reduction in CVD risk resulting from regulr exercise could e ttriuted to trining-induced improvements in conventionl CVD risk fctors [ 26 ]. Among the most promising novel risk fctors recently linked to CVD re oxidtive stress mrkers. Oxidtive stress results from n imlnce of oxidnts nd ntioxidnts tht results in excess rective oxygen species (ROS) production. The excess ROS cn dmge lipids, proteins, nd nucleic cids, nd reserch suggests tht oxidtive stress my contriute to the development of vrious pthologies including ging, dementi, nd therosclerosis [28 ]. Reserch hs linked plsm oxidized LDL (ox- LDL), nitrotyrosine, nd myeloperoxidse (MPO) levels to CVD [12, 17, 32 ]. Ox-LDL produced y lower plsm nitrotyrosine levels thn older sujects (P =.47). Young inctive sujects hd higher ox-ldl levels thn either the young ctive (P =.42) or the older ctive (P =.41) sujects. In ddition, plsm oxidtive stress levels, prticulrly ox-ldl, were correlted with vrious conventionl plsm lipoprotein-lipid levels, nd in older sujects were ssocited with Frminghm risk score (r =.49, P =.15). We found no reltionships etween plsm oxidtive stress mrkers nd NOx levels. The findings suggest tht sedentry lifestyle my e ssocited with higher ox-ldl levels nd tht the levels of oxidtive stress mrkers re relted to levels of other conventionl CVD risk fctors nd overll CVD risk. the oxidtion of LDL y ROS migrtes into the suendothelil spce, is tken up y mcrophges, nd promotes fom cell formtion [ 3 ]. Ox-LDL lso promotes endothelil dysfunction, vsculr remodeling, plque rupture, nd thromosis [35 ]. Plsm levels of ox-ldl hve een shown to predict future CVD events [22 ]. Nitrotyrosine is mrker of protein nitrtion tht hs een ssocited with inflmmtion nd endothelil dysfunction [3 ]. Elevted nitrotyrosine levels hve een found in therosclerotic lesions, nd incresed nitrotyrosine levels re ssocited with vrious CV disese sttes [ 18, 34 ]. The enzyme MPO ctlyzes the formtion of severl oxidnts tht re importnt to the oxidtive modifictions ssocited with therosclerosis [2, 29 ]. Elevted plsm levels of MPO hve een ssocited with CVD [38 ]. While these iomrkers hve een shown to e involved in endothelil dysfunction nd the process of therosclerosis, little is known out their levels in helthy, pre-clinicl popultion. The role of physicl ctivity in reducing CVD risk y improving conventionl risk fctors nd the ge-relted increse in CVD risk is well estlished. However, less is known out the effect of Downloded y: University of Msschusetts - Amherst. Copyrighted mteril. Bjork L et l. Circulting iomrkers of nitro-oxidtive stress Int J Sports Med 212; 33:

3 28 Physiology & Biochemistry long-term physicl ctivity nd the potentil interctive effects of exercise trining nd ge on oxidtive stress iomrkers. Previous studies indicte tht short-term exercise trining (1 18 wks) reduces plsm levels of oxidtive stress mrkers [9, 19, 27 ], ut the effects of long-term exercise trining ( > 26 wks) or ge on these mrkers hve not een investigted. Also, while endothelil dysfunction resulting from elevted ROS levels hs een implicted in CVD [35 ], no studies hve exmined whether the effects of trining or ge on oxidtive stress levels re relted to chnges in nitric oxide (NO) iovilility, indexed in the present study s the plsm levels of oth nitrites nd nitrtes (NOx). We sought to exmine the effects of long-term exercise trining, ge, nd their interction on plsm levels of ox-ldl, nitrotyrosine, MPO, nd NOx. We hypothesized tht plsm levels of ox- LDL, nitrotyrosine, nd MPO would e lower, nd NOx levels higher, in ctive individuls compred to their inctive peers nd in the young compred to the older groups. Methods All recruiting nd screening methods for the sujects in this study were descried previously [14, 15, 37 ]. Sujects were helthy, nonsmoking men with no history of CVD or dietes. Older (62 ± 2 yr) ctive men (n = 12) who hd performed moderte- to high-intensity endurnce exercise for > 4 h/wk for > 3 yers were BMI- nd ge-mtched to inctive men (n = 11). Young (25 ± 4 yers) ctive men (n = 7) who hd performed moderte- to high-intensity endurnce exercise for > 4 h/wk for > 3 yers were lso mtched on the sis of BMI nd ge to inctive men (n = 8). The inctive groups consisted of len, helthy men who hd prticipted in physicl ctivity < 2/wk for < 2 min/ session for > 5 yers with sedentry occuption or retired. All prticipnts provided written informed consent nd ll procedures were pproved y the University of Mrylnd College Prk Institutionl Review Bord nd the study meets the ethicl stndrds of this journl [ 11 ]. Mximl oxygen consumption (VO 2mx ) nd ody composition were mesured s descried previously [15, 37 ] with ll devices clirted prior to usge nd with lortory environmentl conditions stndrdized nd recorded dily. Peripherl venous lood ws smpled in the morning efore 9. m. fter n overnight fst. All sujects voided lcohol, vitmins, cffeine, nd medictions for 24 h efore testing. Sujects were ingesting their hitul diet t the time of testing. Young nd older sujects did not tke ny medictions, vitmins, or ntioxidnt supplementtions for 24 nd 48 h prior to lood smpling, respectively. For ctive sujects lood smpling occurred h fter one of the suject s usul exercise sessions. Plsm lipoprotein-lipid profiles nd fsting glucose levels were mesured (Quest Dignostics, Bltimore, MD). To clculte the suject s overll CVD risk, the conventionl CVD risk fctors tht were mesured were pplied to the equtions sed on the Frminghm study [ 36 ]. Plsm ox-ldl levels in the older sujects were pulished previously nd were mesured with commercilly-ville competitive Enzyme-Linked Immunosorent Assy (ELISA) kit (Mercodi, Uppsl, Sweden) [ 37 ]. The present study used the sme ssy to mesure ox-ldl in the young sujects. In oth ox- LDL ssy runs, 2-level control smples were used to confirm ssy performnce. Previously, ll smples in the older group were nlyzed in single ssy, nd in the present study ll smples from young sujects were nlyzed in single ssy. Thus, within ech group the inter-ssy vriility ws eliminted. All smples were mesured in duplicte. The intr-ssy coefficients of vrition were 6.8 [ 37 ] nd 9.6 % for the older nd young groups, respectively. Nitrotyrosine ws mesured using commercilly-ville ELISA kit (Cell Sciences, Cnton, MA). The kit is solid-phse ELISA sed on the sndwich principle tht detects nitrotyrosine-contining proteins in plsm. For nitrotyrosine, MPO, nd NOx, ll smples were nlyzed in duplicte or triplicte in single ssy to void inter-ssy vriility. The verge intr-ssy coefficient of vrition for nitrotyrosine ws 9.1 %. MPO ws mesured with the Humn MPO ELISA kit (Cell Sciences, Cnton, MA). The intr-ssy coefficient of vrition for MPO ws 5. %. Plsm NOx levels were mesured using Nitrte/Nitrite Colorimetric Assy Kit (Cymn Chemicl Compny, Ann Aror, MI). The intr- nd inter-ssy coefficients of vrition were 7.7 nd 13.2 %, respectively. 2-wy ANOVAs were run with SPSS softwre to exmine the min effects of long-term exercise trining nd ge, long with the interction effect of physicl ctivity nd ge, on plsm levels of ox-ldl, nitrotyrosine, MPO, nd NOx. The normlity of ll vriles ws verified prior to performing sttisticl nlyses. Multiple comprisons etween study groups were nlyzed with Fisher s Lest Significnt Difference method. One-sided p-vlues re presented for tests of priori directionl hypotheses, unless group mens were opposite the direction hypothesized, in which cse 2-tiled p-vlues re presented. Regression nlysis ws performed to determine whether there were ny significnt reltionships etween levels of NOx nd levels of the selected oxidtive stress mrkers. In ddition, Person correltion coefficients were used to ssess reltionships mong study vriles. An α vlue of.5 ws used to indicte sttisticl significnce. Dt re presented s men ± SE. Results nd inctive sujects were successfully mtched for ge nd BMI in oth the young nd the older groups ( Tle 1 ). However, s expected, ctive sujects hd significntly higher Tle 1 Suject Chrcteristics. Young Older Chrcteristic (n = 7) Inctive (n = 8) (n = 12) Inctive (n = 11) ge, yr 25 ± 2 25 ± 1 62 ± 2 64 ± 2 height, m 1.83 ± ± ± ±.1 Weight, kg 81.1 ± ± ± ± BMI, kg/m 24.4 ± ± ± ±.6 ody Ft, % 14.3 ± ± ± ± 1.7 * glucose, mg/dl 86 ± 3 81 ± 3 94 ± 2 11 ± 3 TG, mg/dl 69 ± 7 81 ± ± 8 13 ± 14 * TC, mg/dl 146 ± ± ± ± 11 HDL, mg/dl 53 ± 2 49 ± 4 71 ± 4 51 ± 4 * LDL, mg/dl 79 ± 8 82 ± ± ± 11 TC/HDL 2.8 ± ± ± ±.5 * SBP, mmhg 118 ± ± ± ± 3 DBP, mmhg 78 ± 2 79 ± 2 81 ± 2 86 ± 2 VO 2mx, ml/kg/min 6.4 ± ± ± ± 1.7* Vlues re mens ± SE. BMI, ody mss index; TG, triglycerides; TC, totl cholesterol; HDL, high-density lipoprotein cholesterol; LDL, low-density lipoprotein; SBP, systolic lood pressure; DBP, distolic lood pressure, VO 2mx, mximl oxygen uptke. P <.5 vs. young ctive. * P <.5 vs. older ctive Downloded y: University of Msschusetts - Amherst. Copyrighted mteril. Bjork L et l. Circulting iomrkers of nitro-oxidtive stress Int J Sports Med 212; 33:

4 Physiology & Biochemistry 281 VO 2mx vlues. In ddition, older ctive sujects hd significntly lower ody ft nd significntly etter plsm lipoprotein-lipid profiles, i. e., less therogenic, thn the older inctive sujects. There ws significnt min effect of long-term exercise trining (P =.5), ut not ge, on plsm ox-ldl levels ( Fig. 1 ). Multiple comprisons reveled tht the young inctive group hd significntly higher ox-ldl levels thn either the young ctive (P =.42) or older ctive (P =.42) groups ( Tle 2 ). Among the significnt reltionships etween plsm ox-ldl levels nd conventionl CVD risk fctors were significnt positive correltion etween ox-ldl levels nd LDL cholesterol, nd significnt inverse correltion etween plsm ox-ldl nd HDL cholesterol cross ll study sujects ( Fig. 2 ). There ws significnt min effect of ge on plsm nitrotyrosine levels (P =.47), with older individuls hving significntly higher levels ( Fig. 3 ). There ws no significnt min effect of long-term exercise trining on plsm nitrotyrosine levels nd no significnt interction effect. Multiple comprisons showed no etween-group differences in plsm nitrotyrosine levels ( Tle 2 ). There were no significnt min effects of ge or long-term exercise trining on plsm MPO levels nd no significnt interction effect. Between-group nlyses showed tht the young ctive group hd significntly higher plsm MPO levels (P =.12) compred to the older inctive group ( Tle 2 ). There were no significnt min effects of ge or long-term exercise trining on plsm NOx levels nd no significnt interction effect. Tests of multiple comprisons reveled tht the older inctive group hd significntly higher plsm NOx levels thn either the older ctive (P =.21) or the young ctive (P =.2) groups ( Tle 2 ). There were no significnt correltions etween plsm NOx nd ox-ldl, nitrotyrosine, or MPO levels. Ox-LDL (U/L) * Inctive Fig. 1 Min effect of long-term exercise trining on plsm ox-ldl level etween ctive (n = 19) nd inctive (n = 19) men. Vlues re mens ± SE. *P <.5 vs. young. We lso exmined reltionships etween outcome vriles nd conventionl CVD risk fctors seprtely within the young nd older ge groups. In young sujects, plsm ox-ldl levels were negtively correlted (r =.64, P <.5) with VO 2mx. Significnt positive reltionships etween plsm ox-ldl levels nd the Frminghm risk score nd risk percentge were oserved in older sujects ( Fig. 4 ). When reltionships were nlyzed seprtely y ctivity level groups, plsm MPO levels were positively correlted (r =.51, P <.5) with the ox-ldl to LDL cholesterol rtio in inctive sujects. Discussion Our results suggest tht plsm oxidtive stress mrker levels re ffected y long-term exercise trining nd ge, ut not consistently in the hypothesized direction. The min findings re tht () plsm ox-ldl levels re lower in long-term exercisers, () nitrotyrosine levels re higher in older individuls, (c) MPO levels re lowest nd plsm NOx levels highest in older inctive individuls, (d) oxidtive stress mrker levels re relted to vrious conventionl CVD risk fctors, nd (e) NOx levels re not relted to plsm oxidtive stress iomrker levels. The reltionships etween oxidtive stress mrkers nd other CVD risk fctors suggest tht, t the lest, plsm oxidtive stress mrker levels my serve s indictors of CVD risk. However, the lck of ssocition etween plsm levels of oxidtive stress mrkers nd NOx does not support the hypothesis tht elevtions in oxidtive stress led to incresed CVD risk y reducing NO iovilility. It hs een proposed tht mny pthologic chnges ssocited with ging cn e explined y the irreversile ccumultion of moleculr oxidtive dmge [33 ]. Therefore, one would expect older individuls to hve higher levels of oxidtive stress thn young individuls. In the present study, however, only nitrotyrosine levels were elevted in the older compred to the young sujects. Our findings suggest tht plsm ox-ldl nd MPO my not chnge with ge in the sme mnner s nitrotyrosine. Thus, perhps not ll ROS mrkers contriute to the increse in oxidtive dmge tht hs een hypothesized to occur with ge. However, our study is the first to compre plsm levels of the selected oxidtive stress mrkers cross ge groups, so it is difficult to mke definitive conclusions out how the chosen oxidtive stress iomrkers re ffected y ge. Although cute exercise results in incresed ROS genertion nd elevted oxidtive stress [ 4 ], some previous studies hve found tht exercise trining my reduce systemic levels of oxidtive stress [9, 27 ]. In the present study we found tht ctive individuls hd lower plsm ox-ldl levels thn their sedentry peers. This confirms previous results of reduced plsm ox-ldl levels in older men nd women [ 5 ] nd sedentry, helthy young men Downloded y: University of Msschusetts - Amherst. Copyrighted mteril. Young Older Outcome (n = 7) Inctive (n = 8) (n = 12) Inctive (n = 11) Ox-LDL (U/L) 59.7 ± ± ± ± 4. nitrotyrosine (nm) 41.1 ± ± ± ± MPO (ng/ml) 73. ± ± ± ± 1.4 NOx (μm) 14.9 ± ± ± ± 2. Vlues re mens ± SE. Ox-LDL, oxidized low-density lipoprotein; MPO, myeloperoxidse; NOx, nitrtes/nitrites. Dt with like letters re not sttisticlly different from ech other (P <.5) Tle 2 Plsm Oxidtive Stress Biomrker Levels. Bjork L et l. Circulting iomrkers of nitro-oxidtive stress Int J Sports Med 212; 33:

5 282 Physiology & Biochemistry r =.325 P =.46 2 r =.467 P =.3 Nitrotyrosine (nm) LDL (mg/dl) HDL (mg/dl) Older Inctive Young Inctive Older Young Older Inctive Young Inctive Older Young Fig. 2 Sig n ificnt reltionships (P <.5) cross ll sujects etween plsm levels of ox-ldl nd the conventionl CVD risk fctors LDL cholesterol nd HDL cholesterol ; smple sizes were s follows: older ctive men (n = 12), older inctive men (n = 11), young ctive men (n = 7), nd young inctive men (n = 8). Young nd women [ 8 ] following n exercise intervention. Thus, our lower ox-ldl levels with long-term trining re consistent with these findings. Also, our inverse correltion etween ox-ldl levels nd VO 2mx in young sujects further supports the hypothesis tht regulr physicl ctivity reduces oxidtive stress. We did not find lower nitrotyrosine or MPO levels in the ctive compred to the inctive groups. This finding ws somewht unexpected, s one erlier study found reduction in plsm nitrotyrosine levels with 16-wk exercise intervention in older sujects [ 9 ]. Another study reported decrese in serum MPO levels with 12-wks of endurnce trining in sujects with elevted CVD risk [ 31 ]. Although the present nitrotyrosine nd MPO results do not gree with these previously reported findings, they re not without precedent. One study concluded tht * Older Fig. 3 Min effect of ge on plsm nitrotyrosine levels etween young (n = 15) nd older (n = 23) men. Vlues re mens ± SE. * P <.5 vs. young. older dults who exercise regulrly hve higher levels of systemic oxidtive dmge thn their sedentry peers [ 23 ]. The generl lck of informtion in the literture descriing plsm ox-ldl, nitrotyrosine, nd MPO chnges with exercise trining prevents definitive conclusions from eing drwn reltive to how chronic exercise ffects systemic levels of oxidtive stress. The present findings tht nitrotyrosine nd MPO levels were not lower in ctive compred to the inctive groups could e result of the unique fetures of the present study. First, the present study exmined the effects of long-term exercise trining on oxidtive stress levels, while ll previous studies hve focused on the effects of reltively short-term trining. Second, the inctive sujects in the present study re unique ecuse they hve no redily pprent helth issues nd pper to hve suffered no negtive consequences from their yers of physicl inctivity. Despite their sedentry lifestyles, the young inctive men hd ody weight, BMI, ody composition, lood pressure, nd plsm lipoprotein-lipid vlues similr to those of their ctive peers who hd een exercising regulrly for > 3 yers. The older inctive sujects hd ody weight, BMI, nd lood pressure similr to those of their ctive peers who hd een exercising for > 3 yers. This mkes the individuls in the young nd especilly the older inctive groups quite exceptionl. We found no min effects of ge or ctivity level on plsm NOx levels, lthough there ws some evidence tht the inctive, nd especilly the older inctive, groups hd higher plsm NOx levels thn their trined peers. These results contrst with those from previous studies showing tht NOx levels decline with ge nd tht these declines cn e reversed with short-term exercise trining [7, 21 ]. However, the responses to longer-term trining ( 16 wk) seem to e more vrile. In group of older men nd women, 24 wk of exercise trining did not improve plsm NOx levels [ 1 ]. Given the extremely long durtion of trining in the Downloded y: University of Msschusetts - Amherst. Copyrighted mteril. Bjork L et l. Circulting iomrkers of nitro-oxidtive stress Int J Sports Med 212; 33:

6 Physiology & Biochemistry Inctive Frminghm Risk Score Frminghm Risk Percentge Inctive r =.49 P =.15 present study, the lck of ssocition etween ctivity level nd plsm NOx levels my e less surprising. We found no reltionships etween plsm NOx levels nd plsm ox-ldl, nitrotyrosine, or MPO levels. These findings pper to contrst with previous reserch tht hs linked increses in oxidtive stress to endothelil dysfunction [ 35 ]. However, our plsm NOx mesurement is n ssessment of NO iovilility s opposed to mesurement of endothelil function. Although reduced NO iovilility is one mechnism thought to cuse endothelil dysfunction [1 ], impirments in endothelil function cn occur without decreses in NOx levels. For exmple, n erlier study found tht therosclerotic rits hd higher plsm NO levels, s determined y quntifying nitrosyl compounds in lood, ut impired endothelium-dependent vsodiltion, compred to control rits [ 25 ]. Thus, plsm NOx levels re not necessrily directly relted to endothelil function, nd the present study only llows one to mke conclusions out how NO iovilility is ffected y ge or inctivity. However, the lck of ssocition etween NO iovilility nd plsm oxidtive stress mrker levels in the present study is still surprising. Previous reserch hs shown tht ox-ldl reduces NO iovilility y inctivting NO nd stimulting the relese of 1 r =.42 P =.41 Fig. 4 Sig n ificnt reltionships (P <.5) etween plsm ox-ldl levels nd Frminghm risk score nd Frminghm risk percentge in the ctive (n = 12) nd inctive (n = 11) older sujects. NO scvengers [16 ]. Elevted nitrotyrosine levels hve een ssocited with incresed NO rekdown nd inhiited NO synthesis [ 3 ]. MPO hs previously een shown to decrese NO levels y inhiiting nd uncoupling NOS, nd reking down NO [ 29 ]. 2 possile explntions for why the present study filed to find ny ssocitions etween plsm ox-ldl, nitrotyrosine, nd MPO levels nd plsm NOx levels hve lredy een discussed. Briefly, the prticulrly long durtion of trining undertken y the ctive sujects in this study my hve resulted in different effects on plsm NOx levels thn short-term intervention. Also, the fct tht the inctive sujects in our study were generlly very helthy my indicte tht these iomrkers re not ffected yet in this pre-clinicl popultion. We found severl significnt reltionships etween ox-ldl levels nd conventionl CVD risk fctors. Across ll sujects, plsm ox-ldl ws positively correlted with LDL cholesterol levels nd negtively correlted with HDL cholesterol levels. Such reltionships hve een previously reported [12, 13 ]. The reltionship etween ox-ldl nd LDL cholesterol mkes intuitive sense ecuse ox-ldl molecules re formed y the oxidtive modifiction of ntive LDL, with n increse in LDL likely to e ssocited with n increse in ox-ldl. Also, the negtive ssocition etween ox-ldl nd HDL levels fits with evidence suggesting tht HDL molecules re theroprotective, in prt, ecuse they inhiit LDL oxidtion [ 24 ]. In ddition, within the older sujects ox-ldl levels were positively correlted with Frminghm risk score nd percentge, suggesting tht ox-ldl could hve predictive vlue for CVD, especilly in those t higher risk for CVD. These results re supported y study tht found tht plsm ox-ldl levels were predictive of future CVD events, nd they were stronger predictors thn the conventionl lipoprotein profile, in pprently helthy men [22 ]. In the inctive sujects, positive reltionship ws oserved etween plsm MPO levels nd the rtio of ox-ldl to LDL cholesterol. Previous reserch hs indicted tht MPO cn convert ntive LDL to ox-ldl [ 6 ]. In ddition, one of the secondry oxidtion products generted y MPO, nitrogen dioxide, hs een reported to promote ox-ldl formtion [ 2 ]. Thus, one would expect n increse in MPO to led to n increse in ox-ldl formtion, resulting in the incresed ox-ldl to LDL rtio reported in the present study. The present study is limited y the cross-sectionl design employed to ssess potentil iomrker differences cross ge nd hitul physicl ctivity level groups. This, comined with the reltively smll smple sizes we utilized, hs generted results tht, while hrdly definitive, t the lest provide sustntive frmework for future longitudinl exercise trining intervention studies in lrger popultions. Also, our findings re limited to plsm iomrker levels, keeping in mind tht mesuring the loclized oxidtive stress more directly in tissues or cell comprtments my yield different results. In ddition, the ssessment of plsm levels of oxidtive stress mrkers in the present study my hve enefited from the mesurement of dditionl plsm oxidtive stress mrkers, such s mlondildehyde nd thiorituric cid rection sustnces. And finlly, we hve no mechnistic dt to determine if ny of these oserved reltionships or differences cross groups cn e ttriuted to specific moleculr nd cellulr mechnisms. Such mechnistic dt could include those generted y longitudinl exercise trining, dietry, nd nti-oxidnt studies, leled plsm iomrker turnover studies, nd studies to ssess the Downloded y: University of Msschusetts - Amherst. Copyrighted mteril. Bjork L et l. Circulting iomrkers of nitro-oxidtive stress Int J Sports Med 212; 33:

7 284 Physiology & Biochemistry relese of these iomrkers cross specific tissue eds such s dipose tissue nd skeletl muscle. In conclusion, our findings suggest tht ge nd chronic exercise trining re ssocited with different levels of circulting iomrkers of oxidtive stress. It ppers tht sedentry lifestyle my e ssocited with elevted ox-ldl levels, nd the results indicte tht lower plsm ox-ldl levels in trined individuls re relted to etter CVD risk fctor profiles nd lower overll CVD risk. In ddition, the results did not show link etween plsm oxidtive stress nd plsm NOx levels, underscoring the need for further reserch to elucidte how elevtions in oxidtive stress contriute to increses in CVD risk. Acknowledgements This work ws supported y NIH Predoctorl Trining Grnt T32AG268 (NTJ, SW, JMH) nd the University of Mrylnd Deprtment of Kinesiology Grdute Student Reserch Inititive Fund (NTJ, LB). References 1 Brinkley T E, Fenty-Stewrt NM, Prk JY, Brown MD, Hgerg JM. 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