
 fat redistribution in hiv - infected patients is associated with antiretroviral therapy ( art ) , including protease inhibitors ( pi ) and nonnucleoside reverse transcriptase inhibitors ( nnrti ) [ 1 , 2 ] .
central fat accumulation or lipohypertrophy may be more common in women [ 3 , 4 ] and has been associated with multiple metabolic abnormalities and inflammation in hiv - infected persons on art [ 59 ] .
chronic hiv infection is also associated with persistent inflammation , and treating hiv infection improved endothelial function in treatment - nave subjects with low cardiovascular disease ( cvd ) risk . given the complex interactions between chronic hiv infection and art and the likelihood that traditional framingham prediction may underestimate cardiovascular risk in hiv - infected persons on art [ 12 , 13 ] , novel biomarkers are needed to assess metabolic risk in hiv infection and response to interventions .
eicosanoids are endogenous products of arachidonic acid metabolism involved in oxidant stress , inflammation , and endothelial function , all of which are important in atherosclerosis and cardiovascular disease pathogenesis .
biologic properties and metabolism of eicosanoids are complex and are reviewed elsewhere [ 1517 ] .
briefly , during cellular stress , membrane phospholipids containing arachidonic acid are subjected to nonenzymatic peroxidation by free radical species to generate a variety of biologically active oxidation products , including f2-isoprostanes ( f2-isops ) which can cause vasoconstriction , platelet aggregation , and oxidative tissue damage .
arachidonic acid can also be released from membrane phospholipids and metabolized by oxidizing enzymes during cellular stress .
metabolism by cyclooxygenase ( cox ) enzymes yields a family of products termed prostaglandins ( pgs ) , including pge2 , which causes vasodilation or vasoconstriction and/or vascular smooth muscle proliferation ; thromboxane a2 ( txa2 ) , which causes vasoconstriction , platelet activation , and chemotaxis ; and prostacyclin ( pgi2 ) which causes vasodilation and inhibits platelet aggregation and vascular smooth muscle proliferation . with the exception of f2-isops , parent eicosanoids are unstable in and
prostaglandin metabolites , as well as f2-isops , are stable in urine and accurate indices of endogenous production [ 1821 ] .
the primary pge2 urinary metabolite is 11--hydroxy-9,15-dioxo-2,3,4,5-tetranor - prostane-1,20-dioic acid ( pge - m ) , while the major metabolites used to assess txa2 and pgi2 production are 11-dehydro - thromboxane b2 ( txb2 ) and 2,3-dinor-6-keto - pgf1 ( pgi - m ) , respectively .
urinary assays for pge - m and f2-isop have low intra - individual variation over one year . in hiv - negative populations , urinary f2-isop correlates with traditional cvd risk factors   and surrogate measures of cvd
including brachial artery flow - mediated dilation and carotid intima media thickness [ 24 , 25 ] .
in obese children and adolescents , plasma f2-isop was positively correlated with visceral adipose tissue ( vat ) , a marker of cvd risk .
cigarette smoking is also associated with higher eicosanoids [ 2730 ] . in hiv - infected persons , cross - sectional studies to date have identified associations between higher f2-isop and lipoatrophy , lactic acidosis , virologic suppression on art , heavy smoking , higher body mass index ( bmi ) and waist circumference , elevated liver transaminases and hepatitis c virus ( hcv ) rna , and female sex [ 3135 ] .
early analyses suggested higher f2-isop in persons receiving efavirenz or zidovudine , and lower levels in those on nevirapine compared with other art   but consistent associations with specific art drugs or classes have not been seen .
it is not yet clear why f2-isop levels are consistently higher in women ( hiv - infected and uninfected ) than in men .
a recent study of women in haiti found higher levels of cervical cox-2 and urinary pge - m in hiv - infected women than in uninfected women and a positive correlation between systemic pge - m and both plasma hiv rna and cervical cox-2 levels .
raltegravir ( ral ) is an hiv-1 integrase inhibitor that has not been associated with metabolic perturbations or fat redistribution during short- or long - term therapy [ 3739 ] . a randomized
, open label study was designed to assess the effects of switching from pi- or nnrti - based art to a ral - based regimen in women with lipohypertrophy and suppressed hiv-1 rna on stable therapy .
adipose tissue volumes by computerized tomography ( ct ) , anthropometrics , and fasting metabolic parameters were performed .
the objectives of these secondary analyses were to determine ( a ) the 24-week change in f2-isop and other urinary eicosanoid metabolites and ( b ) correlations between 24-week changes in f2-isop , other urinary eicosanoid metabolites , and changes in vat , the primary outcome of the parent study .
we hypothesized that urinary f2-isop and other eicosanoid metabolites would decrease after 24 weeks in women switching to ral compared to those continuing a pi / nnrti and that these decreases would correlate with decreased vat .
briefly , women with hiv-1 rna < 50 copies / ml , stable art including two nrtis ( tenofovir or abacavir and emtricitabine or lamivudine ) plus a pi or nnrti , and central adiposity ( waist circumference > 94  cm or waist : hip > 0.88 ) were enrolled at five centers in north america from september 2008 to july 2010 and randomized 1  :  1 to switch their pi / nnrti to open label ral 400  mg twice daily ( ral arm ) or continue to present art for 24 weeks ( pi / nnrti arm ) .
relevant exclusion criteria included current use of metformin , thiazolidinediones , or androgen therapy , use of growth hormone or growth hormone - releasing factor in the six months prior to screening , change or initiation of lipid - lowering therapy in the three months prior to screening , and intent to significantly modify diet or exercise habits during the study .
the primary endpoint of the parent trial was between - group change in percent of vat volume 24 weeks following a switch to ral versus continued pi or nnrti .
all study procedures were approved by the institutional review boards of the participating institutions , and all subjects provided informed consent prior to initiation of study procedures .
procedures were performed in accordance with the ethical standards of the responsible committee on human experimentation and with the helsinki declaration of the world medical association .
scans were performed locally but standardized and read centrally by a blinded reader at the tufts university body composition center .
waist , hip , and neck circumferences were performed according to aids clinical trials group standards   at weeks 0 , 12 , and 24 .
fasting ( > 8 hours ) glucose , lipoprotein profile , high - sensitivity c reactive protein ( hscrp ) , and cd4 + t cell counts were assessed at weeks 0 , 12 , and 24 .
hiv-1 rna ( 50 copies / ml assay sensitivity ) was measured at screening and weeks 4 , 8 , 12 , and 24 .
labs were performed at the individual sites in real - time and according to local standards .
clean - catch urine samples were collected , and three - milliliter ( ml ) aliquots of urine were stored at 80c until analysis .
samples were shipped overnight on dry ice to the vanderbilt university eicosanoid core laboratory where analyses were performed .
urinary f2-isop , txb2 , and pgi - m were measured using gas chromatography - negative ion chemical ionization mass spectrometry employing stable isotope dilution methodology , as described elsewhere [ 15 , 18 , 20 , 42 ] .
pge - m was measured by liquid chromatography - mass spectroscopy ( lc - ms ) , as previously described .
results for all urinary metabolites are presented as ng / mg urinary creatinine ( cr ) . at one study site ,
urinary eicosanoid results from these samples were not statistically different from the other sites , and analyses with and without data from these samples were performed ( data not shown ) . as there were no substantive changes in the results , we report results including data from all sites . baseline characteristics of the two randomization groups
were compared using the mann - whitney u test for continuous variables and fisher 's exact test for categorical variables .
median values and interquartile ranges ( iqr ) are reported for continuous variables , and percentages are reported for categorical data .
comparison of median between - group 24-week change scores for eicosanoids was performed using the wilcoxon signed - rank test .
the primary analysis was as - treated , excluding subjects who did not remain on the study regimen and/or did not have an observed primary endpoint .
generalized linear models assessed associations between eicosanoid changes and study arm , adjusting for baseline pi use , bmi , smoking status , study site , and age ( 50 versus < 50 years ) .
all statistical tests were two - sided with a nominal p level of 0.05 . given the exploratory nature of these analyses , we did not adjust results for multiple testing .
data analysis and management was performed using sas 9.2 ( sas institute , inc . , cary , nc , usa ) .
eighteen subjects were randomized to the ral arm and 21 to continue pi / nnrti .
one subject from each arm withdrew for reasons unrelated to the study intervention , leaving 37 subjects who completed the week 24 primary endpoint .
complete demographic and baseline clinical characteristics of the 37 participants included in the as - treated analysis are provided in table 1 . at baseline ,
the study groups were well balanced , with the exception of the pi / nnrti arm having a higher rate of current smoking ( 60% versus 24% ; p = 0.045 ) .
the median age was 43 years , bmi 32  kg / m , and 75% of subjects self - identified as black or hispanic .
sixty - two percent of subjects were on a pi at entry ( versus 38% nnrti ) , and the most commonly prescribed nrti was tenofovir ( 78% ) .
baseline median ( iqr ) urinary f2-isop , pge - m , pgi - m , and txb2 ( ng / mg cr ) were 2.14 ( 1.493.16 ) , 8.07 ( 4.4710.56 ) , 0.10 ( 0.060.15 ) , and 0.46 ( 0.250.73 ) , respectively ( table 1 ) . when comparing study arms , baselines pgi - m , pge - m , and txb2 were all lower , and f2-isop was higher in the ral arm ( table 1 ) , but only pgi - m was statistically different ( p = 0.005 ) .
baseline pge - m tended to be higher in current smokers ( p = 0.1 ; data not shown ) .
eicosanoid levels also tended to differ by baseline nrti , with women receiving abacavir having consistently lower levels ( figure 1 ) , including statistically significantly lower f2-isop ( p = 0.05 ; figure 1(a ) ) and txb2 ( p = 0.04 ; figure 1(d ) ) levels .
pge - m was positively correlated with age ( rho = 0.34 ; p = 0.04 ) and negatively correlated with body weight ( rho = 0.35 ; p = 0.03 ) .
txb2 was positively correlated with vat : sat and vat : total adipose tissue and negatively correlated with sat , body weight , bmi , and hip and neck circumferences ( rho = 0.37 to 0.46 ; p = 0.004 to 0.02 ) .
none of the urinary eicosanoids were correlated with fasting lipids , glucose , insulin resistance , or hscrp in this study population at baseline ( data not shown ) .
median 24-week urinary eicosanoid levels and changes from baseline are shown in figure 2 and table 2 . over 24 weeks
, only pgi - m in ral - treated subjects demonstrated a statistically significant within - group change ( p = 0.04 ; figure 2(c ) ) .
txb2 increased in the ral arm and decreased in the pi / nnrti arm ( + 0.09 [ 0.04 , + 0.13 ] versus 0.02 [ 0.20 , + 0.03 ] ; figure 2(d ) ) , but this difference was of borderline statistical significance ( between - group p = 0.06 ) .
there were no other statistically significant differences between or within study arms over 24 weeks .
age  50 years at baseline was associated with an increase in pge - m ( median change + 3.9 versus 1.3 in subjects < 50 years of age ; p = 0.05 ; figure 3 ) . in the pi / nnrti arm ,
24-week vat change positively correlated with changes in pgi - m ( rho = 0.45 ; p = 0.04 ) and txb2 ( rho = 0.44 ; p = 0.05 ) , with a similar trend seen for pge - m ( rho = 0.41 ; p = 0.07 ) . among persons in the ral arm ,
the change in pge - m correlated with an increase in hdl cholesterol ( rho = 0.56 ; p = 0.02 ) ; this correlation was not observed in the pi / nnrti arm ( rho = 0.17 ; p = 0.48 ) .
changes in eicosanoid levels over 24 weeks were not statistically different by baseline nrti ( abacavir versus tenofovir ; data not shown ) .
changes in urinary eicosanoids from baseline to 24 weeks were assessed in multivariate models adjusting for study arm ( ral versus pi / nnrti ) , baseline bmi , age , pi use , smoking status , and study site .
n = 8) was associated with 24-week pge - m increase (  = 8.3 [ 95% ci 0.3 , 16.3 ] ; p = 0.04 ) , independent of the covariates above ( table 3 ) .
no baseline factors were significantly associated with changes in other urinary eicosanoids ( table 3 ) .
in these hiv - infected women with central adiposity and suppressed hiv rna on art , switching pi- or nnrti - based art to ral did not have significant effects on urinary eicosanoids over 24 weeks .
overall , f2-isop , pge - m , and txb2 levels were higher and pgi - m levels were lower than published levels reported in healthy adults [ 15 , 1820 , 43 , 44 ] .
although formal comparisons were not performed , urinary f2-isop ( lower ) , pge - m ( higher ) , and txb2 ( higher ) levels also differed from those observed in previously studied hiv - infected women who were younger and had lower bmi . of note , several of the markers differed  though not with statistical significance  at baseline between the two study arms . given known effects of smoking on these biomarkers [ 2729 ] , this difference may have been driven in part by the significantly greater number of smokers randomized to the pi / nnrti arm , and this
may therefore have limited our capacity to identify differences in changes over time or due to ral switch .
although baseline pge - m tended to be higher in current smokers ( n = 16 ; median [ iqr ] 9.9 [ 4.914.4 ] ) than nonsmokers ( n = 21 ; median [ iqr ] 7.9 [ 4.0 - 10.0 ] ; p = 0.11 ) , in a multivariate model , neither current smoking nor study arm was significantly associated with 24-week change in pge - m and did not attenuate the relationship between age and change in pge - m .
additionally , f2-isop , which is increased in hiv - infected and uninfected smokers , tended to be higher in smokers at baseline ( p = 0.06 ) but was not significantly higher in the pi / nnrti arm .
age and bmi were lower and higher in the ral than the pi / nnrti arm , respectively ( table 1 ) , and though they were not statistically different , we did include these as covariates in adjusted models . in primary analyses , women in the switch arm had significant improvements in total and ldl cholesterol but did not have a statistically significant improvement in vat compared to women continuing an nnrti or pi .
they also had a significant decrease in soluble cd14 , a marker of monocyte activation .
a recent analysis of extensively treatment experienced , predominantly male , subjects switching enfuvirtide to ral in france reported significant 24-week decreases in interleukin-6 , d - dimer , and hscrp   that were not observed in these less treatment - experienced women with central adiposity . in subjects remaining on pi / nnrti , increasing vat was marginally correlated with increasing pgi - m and pge - m over 24 weeks of follow - up , suggesting a relationship between these markers and central adiposity that was altered by a switch to ral .
older age ( 50 years ) at enrollment was associated with an increase in pge - m independent of study arm , smoking status , pi use at baseline , or other factors .
this was unexpected given associations with f2-isop and txb2 in prior cross - sectional studies of hiv - infected persons [ 33 , 34 ] , and this may be due to the inclusion of both males and females in prior analyses and/or the high prevalence of obesity and relative lack of normal bmi ranges in this study population .
this is the first study to prospectively assess the effects of an art switch on urinary eicosanoid metabolites .
the small sample size of our study and the imbalance of smokers in the pi / nnrti arm likely limited our ability to detect differences between study arms . nonetheless , intriguing trends and preliminary associations were noted .
although not routinely measured in clinical practice , eicosanoids are known markers of cardiovascular and metabolic disease risk in hiv - negative populations . in particular , f2-isop has been associated with cvd disease risk factors , hscrp , carotid intima medial thickness , coronary artery calcium , and angiographic coronary artery obstruction [ 23 , 24 , 47 ] .
pge2 is a complex mediator of inflammation and vasodilation with variable effects on vascular tone depending on the tissue and prostanoid receptor .
increased pge - m has also been associated with malignancies in hiv - negative populations [ 5053 ] , and urinary pge - m correlated with plasma hiv rna and cervical cox-2 levels in a recent study of haitian women .
abacavir exposure has been associated with increased cardiovascular risk in hiv - infected persons , but the association has not been consistent .
although potential mechanism(s ) are not clearly defined , recent studies have reported abnormal platelet reactivity with abacavir [ 56 , 57 ] . to our knowledge ,
lower f2-isop in these abacavir - treated subjects and in a prior cross - sectional analysis of men and women   suggests that if there is excess cardiovascular risk due to abacavir , it is independent of lipid peroxidation - related pathways ; prospective studies would be needed to determine this .
due to providers ' knowledge of potential cardiovascular risk with abacavir and risk of renal toxicity with tenofovir df , it is also possible that abacavir use is simply a marker of some other unmeasured factors associated with risk for these conditions and lower urinary eicosanoid levels .
in addition to the qualifications above , this analysis has other limitations that should be noted .
use of aspirin or nonsteroidal anti - inflammatory drugs ( nsaid ) was not an exclusion criterion for the parent study , and detailed information on dosing was not collected .
given the even distribution of nsaid use across study arms ( table 1 ) , we did not adjust for this variable in multivariate models and do not believe it would explain differential associations within or between groups .
neither menopausal status nor sex hormone levels were ascertained as part of this study . based on data from hiv - infected women ,
the age distribution of our population suggests the majority of subjects were pre- or perimenopausal .
the association between age and pge - m may have been due to postmenopausal changes in the older women ( 50 years ) .
platelet reactivity assays were not performed , so relationships between eicosanoids and platelet function can not be determined .
ct scan was used to assess sat , so we were unable to fully assess peripheral ( limb ) lipoatrophy .
finally , this analysis may have been too small and/or of insufficient duration of follow - up to detect meaningful changes in eicosanoids .
additional studies and longer - term follow - up in hiv - infected persons are needed to further elucidate the role of eicosanoids in metabolic and other aging - related comorbidities and determine their role as useful clinical biomarkers .
 chronic inflammation is a hallmark of hiv infection . 
 eicosanoids reflect inflammation , oxidant stress , and vascular health and vary by sex and metabolic parameters . 
 raltegravir ( ral ) is an hiv-1 integrase inhibitor that may have limited metabolic effects . 
 we assessed urinary f2-isoprostanes ( f2-isops ) , prostaglandin e2 ( pge - m ) , prostacyclin ( pgi - m ) , and thromboxane b2 ( txb2 ) in hiv - infected women switching to ral - containing antiretroviral therapy ( art ) . 
 thirty - seven women ( ral = 17 ; pi / nnrti = 20 ) with a median age of 43 years and bmi 32  kg / m2 completed week 24 . 
 txb2 increased in the ral versus pi / nnrti arm ( + 0.09 versus 0.02 ; p = 0.06 ) . 
 baseline pgi - m was lower in the ral arm ( p = 0.005 ) ; no other between - arm cross - sectional differences were observed . in the pi / nnrti arm , 24-week visceral adipose tissue change correlated with pgi - m ( rho = 0.45 ; p = 0.04 ) and txb2 ( rho = 0.44 ; p = 0.005 ) changes , with a trend seen for pge - m ( rho = 0.41 ; p = 0.07 ) . in an adjusted model , age  50 years ( 
 n = 8) was associated with increased pge - m ( p = 0.04 ) . in this randomized trial , 
 a switch to ral did not significantly affect urinary eicosanoids over 24 weeks . 
 in women continuing pi / nnrti , increased visceral adipose tissue correlated with increased pgi - m and pge - m . 
 older age ( 50 ) was associated with increased pge - m . 
 relationships between aging , adiposity , art , and eicosanoids during hiv - infection require further study . 
1. Introduction
2. Materials and Methods
3. Results
4. Discussion