RESEARCH ARTICLE
Obs uc i e Sleep Apnoea Synd ome,
Endo helial Func ion and Ma ke s o
Endo helializa ion. Changes a e CPAP
Rocio Muñoz-He nandez
1☯‡
, An onio J. Vallejo-Vaz
1☯‡
, Angeles Sanchez A mengol
2‡
,
Ra ael Mo eno-Luna
3‡
, Candela Caballe o-E aso
1,2
, Hada C. Mache
4
, Jose Villa
1,5
,
Ana M Me ino
6
, Ja ie Cas ell
7
, F ancisco Capo e
2
, Pablo S ie el
1,5
*
1Labo a o io de Hipe ensión A e ial e Hipe coles e olemia, Ins i u o de Biomedicina de Se illa (IBiS),
Hospi al Uni e si a io Vi gen del Rocío/CSIC/Uni e sidad de Se illa, Se illa, Spain, 2Unidad del Sueño,
Unidad Medico Qui ú gica de En e medades Respi a o ias, Hospi al Vi gen del Rocío, Se illa, Spain,
3Depa amen o de Fisiopa ología Vascula , Hospi al Nacional de Pa apléjicos, SESCAM, Toledo, Spain,
4Se icio de Bioquímica Clínica, Hospi al Vi gen del Rocío, Se illa, Spain, 5Unidad Clínico Expe imen al
de Riesgo Vascula (UCAMI), Hospi al Vi gen del Rocío, Se illa, Spain, 6Ins i u o de In es igación
Biomédica Bell i ge (IDIBELL), L'Hospi ale de Llob ega , Ba celona, Spain, 7UGC de Radiología, Hospi al
Vi gen del Rocío, Se illa, Spain
☯These au ho s con ibu ed equally o his wo k.
‡RMH and AJV a e sha ed i s au ho s on his wo k. ASA and RML a e second au ho s on his wo k. ASA
and RML also con ibu ed equally o his wo k.
*[email p o ec ed]
Abs ac
S udy objec i es
This s udy ies o assess he endo helial unc ion in i o using low-media ed dila a ion
(FMD) and se e al bioma ke s o endo helium o ma ion/ es o a ion and damage in pa ien s
wi h obs uc i e sleep apnoea (OSA) synd ome a baseline and a e h ee mon hs wi h
CPAP he apy.
Design
Obse a ional s udy, be o e and a e CPAP he apy.
Se ing and Pa ien s
We s udied 30 pa ien s wi h apnoea/hypopnoea index (AHI) >15/h ha we e compa ed wi h
hemsel es a e h ee mon hs o CPAP he apy. FMD was assessed non-in asi ely in i o
using he Lase -Dopple lowme y. Ci cula ing cell- ee DNA (c -DNA) and mic opa icles
(MPs) we e measu ed as ma ke s o endo helial damage and he ascula endo helial
g ow h ac o (VEGF) was de e mined as a ma ke o endo helial es o a ion p ocess.
Measu emen s and esul s
A e h ee mon h wi h CPAP, FMD signi ican ly inc eased (1072.26 ±483.21 s.
1604.38 ±915.69 PU, p<0.005) c -DNA and MPs signi ican ly dec eased (187.93 ±115.81
PLOS ONE | DOI:10.1371/jou nal.pone.0122091 Ma ch 27, 2015 1/13
OPEN ACCESS
Ci a ion: Muñoz-He nandez R, Vallejo-Vaz AJ,
Sanchez A mengol A, Mo eno-Luna R,
Caballe o-E aso C, Mache HC, e al. (2015)
Obs uc i e Sleep Apnoea Synd ome, Endo helial
Func ion and Ma ke s o Endo helializa ion. Changes
a e CPAP. PLoS ONE 10(3): e0122091.
doi:10.1371/jou nal.pone.0122091
Academic Edi o : Wing-ho Yung, The Chinese
Uni e si y o Hong Kong, HONG KONG
Recei ed: Decembe 23, 2014
Accep ed: Feb ua y 22, 2015
Published: Ma ch 27, 2015
Copy igh : © 2015 Muñoz-He nandez e al. This is
an open access a icle dis ibu ed unde he e ms o
he C ea i e Commons A ibu ion License, which
pe mi s un es ic ed use, dis ibu ion, and
ep oduc ion in any medium, p o ided he o iginal
au ho and sou ce a e c edi ed.
Da a A ailabili y S a emen : All ele an da a a e in
he pape .
Funding: This wo k was suppo ed by wo g an s: 1)
Ins i u o de Salud Ca los III, PI/0044/07, o
D . P. S ie el. 2) Conseje ia de Salud, Jun a de
Andalucía, P09-CTS-4971, o D a. M.A Sánchez-
A mengol. The unde s had no ole in s udy design,
da a collec ion and analysis, decision o publish, o
p epa a ion o he manusc ip .
s. 121.28 ±78.98 pg/ml, p<0.01, and 69.60 ±62.60 s. 39.82 ±22.14 U/μL, p<0.05, espec-
i ely) and VEGF le els inc eased (585.02 ±246.06 s. 641.11 ±212.69 pg/ml, p<0.05).
These changes we e highe in pa ien s wi h mo e se e e disease. The e was a ela ionship
be ween ma ke s o damage ( = -0.53, p<0.005) bu no be ween ma ke s o damage and
es o a ion, hus sugges ing ha bo h ypes o ma ke s should be measu ed oge he .
Conclusions
CPAP he apy imp o es FMD. This imp o emen may be ela ed o an inc ease o endo he-
lial es o a ion p ocess and a dec ease o endo helial damage.
In oduc ion
Obs uc i e sleep apnoea (OSA) synd ome is a espi a o y diso de cha ac e ized by he p es-
ence o apnoeas-hypopneas du ing sleep ha lead o episodes o in e mi en hypoxia. A ela-
ionship among OSA, ascula isk ac o s and ascula disease has been desc ibed in la ge
p ospec i e s udies [1,2]. Howe e , he unde lying mechanisms linking OSA synd ome and
ascula pa hology emain unclea . Se e al s udies ha e sugges ed ha pa ien s wi h OSA syn-
d ome ha e an endo helial dys unc ion [3,4] ela ed o he in e mi en hypoxemia and conse-
quen gene a ion o eac i e oxygen species (ROS), p o-in lamma o y molecules, ma ke s o
inc eased oxida i e s ess, as well as se e al coagula ion and lipid me abolism diso de s [5]. In-
e es ingly, ea men o he in e mi en hypoxemia wi h con inuous posi i e ai way p essu e
(CPAP) leads o an imp o emen in endo helial unc ion [6].
Endo helial dys unc ion can be assessed by measu ing se e al bioma ke s in plasma, bu
also by assessing endo helial unc ion in i o measu ing low-media ed dila a ion (FMD) using
high-de ini ion ul asonog aphy [7] o Lase -Dopple lowme y [8]. Mo e ecen ly, new
ma ke s o endo helial de elopmen and damage ha e been sugges ed. The p oduc ion and e-
lease o p o-angiogenic ac o s such as he ascula endo helial g ow h ac o (VEGF) can mo-
bilize endo helial p ogeni o cells and may enhance he ec ui men o hese cells o he inju ed
ascula issue [9]. Con e sely, ci cula ing mic opa icles (MPs) a e small esicles (<1μm) e-
leased om he endo helium in esponse o se e al inju ies and ha ha e been epo ed o be
inc eased in ce eb o ascula disease, hype ension, diabe es, smoking and co ona y a e y dis-
ease [5]. Recen e idence sugges ha CD31
+
/annexin V
+
MPs a e inc eased in condi ions o
sys emic endo helial cell damage and ca dio ascula disease [10]. Finally, ci cula ing cell- ee
DNA (c -DNA) is conside ed as ano he ma ke o cell damage measu able in he bloods eam
ha may inc ease in di e en si ua ions in ol ing hypoxia [11].
Ou pu pose in his s udy was o assess he endo helial unc ion in i o, using FMD by Lase -
Dopple lowme y, and o measu e di e en ma ke s o endo helium o ma ion and damage
(VEGF,CD31
+
/annexin V
+
MPs and ci cula ing c -DNA) in a g oup o pa ien s diagnosed wi h
OSA, a baseline and a e h ee mon h wi h CPAP he apy. As a seconda y objec i e we aimed o
co ela e he changes obse ed in hose pa ame e s wi h he se e i y o OSA synd ome a baseline.
Ma e ials and Me hods
Subjec s
We included 30 consecu i e pa ien s diagnosed wi h OSA and ha ing an apnoea-hypopnea
index (AHI) g ea e han 15 pe hou . Each pa ien was his own con ol a e h ee mon hs o
OSA Synd ome and Ma ke s o Endo helializa ion
PLOS ONE | DOI:10.1371/jou nal.pone.0122091 Ma ch 27, 2015 2/13
Compe ing In e es s: The au ho s ha e decla ed
ha no compe ing in e es s exis .
CPAP he apy. Exclusion c i e ia we e as ollows: 1) absence o indica ion o CPAP, 2) p io
ea men wi h CPAP, 3) e usal, in ole ance, o less han ou hou s use o CPAP in he nigh -
ime, 4) any change o d ugs ha may ha e in luence on blood p essu e o endo helial unc ion
wi hin he h ee mon hs o ollow-up, 5) glome ula il a ion a e (MDRD) <30 ml/min/
1,73m
2
. E idence o any o he acu e o ch onic condi ion ha in he in es iga o ’s opinion may
ha e in luence on he pa ame e s assessed in ou s udy was also conside ed o exclusion c i e ia.
This s udy was app o ed by he Human Resea ch Re iew Commi ee a he Vi gen del
Rocío Uni e si y Hospi al and all he pa icipan s p o ided hei w i en in o med consen
be o e inclusion.
Polyg aphy
All he included pa ien s unde wen a ended espi a o y polyg aphy [“Sibelhome plus”
1
(Sibelmed, SIBEL S.A, Ba celona)] in he sleep labo a o y o ou cen e . Respi a o y polyg aphy
included con inuous eco ding o o onasal low and p essu e, hea a e, ho acic and abdomi-
nal espi a o y mo emen s, and oxygen sa u a ion (SaO2). Polyg aphy da a we e sco ed manu-
ally by ained pe sonnel ( he Spanish Socie y o Pulmonology and Tho acic Su ge y guidelines
we e ollowed o he polyg aphy eadings [12]). Apnea was de ined as an in e up ion o o o-
nasal ai low o mo e han 10 seconds. Hypopnea was de ined as a 50% educ ion in o onasal
ai low o mo e han 10 seconds associa ed wi h an oxygen desa u a ion o 4% o highe .
Apnea-hypopnea index (AHI) was de ined as he numbe o apneas plus hypopneas pe
hou o eco ding, desa u a ion index was de ined as he he numbe o oxygen desa u a ion
pe hou o eco ding. The a e age SaO2 was also measu ed.
CPAP P essu e Ti a ion
In he pa ien s diagnosed o OSA, CPAP indica ion was pe o med acco ding o he Spanish
Socie y o Pulmonology and Tho acic Su ge y guidelines [12]. In hese pa ien s, op imal CPAP
p essu e was i a ed a home by an au o CPAP de ice (REMs a au o, Philips Respi onics,
Pennsyl ania, USA) wi hin a pe iod o less han 15 days a e he diagnos ic s udy, acco ding
o a p ocedu e p e iously alida ed [13]. The op imal ixed p essu e was de e mined by ained
pe sonnel, based on he isual e alua ion o he aw da a eco ding om he nigh s udy.
24-hou - Ambula o y blood p essu e moni o ing
Blood p essu e (BP) was measu ed using a alida ed 24-h de ice (SpaceLabs 90207, Redmond,
WA, USA). I was p og ammed o measu e BP e e y 20 min du ing 24 h. The limi s o conside
he diagnosis o hype ension and p esence o absence o dippe pa e n we e hose p oposed
in he ESC/ESH guidelines [14].
Assessmen o low-media ed dila ion (FMD) by Lase -Dopple
lowme y
A Lase -Dopple linea Pe i lux Sys em 5000 (Pe imed SA) was used o measu e FMD in he
o ea m a 15 cm om he w is a e 4 minu es o ischemia p o oked in he a m wi h a BP
cu ( o de ails see e e ence [8]).
Measu emen o VEGF
Le els o ascula endo helial g ow h ac o (VEGF) we e analyzed in se um by ELISA using
he Quan ikine VEGF ELISA Ki (R&D Sys ems, Minneapolis, MN).
OSA Synd ome and Ma ke s o Endo helializa ion
PLOS ONE | DOI:10.1371/jou nal.pone.0122091 Ma ch 27, 2015 3/13
Ci cula ing CD31+/annexin V+ Mic opa icles
Hepa in-bu e ed blood samples we e ob ained and p ocessed and immedia ely spun a 13,000
×g o 20 minu es o sepa a e he pla ele s.
Pla ele -poo plasma was incuba ed o 15 min wi h a monoclonal an ibody agains FITC-
labeled an i-CD31 an ibody (BD Pha mingen. BD Bioscience, CA), ollowed by incuba ion
wi h PE-conjuga ed Annexin V ki s acco ding o he manu ac u e ’s ins uc ions (BD Pha -
mingen, BD bioscience, CA). The nega i e con ol (ze o alue) was ob ained using he iso ype
an ibodies. Flow Coun Calib a o beads (Beckman Coul e , Ma seille, F ance) we e added.
Analyses we e pe o med in a Coul e Cy omic FC 500 low cy ome e (Beckman Coul e )
wi h CXP so wa e (Beckman Coul e ). Each sample was measu ed in iplica e and he mean
was aken as he inal esul .
Ci cula ing cell- ee DNA measu emen (c -DNA)
Once collec ed, blood samples we e spun o 8 minu es a 3,500 pm, and he se um was ozen
a −80°C o la e DNA ex ac ion. DNA was ex ac ed au oma ically om he s o ed se um
samples (400 μl) using a Compac MagnaPu e Ins umen and he nucleic acid isola ion
MagNA Pu e Compac Nucleic Acid Isola ion Ki I (Roche Diagnos ics, Basel, Swi ze land),
acco ding o he To al Plasma NA 100 400 V3 1 p o ocol. The DNA was esuspended in a inal
olume o 50 μl o wa e , and he se um DNA empla e was ampli ied in a inal olume o 20 μl
using a eal- ime quan i a i e polyme ase chain eac ion (PCR) assay o he be a-globin gene
on a Ligh -Cycle 480 Real-Time PCR ins umen (Roche Diagnos ics), acco ding o he manu-
ac u e ’s ins uc ions. The β-globin Taqman sys em uses he ollowing p ime s: be a-globin-
354F (50- TG CAC CTG ACT CCT GAG GAG A-30); be a-globin-455R (50-CCT TGA TAC
CAA CCT GCC CAG-30); and a dual-labeled luo escen p obe be a-globin-402T (50-(FAM)
TCT GGC CAA GTT TCA ACT CTG CTC GCT (TAMRA)-30). Ampli ica ion was ca ied ou
o e 48 cycles a 95°C o 5 minu es and a 62°C o 20 minu es, and he inal size o he ampli-
con was 102 base pai s.
S a is ical analysis
Quan i a i e da a we e exp essed as mean ± SD. The Shapi o–Wilk es was used o assess no -
mali y o dis ibu ions. We compa ed he changes a e 3 mon hs o CPAP he apy wi h espec
o he baseline alues using S uden ’s - es o pai ed samples in case o no mali y. When sam-
ples we e no no mally dis ibu ed, a Wilcoxon pai ed ank es was used. Pea son co ela ions
we e ca ied ou o explo e he possible linea ela ionship be ween a iables a baseline and
changes a e he CPAP- ea men pe iod. Di e ences we e conside ed o be signi ican when
p alues we e less han 0.05. All analyses we e pe o med wi h SSPS 15.0 so wa e.
Resul s
The baseline cha ac e is ics o he s udied pa ien s a e shown in Table 1. The a e age compli-
ance wi h CPAP he apy was 5.26 ± 1.61 hou s/nigh . Table 2 illus a es he changes obse ed
in se e al a iables a e he 3-mon h pe iod wi h CPAP ea men . B ie ly, BP dec eased, pa -
icula ly du ing he nigh - ime, hus causing an inc ease in he numbe o subjec s ha ing a
“dippe ”pa e n.
A he same ime, he endo helial unc ion assessed by FMD by Lase -Dopple lowme y
imp o ed, since he a ea o hype aemia a e he ischemia signi ican ly inc eased. Mo eo e ,
ma ke s o endo helial damage (CD31
+
/annexin V
+
MPs and ci cula ing c -DNA) dec eased,
whe eas a ma ke o eendo elializa ion (VGEF) inc eased.
OSA Synd ome and Ma ke s o Endo helializa ion
PLOS ONE | DOI:10.1371/jou nal.pone.0122091 Ma ch 27, 2015 4/13
Table 1. Baseline cha ac e is ics o he pa ien s.
Age (yea s) 51.70 ±11.54 (95%CI: 47.39, 56.01)
Sex (Male/Female) M: 19 (63.3%) / F: 11 (36.7%)
Epwo h's Ques ionnai e (poin s) 10.73 ±3.69 (95%CI: 9.35, 12.11)
Apnoea-hypopnea index (No/h) 56.28 ±25.53 (95%CI: 46.74, 65.82)
Oxygen desa u a ion Index 54.49 ±24.84 (95%CI: 45.21, 63.76)
Mean oxygen sa u a ion (%) 90.57 ±4.47 (95%CI: 88.9, 92.24)
A e ial Hype ension 17 (56.7%)
Type 2 Diabe es Melli us 7 (23.3%)
Hype choles e olemia 17 (56.7%)
Cu en smoke s 9 (30.0%)
Ex-smoke s 8 (26.7%)
Co ona y a e y disease 1 (3.3%)
Ic us 1 (3.3%)
Body mass index (kg/m
2
) 35.83 ±6.56 (95%CI: 33.38, 38.28)
Wais ci cum e ence (cm) 115.21 ±12.53 (95%CI: 110.53, 119.89)
Me abolic synd ome (ATP-III c i e ia) 16 (53.3%)
Mean numbe o Me abolic Synd ome c i e ia (ATP-III) 2.66 ±1.12 (95%CI: 2.24, 3.08)
Da a a e shown as mean ±SD (95% confidence in e als) o n (%).
doi:10.1371/jou nal.pone.0122091. 001
Table 2. Changes o blood p essu e, low-media ed dila ion, CD31
+
/annexin V
+
mic opa icles, ci cula ing cell- ee DNA and VEGF a e CPAP.
Baseline A e 3 mon hs wi h CPAP he apy p
24 h SBP (mmHg) 125.36 ±12.28 121.27 ±12.64 <0.01
24 h DBP (mmHg) 76.06 ±10.54 72.58 ±10.93 <0.005
Day- ime SBP (mmHg) 128.20 ±12.74 125.58 ±14.20 <0.05
Day- ime DBP (mmHg) 79.13 ±11.18 76.10 ±12.15 <0.005
Nigh - ime SBP (mmHg) 118.24 ±14.59 111.44 ±11.95 <0.005
Nigh - ime DBP (mmHg) 68.62 ±11.95 64.65 ±10.53 <0.05
24 h. PP (mm Hg) *49.30 ±7.61 48.68 ±7.56 NS
Day- ime PP (mm Hg) *49.06 ±8.21 49.48 ±8.42 NS
Nigh - ime PP (mm Hg) *49.62 ±7.58 46.79 ±7.04 <0.005
AR-SBP (%) 35.81 ±25.95 27.11 ±25.14 NS
AR-DBP (%) 32,52 ±28,02 25,63 ±27,08 <0.05
Dippe (%) 20.7% 55.2% <0.05
Non-Dippe (%) 79.,3% 44.8% <0.05
FMD, hype emic a ea (PU) 1072.2 ±483.2 1604.3 ±915.6 <0.005
CD31
+
/Annexin V
+
MPs (U/μL) 69.60 ±62.60 39.82 ±22.14 <0.05
c -DNA (ng/ml) 187.93 ±115.81 121.28 ±78.98 <0.01
VEGF (pg/ml) 585.02 ±246.06 641.11 ±212.69 <0.05
Da a a e shown as mean ±SD (95% confidence in e als) o n (%).
(*) Pulse p essu e (PP) is defined as sys olic minus dias olic blood p essu e. AR: abno mal eadings; c -DNA: ci cula ing cell- ee DNA; CPAP: con inuous
posi i e p essu e ai way; DBP: Dias olic Blood P essu e; FMD: Flow media ed dila a ion; MPs: mic opa icles; PP: pulse p essu e; SBP: Sys olic Blood
P essu e; VEGF: Vascula endo helial g ow h ac o .
doi:10.1371/jou nal.pone.0122091. 002
OSA Synd ome and Ma ke s o Endo helializa ion
PLOS ONE | DOI:10.1371/jou nal.pone.0122091 Ma ch 27, 2015 5/13

Fig 1 shows how he hype aemic a ea a e ischemia was nega i ely ela ed o VEGF. How-
e e FMD did no co ela e wi h CD31
+
/annexin V
+
MPs o ci cula ing c -DNA. BP le els did
no co ela e wi h VEGF. CD31
+
/annexin V
+
MPs and ci cula ing c -DNA co ela ed wi h
24-h dias olic BP ( = 0.39, p<0.05, and = 0.41, p<0.05, espec i ely), day ime dias olic BP
( = 0.38, p<0.05; = 0.41, p<0.05), 24-h mean BP ( = 0.43, p<0.01; = 0.39, p<0.05) and
day ime mean BP ( = 0.44, p<0.01; = 0.39, p<0.05). 24-h sys olic BP also co ela ed wi h ci -
cula ing c -DNA ( = 0.42, p<0.01).
As i is obse ed in he Fig 2, bo h ma ke s o endo helial damage a e posi i ely ela ed each
o he . Howe e , he e was no any signi ican ela ionship be ween CD31
+
/annexin V
+
MPs
and VEGF ( = -0.27, p = 0.25) o be ween ci cula ing c -DNA and VEGF ( = 0.02, p = 0.93).
Figs 3and 4show how he apnoea-hypopnea index and he oxygen desa u a ion index,
espec i ely, we e co ela ed in e sely wi h he changes obse ed in he alues o ci cula ing
Fig 1. Rela ionship be ween low media ed dila a ion measu ed by Lase -Dopple lowme y
(hype aemic a ea) and ascula endo helial g ow h ac o (VEGF).
doi:10.1371/jou nal.pone.0122091.g001
Fig 2. Rela ionship be ween CD31
+
/annexin V
+
MPs and ci cula ing c -DNA.
doi:10.1371/jou nal.pone.0122091.g002
OSA Synd ome and Ma ke s o Endo helializa ion
PLOS ONE | DOI:10.1371/jou nal.pone.0122091 Ma ch 27, 2015 6/13
Fig 3. Rela ionship among he se e i y o he disease measu ed acco ding o he apnoea-hypopnea
index and changes om baseline in ci cula ing c -DNA, CD31
+
/annexin V
+
MPs and ascula
endo helial g ow h ac o (VEGF).
doi:10.1371/jou nal.pone.0122091.g003
OSA Synd ome and Ma ke s o Endo helializa ion
PLOS ONE | DOI:10.1371/jou nal.pone.0122091 Ma ch 27, 2015 7/13
Fig 4. Rela ionship among he se e i y o he disease measu ed acco ding o he oxygen
desa u a ion index, and changes om baseline in ci cula ing c -DNA, CD31
+
/annexin V
+
MPs and
ascula endo helial g ow h ac o (VEGF).
doi:10.1371/jou nal.pone.0122091.g004
OSA Synd ome and Ma ke s o Endo helializa ion
PLOS ONE | DOI:10.1371/jou nal.pone.0122091 Ma ch 27, 2015 8/13
c -DNA o CD31
+
/annexin V
+
MPs and posi i ely wi h he changes in VEGF le els. Con e se-
ly, Fig 5 displays mean SaO2 co ela ing posi i ely wi h he changes obse ed in c -DNA o
CD31
+
/annexin V
+
MPs and co ela ing in e sely wi h changes in VEGF alues (al hough he
las co ela ion did no each s a is ical signi icance (p<0.10).
Discussion
Vascula unc ion has been assessed in i o using he FMD on b achial a e y wi h ul asounds.
Howe e , mo e ecen ly a new echnique has appea ed o measu ing FMD using Lase -
Dopple lowme y. This echnique is mo e obse e -independen and i measu es mic oci cu-
la ion a he han ascula unc ion in highe a e ies. As a as we know, he e a e ew s udies
wi h his echnique in pa ien s wi h OSA, and only one o hem e alua ing he e ec o CPAP
on FMD [15].
In he p esen s udy we ha e obse ed a ma ked imp o emen in FMD (exp essed as he inc e-
men in he hype emia a ea measu ed in PU) a e CPAP he apy (Table 2). This imp o emen
was no ela ed o he wo ma ke s o endo helium damage ha we measu ed (CD31
+
/annexin
V
+
MPs and ci cula ing c -DNA); howe e , i was nega i ely ela ed o a ma ke o es o a ion o
he endo helium (VEGF) (Fig 1), pe haps indica ing ha pa ien s wi h a good endo helial unc ion
do no need o ha e high alues o VEGF and ice e sa. Howe e , he in luence o addi ional ac-
o s ha may con use his ela ionship canno be uled ou ; o ins ance, he ime o exposu e o
he sleep apnea: a sho e ime may lead o lesse impai men o endo helial unc ion and lowe
VGEF le els. Ne e heless, pa ien s we e all included soon jus a e he diagnosis, o minimize
di e ences among pa ien s in his ega d.
MPs a e small (<1μm) esicles ha a e eleased om he endo helium in esponse o
se e al inju ies. The le el o ci cula ing MPs in pe iphe al blood has been epo ed o be
inc eased in se e al ischemic diseases [7]. I seems ha MPs om ac i a ed leukocy es
(CD62L_ MPs) a e highe in pa ien s wi h OSA and a posi i e co ela ion be ween ci cula ing
le els o CD62L_ MPs and noc u nal hypoxemia se e i y has been epo ed [16,17,18]. Ne e -
heless, o he au ho s s udying child en wi h sleep b ea hing diso de s obse ed ha leukocy e
CD11b+ MPs and pla ele CD41a+ MPs we e he mic opa icles ha co ela ed he bes wi h
he apnoea-hypopnea index [19].
In ou s udy we measu e CD31
+
/annexin V
+
MPs (mic opa icles ma ked wi h an i-CD31
an ibody, ollowed by incuba ion wi h annexin V) in plasma and we ound a signi ican de-
c ease a e CPAP he apy (Table 2), which was mo e e iden in pa ien s wi h a mo e se e e
disease (Figs 3–5). We chose hese ypes o CD31
+
/annexin V
+
MPs because hey ha e been
ound o be inc eased in o he ca dio ascula pa hologies [10,20,21] and, o ou bes knowl-
edge, hey ha e ne e been in es iga ed in OSA synd ome. In ou wo k we also ound ha his
ype o CD31
+
/annexin V
+
MPs was well ela ed o o he ma ke s o endo helial dys unc ion,
such as ci cula ing c -DNA (Fig 2).
On he o he hand, i has also been obse ed ha c -DNA le els ise in pa hologies in ol ing
ischemia, such as acu e co ona y synd ome, ischemic hea ailu e, s oke, mesen e ic ischemia
o in pa ien s who ha e su e ed a ca diac a es ou side he hospi al [22,23]; we ha e also e-
cen ly epo ed highe le els o his ma ke o endo helial damage in p eeclampsia and HELLP
synd ome [11]. These bo h pa hologies ha e some simila i ies wi h he OSA synd ome, since
in bo h condi ions he hypoxia (sys emic o speci ic o gan- ela ed) can play a ole ( he o igin
o he a o emen ioned pa hologies has been sugges ed o be placen al hypoxia) and pa ien s
can be s udied du ing he pa hologic s a e as well as a e he imp o emen o he in e mi en
hypoxia (OSA synd ome) o he eco e y o he disease (a e deli e y, when he placen a is e-
mo ed). The in e es ing esul s ob ained in ha s udy encou aged us o measu e c -DNA in
OSA Synd ome and Ma ke s o Endo helializa ion
PLOS ONE | DOI:10.1371/jou nal.pone.0122091 Ma ch 27, 2015 9/13