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Mark A. Skidmore, Khairul Mohd Fadzli Mustaffa, Lynsay C. Cooper, Scott E. Guimond, Edwin A. Yates, Alister G. Craig
A semi-synthetic glycosaminoglycan analogue inhibits and reverses Plasmodium falciparum cytoadherence
www.plosone.org
www.plosone.org
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ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 528.0377 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(Primary)Tj
3.4979 0 Td
(human)Tj
3.0784 0 Td
(umbilical)Tj
3.9571 0 Td
(vein)Tj
1.9106 0 Td
(vascular)Tj
3.4752 0 Td
(endothelial)Tj
4.6715 0 Td
(cells)Tj
1.9162 0 Td
(\(HUVEC\))Tj
4.3313 0 Td
(and)Tj
1.7291 0 Td
(human)Tj
3.0784 0 Td
(dermal)Tj
3.0444 0 Td
(micro-)Tj
-34.6901 -1.2982 Td
(vascular)Tj
3.4695 0 Td
(endothelial)Tj
4.6715 0 Td
(cells)Tj
1.9219 0 Td
(\(HDMEC\))Tj
4.5297 0 Td
(obtained)Tj
3.7304 0 Td
(from)Tj
2.2053 0 Td
(Promocell)Tj
4.3257 0 Td
(were)Tj
2.1203 0 Td
(cultured)Tj
3.5659 0 Td
(as)Tj
1.0148 0 Td
(per)Tj
1.5364 0 Td
(the)Tj
1.4683 0 Td
(manu-)Tj
-34.5597 -1.2983 Td
(facturer's)Tj
3.9344 0 Td
(instructions.)Tj
5.2214 0 Td
(The)Tj
1.7858 0 Td
(Human)Tj
3.3165 0 Td
(Brain)Tj
2.4208 0 Td
(Endothelial)Tj
4.8132 0 Td
(Cell)Tj
1.8028 0 Td
(line)Tj
1.7008 0 Td
(\(HBEC5i\))Tj
4.2236 0 Td
(was)Tj
1.7008 0 Td
(cultured)Tj
3.5659 0 Td
(as)Tj
-34.486 -1.3039 Td
(described)Tj
4.0478 0 Td
(by)Tj
1.1792 0 Td
(Tripathi)Tj
/F15 1 Tf
3.5093 0 Td
[()8()]TJ
0.9127 0 Td
[()11()]TJ
/F10 1 Tf
(.)Tj
1.1622 0 Td
([)Tj
0.83 0.64 0.02 0 k
(5)Tj
0 g
(].)Tj
1.5988 0 Td
(Primary)Tj
3.4922 0 Td
(endothelial)Tj
4.6715 0 Td
(cells)Tj
1.9162 0 Td
(at)Tj
0.9524 0 Td
(passage)Tj
3.2088 0 Td
(46)Tj
1.6895 0 Td
(were)Tj
2.1203 0 Td
(used)Tj
2.0522 0 Td
(for)Tj
1.389 0 Td
(all)Tj
1.1395 0 Td
(exper-)Tj
-35.0416 -1.2983 Td
(iments.)Tj
3.1634 0 Td
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2.296 0 Td
(to)Tj
1.0262 0 Td
(use,)Tj
1.7574 0 Td
(cells)Tj
1.9163 0 Td
(were)Tj
2.1259 0 Td
(stimulated)Tj
4.422 0 Td
(by)Tj
1.1793 0 Td
(addition)Tj
3.5999 0 Td
(of)Tj
1.0148 0 Td
(1)Tj
0.6973 0 Td
(ng.ml)Tj
ET
Q
q
1 j
1 J
0 w
7.4999 0 0 6.6659 455.0173 480.4724 cm
BT
/F10 1 Tf
1 TL
-0.0053 Tc
0 0 Td
(-1)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 463.4078 476.0503 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(TNF)Tj
2.0976 0 Td
(for)Tj
1.3833 0 Td
(18)Tj
1.1735 0 Td
(h)Tj
0.7484 0 Td
(to)Tj
1.0204 0 Td
(allow)Tj
-32.7625 -1.2982 Td
(enhanced)Tj
4.0762 0 Td
(ICAM-1)Tj
3.6283 0 Td
(expression)Tj
4.4617 0 Td
(on)Tj
1.2756 0 Td
(the)Tj
1.4683 0 Td
(surface)Tj
3.0501 0 Td
(of)Tj
1.0148 0 Td
(the)Tj
1.4683 0 Td
(endothelial)Tj
4.6715 0 Td
(cells.)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 435.685 cm
BT
/F0 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(Parasite)Tj
3.8456 0 Td
(culture)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 418.6771 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(ItG)Tj
1.5703 0 Td
(\(ITvar16\))Tj
4.0649 0 Td
([)Tj
0.83 0.64 0.02 0 k
(20)Tj
0 g
(])Tj
1.8538 0 Td
(and)Tj
1.7292 0 Td
(A4)Tj
1.3776 0 Td
(\(ITvar14\))Tj
4.0648 0 Td
([)Tj
0.83 0.64 0.02 0 k
(21)Tj
0 g
(])Tj
1.8482 0 Td
(laboratory)Tj
4.354 0 Td
(parasite)Tj
3.3222 0 Td
(lines,)Tj
2.296 0 Td
(which)Tj
2.6476 0 Td
(are)Tj
1.4513 0 Td
(well)Tj
1.8198 0 Td
(character-)Tj
-32.3997 -1.2982 Td
(ized)Tj
1.8481 0 Td
(for)Tj
1.389 0 Td
(their)Tj
2.109 0 Td
(binding)Tj
3.3505 0 Td
(to)Tj
1.0261 0 Td
(ICAM-1)Tj
3.6227 0 Td
(and)Tj
1.7348 0 Td
(CD36)Tj
2.5681 0 Td
([)Tj
0.83 0.64 0.02 0 k
(22)Tj
0 g
(],)Tj
2.075 0 Td
(were)Tj
2.126 0 Td
(cultured)Tj
3.5659 0 Td
(under)Tj
2.6192 0 Td
(standard)Tj
3.7191 0 Td
(conditions)Tj
4.4787 0 Td
(in)Tj
-36.2322 -1.2983 Td
(RPMI)Tj
2.6305 0 Td
(1640)Tj
2.126 0 Td
(medium)Tj
3.6056 0 Td
(supplemented)Tj
5.8507 0 Td
(with)Tj
2.0012 0 Td
(37.5)Tj
1.8709 0 Td
(mM)Tj
1.9332 0 Td
(HEPES,)Tj
3.3845 0 Td
(7)Tj
0.6974 0 Td
(mM)Tj
1.9332 0 Td
(D-Glucose,)Tj
4.7338 0 Td
(25)Tj
1.1735 0 Td
(ug.ml)Tj
ET
Q
q
1 j
1 J
0 w
7.4999 0 0 6.6659 542.2677 397.1338 cm
BT
/F10 1 Tf
1 TL
-0.0053 Tc
0 0 Td
(-1)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 550.6015 392.7118 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(genta-)Tj
-35.0586 -1.3039 Td
(mycin)Tj
2.7326 0 Td
(sulfate,)Tj
3.0217 0 Td
(2)Tj
0.6973 0 Td
(mM)Tj
1.9332 0 Td
(L-glutamine)Tj
5.1363 0 Td
(and)Tj
1.7348 0 Td
(10%)Tj
1.9276 0 Td
(\(v/v\))Tj
2.1373 0 Td
(pooled)Tj
2.948 0 Td
(human)Tj
3.0784 0 Td
(serum)Tj
2.7213 0 Td
(at)Tj
0.9524 0 Td
(pH)Tj
1.508 0 Td
(7.2)Tj
1.3947 0 Td
(in)Tj
1.0374 0 Td
(a)Tj
0.652 0 Td
(gas)Tj
1.4796 0 Td
(mix-)Tj
-35.0926 -1.2982 Td
(ture)Tj
1.8425 0 Td
(comprising)Tj
4.7622 0 Td
(96%)Tj
1.9332 0 Td
(nitrogen,)Tj
3.8721 0 Td
(3%)Tj
1.4513 0 Td
(carbon)Tj
3.016 0 Td
(dioxide,)Tj
3.4299 0 Td
(and)Tj
1.7292 0 Td
(1%)Tj
1.4513 0 Td
(oxygen.)Tj
3.3108 0 Td
(Additionally,)Tj
5.4539 0 Td
(recently)Tj
-32.2524 -1.2983 Td
(culture)Tj
3.0387 0 Td
(adapted)Tj
3.3789 0 Td
(patient)Tj
3.0103 0 Td
(isolates:)Tj
3.3449 0 Td
(PO69,)Tj
2.6986 0 Td
(8146)Tj
2.1259 0 Td
(and)Tj
1.7292 0 Td
(8026)Tj
2.1203 0 Td
(were)Tj
2.1259 0 Td
(also)Tj
1.7632 0 Td
(included)Tj
3.7133 0 Td
(in)Tj
1.0375 0 Td
(the)Tj
1.4684 0 Td
(study)Tj
2.3924 0 Td
([)Tj
0.83 0.64 0.02 0 k
(23)Tj
0 g
(].)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 326.3244 cm
BT
/F0 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(Plasmagel)Tj
4.8283 0 Td
(trophozoite)Tj
5.1072 0 Td
(enrichment)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 309.3732 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(Parasite)Tj
3.3675 0 Td
(culture)Tj
3.0387 0 Td
(at)Tj
0.9524 0 Td
(trophozoite)Tj
4.8586 0 Td
(stage)Tj
2.1997 0 Td
(was)Tj
1.7007 0 Td
(centrifuged)Tj
4.7849 0 Td
(\(500)Tj
1.9842 0 Td
(g,)Tj
0.9128 0 Td
(5)Tj
0.6916 0 Td
(min\))Tj
2.1997 0 Td
(and)Tj
1.7291 0 Td
(the)Tj
1.4684 0 Td
(pRBC)Tj
2.6022 0 Td
(pellet)Tj
2.3754 0 Td
(was)Tj
-34.8659 -1.3039 Td
(resuspended)Tj
5.2327 0 Td
(in)Tj
1.0318 0 Td
(a)Tj
0.6576 0 Td
(ratio)Tj
2.0976 0 Td
(of)Tj
1.0148 0 Td
(2)Tj
0.6974 0 Td
(volumes)Tj
3.5546 0 Td
(pellet)Tj
2.3754 0 Td
(to)Tj
1.0261 0 Td
(3)Tj
0.6973 0 Td
(volumes)Tj
3.5603 0 Td
(RPMI-based)Tj
5.2384 0 Td
(growth)Tj
3.0728 0 Td
(media)Tj
2.6929 0 Td
(without)Tj
-32.9497 -1.2982 Td
(human)Tj
3.0784 0 Td
(serum)Tj
2.7212 0 Td
(\(incomplete)Tj
5.0343 0 Td
(medium\))Tj
3.9458 0 Td
(and)Tj
1.7291 0 Td
(5)Tj
0.6973 0 Td
(volumes)Tj
3.5603 0 Td
(Plasmion)Tj
3.9628 0 Td
(\(Laboratoire)Tj
5.2158 0 Td
(Fresenius)Tj
4.0308 0 Td
(Kabi,)Tj
-33.9758 -1.2983 Td
(France\),)Tj
3.5149 0 Td
(and)Tj
1.7291 0 Td
(allowed)Tj
3.2882 0 Td
(to)Tj
1.0261 0 Td
(settle)Tj
2.2677 0 Td
(for)Tj
1.389 0 Td
(2030)Tj
2.6362 0 Td
(min)Tj
1.8595 0 Td
(at)Tj
0.9524 0 Td
(37)Tj
1.1679 0 Td
(C.)Tj
1.1112 0 Td
(Trophozoite)Tj
5.1703 0 Td
(stage)Tj
2.1997 0 Td
(pRBC)Tj
2.6022 0 Td
(in)Tj
1.0375 0 Td
(the)Tj
1.4683 0 Td
(top)Tj
1.542 0 Td
(layer)Tj
-34.9622 -1.3039 Td
(were)Tj
2.1203 0 Td
(washed)Tj
3.1804 0 Td
(three)Tj
2.262 0 Td
(times)Tj
2.3925 0 Td
(in)Tj
1.0318 0 Td
(incomplete)Tj
4.6941 0 Td
(medium)Tj
3.6057 0 Td
(and)Tj
1.7347 0 Td
(the)Tj
1.4684 0 Td
(parasitaemia)Tj
5.2781 0 Td
(assessed)Tj
3.4752 0 Td
(by)Tj
1.1792 0 Td
(Giemsa)Tj
-32.4224 -1.2983 Td
(stained)Tj
3.0784 0 Td
(smear.)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 217.0204 cm
BT
/F0 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(Selection)Tj
4.2803 0 Td
(of)Tj
1.0441 0 Td
(pRBC)Tj
2.863 0 Td
(on)Tj
1.3134 0 Td
(ICAM-1)Tj
3.5859 0 Td
(purified)Tj
3.4488 0 Td
(protein)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 200.0125 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(To)Tj
1.3379 0 Td
(increase)Tj
3.4696 0 Td
(the)Tj
1.474 0 Td
(homogeneity)Tj
5.4595 0 Td
(of)Tj
1.0148 0 Td
(the)Tj
1.4683 0 Td
(ItG)Tj
1.5761 0 Td
(parasite)Tj
3.3278 0 Td
(population)Tj
4.5864 0 Td
(that)Tj
1.7802 0 Td
(expresses)Tj
3.9288 0 Td
(a)Tj
0.652 0 Td
(PfEMP-1)Tj
3.9117 0 Td
(protein)Tj
-33.9871 -1.3039 Td
(with)Tj
1.9955 0 Td
(high)Tj
2.0069 0 Td
(affinity)Tj
3.0671 0 Td
(for)Tj
1.389 0 Td
(ICAM-1,)Tj
3.8494 0 Td
(the)Tj
1.4683 0 Td
(population)Tj
4.5921 0 Td
(was)Tj
1.7008 0 Td
(subjected)Tj
3.9685 0 Td
(to)Tj
1.0205 0 Td
(selection)Tj
3.719 0 Td
(on)Tj
1.2756 0 Td
(ICAM-1)Tj
3.6283 0 Td
(protein.)Tj
-33.681 -1.2982 Td
(2.5)Tj
/F16 1 Tf
1.3946 0 Td
[()]TJ
/F10 1 Tf
(g)Tj
1.1905 0 Td
(of)Tj
1.0148 0 Td
(ICAM-1)Tj
3.6284 0 Td
(protein)Tj
3.1521 0 Td
(was)Tj
1.7007 0 Td
(coated)Tj
2.8347 0 Td
(on)Tj
1.2755 0 Td
(to)Tj
1.0262 0 Td
(50)Tj
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1.1678 0 Td
[()]TJ
/F10 1 Tf
(l)Tj
0.9752 0 Td
(of)Tj
1.0148 0 Td
(protein)Tj
3.1521 0 Td
(A)Tj
0.907 0 Td
(Dynabeads)Tj
4.6432 0 Td
(\(Invitrogen\))Tj
5.125 0 Td
(in)Tj
-34.2026 -1.2983 Td
(200)Tj
/F16 1 Tf
1.644 0 Td
[()]TJ
/F10 1 Tf
(l)Tj
0.9751 0 Td
(of)Tj
1.0148 0 Td
(1%)Tj
1.4514 0 Td
(\(v/v\))Tj
2.143 0 Td
(Bovine)Tj
2.999 0 Td
(Serum)Tj
2.8346 0 Td
(Albumin)Tj
3.8211 0 Td
(\(BSA\))Tj
2.6419 0 Td
(in)Tj
1.0374 0 Td
(PBS)Tj
1.8312 0 Td
(and)Tj
1.7291 0 Td
(incubated)Tj
4.1783 0 Td
(for)Tj
1.3889 0 Td
(1)Tj
0.6917 0 Td
(hour)Tj
2.1543 0 Td
(at)Tj
0.9524 0 Td
(room)Tj
-33.4882 -1.2983 Td
(temperature)Tj
5.125 0 Td
(with)Tj
2.0012 0 Td
(gentle)Tj
2.6249 0 Td
(rotation)Tj
3.4525 0 Td
(\(15)Tj
1.5081 0 Td
(rpm\).)Tj
2.4944 0 Td
(Dynabeads)Tj
4.6488 0 Td
(were)Tj
2.1203 0 Td
(washed)Tj
3.1805 0 Td
(with)Tj
1.9956 0 Td
(1%)Tj
1.4513 0 Td
(BSA/PBS)Tj
3.9061 0 Td
(and)Tj
1.7291 0 Td
(a)Tj
-36.2378 -1.3039 Td
(magnetic)Tj
3.9004 0 Td
(stand.)Tj
2.6135 0 Td
(50)Tj
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1.1736 0 Td
[()]TJ
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(l)Tj
0.9694 0 Td
(of)Tj
1.0148 0 Td
(synchronized)Tj
5.5956 0 Td
(and)Tj
1.7291 0 Td
(enriched)Tj
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(ItG)Tj
1.5704 0 Td
(parasite)Tj
3.3279 0 Td
(culture)Tj
3.0387 0 Td
(using)Tj
2.3867 0 Td
(Plasmion)Tj
3.9628 0 Td
(were)Tj
-35.0189 -1.2982 Td
(incubated)Tj
4.1782 0 Td
(with)Tj
1.9956 0 Td
(the)Tj
1.4683 0 Td
(coated)Tj
2.8346 0 Td
(beads)Tj
2.4718 0 Td
(in)Tj
1.0375 0 Td
(400)Tj
/F16 1 Tf
1.6441 0 Td
[()]TJ
/F10 1 Tf
(l)Tj
0.9751 0 Td
(1%)Tj
1.4513 0 Td
(BSA/PBS)Tj
3.9061 0 Td
(for)Tj
1.3833 0 Td
(45)Tj
1.1736 0 Td
(min)Tj
1.8538 0 Td
(at)Tj
0.9525 0 Td
(room)Tj
2.4264 0 Td
(temperature)Tj
5.1307 0 Td
(by)Tj
-34.8829 -1.2983 Td
(gentle)Tj
2.6248 0 Td
(rotation.)Tj
3.6737 0 Td
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2.914 0 Td
(pRBC)Tj
2.6022 0 Td
(were)Tj
2.1203 0 Td
(washed)Tj
3.1804 0 Td
(3)Tj
0.6974 0 Td
(times)Tj
2.3867 0 Td
(with)Tj
1.9956 0 Td
(1%)Tj
1.457 0 Td
(BSA/PBS)Tj
3.9004 0 Td
(using)Tj
2.3925 0 Td
(the)Tj
1.4683 0 Td
(magnetic)Tj
-31.4133 -1.3039 Td
(stand.)Tj
2.6135 0 Td
(Beads)Tj
2.5455 0 Td
(were)Tj
2.1259 0 Td
(resuspended)Tj
5.2328 0 Td
(in)Tj
1.0318 0 Td
(5)Tj
0.6973 0 Td
(ml)Tj
1.2869 0 Td
(of)Tj
1.0148 0 Td
(complete)Tj
3.8778 0 Td
(RPMI)Tj
2.6305 0 Td
(media)Tj
2.6929 0 Td
(and)Tj
1.7291 0 Td
(transferred)Tj
4.6432 0 Td
(to)Tj
1.0261 0 Td
(culture)Tj
3.033 0 Td
(in)Tj
-36.1811 -1.2983 Td
(a)Tj
0.6519 0 Td
(T25)Tj
1.7858 0 Td
(culture)Tj
3.0331 0 Td
(flask)Tj
2.0466 0 Td
(with)Tj
2.0012 0 Td
(the)Tj
1.4684 0 Td
(addition)Tj
3.5943 0 Td
(of)Tj
1.0148 0 Td
(100)Tj
/F16 1 Tf
1.6497 0 Td
[()]TJ
/F10 1 Tf
(l)Tj
0.9751 0 Td
(of)Tj
1.0148 0 Td
(washed)Tj
3.1748 0 Td
(red)Tj
1.5421 0 Td
(blood)Tj
2.5171 0 Td
(cells.)Tj
ET
Q
q
1 j
1 J
0 w
576 737.1 m
36 737.1 l
36 737.6 l
576 737.6 l
f*
36 741.2598 107.1496 23.6976 re
W* n
q
107.0929 0 0 23.6409 36 741.3165 cm
q
/I0 Do
Q
Q
Q
q
0 0 612 792 re
W* n
1 j
1 J
0 w
7.9999 0 0 7.9999 464.0314 745.7952 cm
BT
/F0 1 Tf
1 TL
-0.005 Tc
0 0 Td
(GAG)Tj
2.4307 0 Td
(inhibition)Tj
4.0748 0 Td
(of)Tj
1.0418 0 Td
(cytoadher)Tj
4.3157 0 Td
(ence)Tj
ET
Q
q
1 j
1 J
0 w
36 48.0002 m
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(were)Tj
2.126 0 Td
(assayed)Tj
3.2145 0 Td
(as)Tj
1.0148 0 Td
(controls)Tj
3.4809 0 Td
(The)Tj
1.7858 0 Td
(EC)Tj
ET
Q
q
1 j
1 J
0 w
7.4999 0 0 6.6659 558.8787 543.1181 cm
BT
/F10 1 Tf
1 TL
-0.0053 Tc
0 0 Td
(50)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 531.7795 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(values)Tj
2.6645 0 Td
(of)Tj
1.0148 0 Td
(all)Tj
1.1452 0 Td
(semi-synthetic,)Tj
6.2702 0 Td
(sulphated)Tj
4.0875 0 Td
(carbohydrates)Tj
5.862 0 Td
(were)Tj
2.1203 0 Td
(determined)Tj
4.8529 0 Td
(using)Tj
2.3868 0 Td
(a)Tj
0.6519 0 Td
(sigmoidal)Tj
4.1159 0 Td
(dose)Tj
-35.172 -1.2982 Td
(response)Tj
3.7303 0 Td
(curved)Tj
2.948 0 Td
(fitted)Tj
2.3244 0 Td
(post)Tj
1.9049 0 Td
(normalisation)Tj
5.828 0 Td
(\(with)Tj
2.3357 0 Td
(a)Tj
0.6577 0 Td
(100%)Tj
2.4037 0 Td
(upper)Tj
2.5739 0 Td
(maximal)Tj
3.7077 0 Td
(at)Tj
0.9524 0 Td
(2)Tj
0.6973 0 Td
(mins;)Tj
2.4378 0 Td
(0%)Tj
1.457 0 Td
(lower)Tj
-33.9588 -1.2983 Td
(maximal)Tj
3.702 0 Td
(represented)Tj
4.9095 0 Td
(by)Tj
1.1849 0 Td
(the)Tj
1.4684 0 Td
(time)Tj
2.0296 0 Td
(required)Tj
3.6396 0 Td
(for)Tj
1.3833 0 Td
(the)Tj
1.474 0 Td
(water)Tj
2.4265 0 Td
(control)Tj
3.1237 0 Td
(to)Tj
1.0262 0 Td
(clot)Tj
1.6951 0 Td
(normal)Tj
3.1464 0 Td
(human)Tj
3.0784 0 Td
(citrated)Tj
-34.2876 -1.3039 Td
(plasma\))Tj
3.3732 0 Td
(with)Tj
1.9955 0 Td
(GraphPad)Tj
4.3087 0 Td
(Prism)Tj
2.5965 0 Td
(6)Tj
0.6916 0 Td
(software)Tj
3.5887 0 Td
(and)Tj
1.7291 0 Td
(compared)Tj
4.2463 0 Td
(to)Tj
1.0261 0 Td
(those)Tj
2.3357 0 Td
(obtained)Tj
3.7361 0 Td
(for)Tj
1.3833 0 Td
(the)Tj
1.4683 0 Td
(heparin)Tj
-32.4791 -1.2983 Td
(control.)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 451.2188 cm
BT
/F0 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(Activated)Tj
4.2897 0 Td
(partial)Tj
2.9009 0 Td
(thromboplastin)Tj
6.6992 0 Td
(time)Tj
2.093 0 Td
(\(aPTT\))Tj
3.3118 0 Td
(coagulation)Tj
5.263 0 Td
(assay)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 434.211 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(Serially)Tj
3.135 0 Td
(diluted,)Tj
3.2655 0 Td
(sulphated)Tj
4.0819 0 Td
(carbohydrate)Tj
5.5049 0 Td
(samples)Tj
3.3788 0 Td
(\(25)Tj
/F16 1 Tf
1.5137 0 Td
[()]TJ
/F10 1 Tf
(l\))Tj
1.3096 0 Td
(were)Tj
2.126 0 Td
(incubated)Tj
4.1782 0 Td
(with)Tj
1.9956 0 Td
(normal)Tj
3.1464 0 Td
(human)Tj
-33.6356 -1.2982 Td
(citrated)Tj
3.2598 0 Td
(plasma)Tj
3.033 0 Td
(\(50)Tj
/F16 1 Tf
1.5137 0 Td
[()]TJ
/F10 1 Tf
(l;)Tj
1.1962 0 Td
(NHSBT\))Tj
3.7304 0 Td
(and)Tj
1.7348 0 Td
(Pathromtin)Tj
4.8472 0 Td
(SL)Tj
1.2359 0 Td
(reagent)Tj
3.1861 0 Td
(\(50)Tj
/F16 1 Tf
1.508 0 Td
[()]TJ
/F10 1 Tf
(l;)Tj
1.2019 0 Td
(Siemens\))Tj
3.8778 0 Td
(for)Tj
1.3833 0 Td
(2)Tj
0.6973 0 Td
(min)Tj
1.8539 0 Td
(at)Tj
0.9524 0 Td
(37)Tj
1.1735 0 Td
(C)Tj
-36.3852 -1.3039 Td
(prior)Tj
2.2506 0 Td
(to)Tj
1.0262 0 Td
(the)Tj
1.4683 0 Td
(addition)Tj
3.6 0 Td
(of)Tj
1.0148 0 Td
(calcium)Tj
3.3619 0 Td
(chloride)Tj
3.5149 0 Td
(\(25)Tj
/F16 1 Tf
1.508 0 Td
[()]TJ
/F10 1 Tf
(l;)Tj
1.2019 0 Td
(50)Tj
1.1679 0 Td
(mM\).)Tj
2.5001 0 Td
(The)Tj
1.7802 0 Td
(time)Tj
2.0296 0 Td
(taken)Tj
2.4207 0 Td
(for)Tj
1.389 0 Td
(clot)Tj
1.6951 0 Td
(formations)Tj
4.6204 0 Td
(to)Tj
-36.5496 -1.2983 Td
(occur)Tj
2.4717 0 Td
(\(an)Tj
1.5421 0 Td
(upper)Tj
2.5795 0 Td
(maximal)Tj
3.702 0 Td
(of)Tj
1.0148 0 Td
(2)Tj
0.6973 0 Td
(mins)Tj
2.2167 0 Td
(was)Tj
1.7008 0 Td
(observed\))Tj
4.1329 0 Td
(were)Tj
2.1259 0 Td
(recorded)Tj
3.7928 0 Td
(using)Tj
2.3924 0 Td
(a)Tj
0.652 0 Td
(Thrombotrak)Tj
5.6692 0 Td
(Solo)Tj
-34.6901 -1.2983 Td
(coagulometer)Tj
5.6635 0 Td
(\(Axis-Shield\))Tj
5.4992 0 Td
(as)Tj
1.0148 0 Td
(per)Tj
1.5307 0 Td
(the)Tj
1.4683 0 Td
(manufacturer's)Tj
6.2646 0 Td
(instructions.)Tj
5.227 0 Td
(Water)Tj
2.7099 0 Td
(and)Tj
1.7292 0 Td
(sodium)Tj
3.2314 0 Td
(porcine)Tj
-34.3386 -1.3039 Td
(mucosal)Tj
3.5376 0 Td
(heparin)Tj
3.3108 0 Td
(\(203)Tj
1.9843 0 Td
(IU/mg;)Tj
3.135 0 Td
(VWR\))Tj
2.846 0 Td
(were)Tj
2.1203 0 Td
(assayed)Tj
3.2145 0 Td
(as)Tj
1.0148 0 Td
(controls.)Tj
3.7077 0 Td
(The)Tj
1.7858 0 Td
(EC)Tj
ET
Q
q
1 j
1 J
0 w
7.4999 0 0 6.6659 478.885 367.5401 cm
BT
/F10 1 Tf
1 TL
-0.0053 Tc
0 0 Td
(50)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 488.2393 369.1842 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(values)Tj
2.6645 0 Td
(of)Tj
1.0148 0 Td
(all)Tj
1.1452 0 Td
(semi-syn-)Tj
-33.6469 -1.2982 Td
(thetic,)Tj
2.6815 0 Td
(sulphated)Tj
4.0819 0 Td
(carbohydrates)Tj
5.862 0 Td
(were)Tj
2.126 0 Td
(determined)Tj
4.8472 0 Td
(using)Tj
2.3924 0 Td
(a)Tj
0.652 0 Td
(sigmoidal)Tj
4.1158 0 Td
(dose)Tj
2.041 0 Td
(response)Tj
3.7302 0 Td
(curved)Tj
2.9481 0 Td
(fitted)Tj
-35.4781 -1.2983 Td
(post)Tj
1.9048 0 Td
(normalisation)Tj
5.828 0 Td
(\(with)Tj
2.3358 0 Td
(a)Tj
0.6519 0 Td
(100%)Tj
2.4095 0 Td
(upper)Tj
2.5738 0 Td
(maximal)Tj
3.7077 0 Td
(at)Tj
0.9524 0 Td
(2)Tj
0.6917 0 Td
(mins;)Tj
2.4434 0 Td
(0%)Tj
1.4513 0 Td
(lower)Tj
2.4548 0 Td
(maximal)Tj
3.7076 0 Td
(represented)Tj
4.9096 0 Td
(by)Tj
-36.0223 -1.3039 Td
(the)Tj
1.4683 0 Td
(time)Tj
2.0296 0 Td
(required)Tj
3.6396 0 Td
(for)Tj
1.3833 0 Td
(the)Tj
1.4684 0 Td
(water)Tj
2.4321 0 Td
(control)Tj
3.1238 0 Td
(to)Tj
1.0261 0 Td
(clot)Tj
1.6951 0 Td
(normal)Tj
3.1464 0 Td
(human)Tj
3.0784 0 Td
(citrated)Tj
3.2599 0 Td
(plasma\))Tj
3.3731 0 Td
(with)Tj
2.0012 0 Td
(GraphPad)Tj
-33.1253 -1.2983 Td
(Prism)Tj
2.5965 0 Td
(6)Tj
0.6973 0 Td
(software)Tj
3.583 0 Td
(and)Tj
1.7348 0 Td
(compared)Tj
4.2406 0 Td
(to)Tj
1.0261 0 Td
(those)Tj
2.3357 0 Td
(obtained)Tj
3.7361 0 Td
(for)Tj
1.3833 0 Td
(the)Tj
1.474 0 Td
(heparin)Tj
3.3051 0 Td
(control.)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 288.6236 cm
BT
/F0 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(MTT)Tj
2.2677 0 Td
(cell)Tj
1.6961 0 Td
(proliferation)Tj
5.3717 0 Td
(assay)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 271.6157 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(Potential)Tj
3.7984 0 Td
(toxic)Tj
2.1826 0 Td
(effects)Tj
2.7269 0 Td
(of)Tj
1.0148 0 Td
(sulphated)Tj
4.0819 0 Td
(carbohydrates)Tj
5.862 0 Td
(on)Tj
1.2756 0 Td
(endothelial)Tj
4.6714 0 Td
(cells)Tj
1.9163 0 Td
(were)Tj
2.1259 0 Td
(screened)Tj
3.7191 0 Td
(against)Tj
-33.3749 -1.3039 Td
(utilising)Tj
3.4695 0 Td
(the)Tj
1.4684 0 Td
(tetrazolium)Tj
4.8245 0 Td
(salt,)Tj
1.7858 0 Td
(34,5)Tj
2.3868 0 Td
(dimethylthiazol-2,5)Tj
8.0106 0 Td
(diphenyl)Tj
3.7304 0 Td
(tetrazolium)Tj
4.8245 0 Td
(bromide)Tj
3.6454 0 Td
(\(MTT\))Tj
-34.1459 -1.2982 Td
(proliferation)Tj
5.2667 0 Td
(assay.)Tj
2.4888 0 Td
(Briefly,)Tj
3.0784 0 Td
(a)Tj
0.652 0 Td
(serial)Tj
2.3244 0 Td
(dilution)Tj
3.4072 0 Td
(of)Tj
1.0148 0 Td
(the)Tj
1.4683 0 Td
(test)Tj
1.6044 0 Td
(carbohydrate)Tj
5.4992 0 Td
(was)Tj
1.7008 0 Td
(prepared)Tj
3.804 0 Td
(and)Tj
1.7292 0 Td
(added)Tj
2.6588 0 Td
(to)Tj
-36.697 -1.2983 Td
(HUVEC)Tj
3.651 0 Td
(cells)Tj
1.9218 0 Td
(\(2)Tj
/F16 1 Tf
[()]TJ
/F10 1 Tf
(10)Tj
ET
Q
q
1 j
1 J
0 w
7.4999 0 0 6.6659 279.0992 237.033 cm
BT
/F10 1 Tf
1 TL
-0.0053 Tc
0 0 Td
(4)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 284.8251 232.611 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(cells\))Tj
2.262 0 Td
(in)Tj
1.0375 0 Td
(a)Tj
0.6519 0 Td
(multiwell)Tj
3.9798 0 Td
(plate)Tj
2.1373 0 Td
(\(Greiner\);)Tj
4.2463 0 Td
(a)Tj
0.6577 0 Td
(positive)Tj
3.3108 0 Td
(control,)Tj
3.3505 0 Td
(the)Tj
1.4684 0 Td
(Golgi)Tj
2.4207 0 Td
(disrup-)Tj
-34.0041 -1.2982 Td
(tor)Tj
1.3947 0 Td
(brefeldin)Tj
3.8211 0 Td
(A)Tj
0.907 0 Td
(\(10)Tj
/F16 1 Tf
1.5137 0 Td
[()]TJ
/F10 1 Tf
(l,)Tj
1.1961 0 Td
(10)Tj
1.1736 0 Td
(ng.ml)Tj
ET
Q
q
1 j
1 J
0 w
7.4999 0 0 6.6659 323.0929 224.0503 cm
BT
/F10 1 Tf
1 TL
-0.0053 Tc
0 0 Td
(-1)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 331.4834 219.6283 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(in)Tj
1.0374 0 Td
(PBS\),)Tj
2.3981 0 Td
(and)Tj
1.7291 0 Td
(PBS)Tj
1.8369 0 Td
(\(10)Tj
/F16 1 Tf
1.508 0 Td
[()]TJ
/F10 1 Tf
(l,)Tj
1.2019 0 Td
(as)Tj
1.0148 0 Td
(a)Tj
0.6519 0 Td
(negative)Tj
3.5377 0 Td
(control\))Tj
3.4582 0 Td
(were)Tj
2.126 0 Td
(also)Tj
-33.647 -1.3039 Td
(included.)Tj
3.9345 0 Td
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1.9446 0 Td
(48)Tj
1.1735 0 Td
(hr)Tj
1.1168 0 Td
(incubation,)Tj
4.7792 0 Td
(MTT)Tj
2.33 0 Td
(solution)Tj
3.4866 0 Td
(\(10)Tj
/F16 1 Tf
1.508 0 Td
[()]TJ
/F10 1 Tf
(l,)Tj
1.2019 0 Td
(0.5%)Tj
2.1543 0 Td
(w/v)Tj
1.6838 0 Td
(in)Tj
1.0374 0 Td
(PBS\))Tj
2.177 0 Td
(was)Tj
1.6951 0 Td
(added)Tj
2.6646 0 Td
(to)Tj
1.0261 0 Td
(all)Tj
1.1452 0 Td
(wells)Tj
-35.0586 -1.2982 Td
(for)Tj
1.3833 0 Td
(4)Tj
0.6974 0 Td
(hr)Tj
1.1168 0 Td
(at)Tj
0.9524 0 Td
(37)Tj
1.1736 0 Td
(C.)Tj
1.1112 0 Td
(Finally,)Tj
3.1691 0 Td
(the)Tj
1.4683 0 Td
(supernatants)Tj
5.3403 0 Td
(were)Tj
2.126 0 Td
(discarded,)Tj
4.3086 0 Td
(the)Tj
1.4684 0 Td
(cells)Tj
1.9219 0 Td
(washed)Tj
3.1747 0 Td
(\(PBS\))Tj
2.5172 0 Td
(and)Tj
1.7291 0 Td
(treated)Tj
-33.6583 -1.2983 Td
(with)Tj
1.9956 0 Td
(dimethylsulfoxide)Tj
7.3984 0 Td
(\(10)Tj
/F16 1 Tf
1.5137 0 Td
[()]TJ
/F10 1 Tf
(l\).)Tj
1.5364 0 Td
(Cell)Tj
1.797 0 Td
(proliferation)Tj
5.2668 0 Td
(levels)Tj
2.3754 0 Td
(were)Tj
2.1259 0 Td
(ascertained)Tj
4.7339 0 Td
(indirectly)Tj
4.0478 0 Td
(by)Tj
1.1792 0 Td
(spectro-)Tj
-33.9701 -1.3039 Td
(photometry)Tj
4.9436 0 Td
(at)Tj
0.9525 0 Td
(a)Tj
/F17 1 Tf
0.6519 0 Td
()Tj
/F10 1 Tf
(abs)Tj
2.0183 0 Td
(of)Tj
1.0148 0 Td
(540)Tj
1.6497 0 Td
(nm.)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 139.011 cm
BT
/F0 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(Statistical)Tj
4.4504 0 Td
(analyses)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 122.0031 cm
BT
/F10 1 Tf
1 TL
-0.004 Tc
0 0 Td
(Results)Tj
3.0557 0 Td
(shown)Tj
2.8516 0 Td
(are)Tj
1.4457 0 Td
(the)Tj
1.474 0 Td
(mean)Tj
2.4434 0 Td
(of)Tj
1.0148 0 Td
(two)Tj
1.7121 0 Td
(independent)Tj
5.2724 0 Td
(experiments)Tj
/F16 1 Tf
5.1364 0 Td
[()]TJ
/F10 1 Tf
0.7937 0 Td
(Standard)Tj
3.8324 0 Td
(Deviation)Tj
4.1612 0 Td
(\(SD\).)Tj
2.3358 0 Td
(A)Tj
-35.5292 -1.2982 Td
(standard)Tj
3.719 0 Td
(unpaired)Tj
3.8381 0 Td
(t-test)Tj
2.2507 0 Td
(was)Tj
1.7007 0 Td
(performed)Tj
4.4674 0 Td
(\(Figs)Tj
0.83 0.64 0.02 0 k
2.1827 0 Td
(1)Tj
0 g
(,)Tj
0.83 0.64 0.02 0 k
0.9184 0 Td
(2)Tj
0 g
0.6973 0 Td
(and)Tj
0.83 0.64 0.02 0 k
1.7291 0 Td
(3)Tj
0 g
(\),)Tj
1.2643 0 Td
(ANOVA)Tj
3.7644 0 Td
(\(Kruskal)Tj
3.668 0 Td
(Wallis)Tj
2.7439 0 Td
(with)Tj
1.9956 0 Td
(post-)Tj
-34.9396 -1.3039 Td
(test\))Tj
1.9388 0 Td
(\()Tj
0.83 0.64 0.02 0 k
(Fig)Tj
1.8199 0 Td
(4)Tj
0 g
(\))Tj
1.0318 0 Td
(and)Tj
1.7348 0 Td
(unpaired)Tj
3.8324 0 Td
(t-test)Tj
2.2507 0 Td
(\()Tj
0.83 0.64 0.02 0 k
(Fig)Tj
1.8198 0 Td
(5)Tj
0 g
(\))Tj
1.0318 0 Td
(using)Tj
2.3924 0 Td
(GraphPad)Tj
4.303 0 Td
(Instat3)Tj
2.9764 0 Td
(software)Tj
3.5886 0 Td
(and)Tj
1.7291 0 Td
(considered)Tj
4.5978 0 Td
(signif-)Tj
-35.0473 -1.2983 Td
(icant)Tj
2.1883 0 Td
(when)Tj
/F15 1 Tf
2.4037 0 Td
[()]TJ
/F18 1 Tf
0 Tc
(<)Tj
/F10 1 Tf
-0.004 Tc
(0.05.)Tj
ET
Q
q
1 j
1 J
0 w
576 737.1 m
36 737.1 l
36 737.6 l
576 737.6 l
f*
36 741.2598 107.1496 23.6976 re
W* n
q
107.0929 0 0 23.6409 36 741.3165 cm
q
/I0 Do
Q
Q
Q
q
0 0 612 792 re
W* n
1 j
1 J
0 w
7.9999 0 0 7.9999 464.0314 745.7952 cm
BT
/F0 1 Tf
1 TL
-0.005 Tc
0 0 Td
(GAG)Tj
2.4307 0 Td
(inhibition)Tj
4.0748 0 Td
(of)Tj
1.0418 0 Td
(cytoadher)Tj
4.3157 0 Td
(ence)Tj
ET
Q
q
1 j
1 J
0 w
36 48.0002 m
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