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Margherita Cortini, Annamaria Massa, Sofia Avnet, Gloria Bonuccelli, Nicola Baldini
Tumor-Activated Mesenchymal Stromal Cells Promote Osteosarcoma Stemness and Migratory Potential via IL-6 Secretion
www.plosone.org
www.plosone.org
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(nM\),)Tj
2.228 0 Td
(putrescine)Tj
4.3767 0 Td
(\(10)Tj
1.5136 0 Td
(mg/mL\),)Tj
3.7588 0 Td
(sodium)Tj
3.2314 0 Td
(selenite)Tj
3.2088 0 Td
(\(30)Tj
1.5137 0 Td
(nM\),)Tj
2.2224 0 Td
(apo-)Tj
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(transferrin)Tj
4.5013 0 Td
(\(100)Tj
1.9899 0 Td
(mg/mL\),)Tj
3.7531 0 Td
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1.7348 0 Td
(insulin)Tj
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(\(25)Tj
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(mg/mL\),)Tj
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(\(20)Tj
1.5137 0 Td
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(mL\))Tj
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1.7292 0 Td
(basic)Tj
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(fibroblast)Tj
4.0025 0 Td
(growth)Tj
3.0728 0 Td
(factor)Tj
2.5455 0 Td
(\(10)Tj
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5.4935 0 Td
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1.0318 0 Td
(low-attachment)Tj
6.514 0 Td
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1.389 0 Td
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(6)Tj
0.6916 0 Td
(passages)Tj
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Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 563.9244 cm
BT
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1 TL
-0.0033 Tc
0 0 Td
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4.7764 0 Td
(experiments)Tj
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Q
q
1 j
1 J
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BT
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1 TL
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1.7971 0 Td
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(twice)Tj
2.313 0 Td
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(in)Tj
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1.0375 0 Td
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1.3833 0 Td
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(MEM)Tj
3.1408 0 Td
(plus)Tj
1.8765 0 Td
(0.1%)Tj
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1.0375 0 Td
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-34.1119 -1.3039 Td
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(conditions)Tj
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1.3833 0 Td
(the)Tj
1.4683 0 Td
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3.9799 0 Td
(24)Tj
1.1735 0 Td
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2.5115 0 Td
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1.0261 0 Td
(allow)Tj
2.3414 0 Td
(cytokine)Tj
3.634 0 Td
(secretion)Tj
3.8438 0 Td
(and)Tj
1.7348 0 Td
(accumulation.)Tj
-30.2965 -1.2983 Td
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1.7801 0 Td
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3.9798 0 Td
(day,)Tj
1.8595 0 Td
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1.9219 0 Td
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4.0308 0 Td
(counted)Tj
3.4753 0 Td
(and)Tj
1.7348 0 Td
(seeded)Tj
2.9026 0 Td
(at)Tj
0.9524 0 Td
(a)Tj
0.6577 0 Td
(density)Tj
3.1011 0 Td
(of)Tj
-34.1856 -1.2983 Td
(10,000)Tj
2.8232 0 Td
(cells/mL)Tj
3.6057 0 Td
(in)Tj
1.0375 0 Td
(CSC)Tj
2.0239 0 Td
(complete)Tj
3.8778 0 Td
(medium)Tj
3.6056 0 Td
(on)Tj
1.2756 0 Td
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4.5411 0 Td
(coated)Tj
2.8346 0 Td
(plates)Tj
2.5001 0 Td
(in)Tj
1.0375 0 Td
(the)Tj
1.4683 0 Td
(lower)Tj
2.4492 0 Td
(compart-)Tj
-33.0801 -1.2982 Td
(ment)Tj
2.3073 0 Td
(of)Tj
1.0148 0 Td
(a)Tj
0.6577 0 Td
(boyden)Tj
3.1861 0 Td
(chamber.)Tj
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1.7858 0 Td
(pretreated)Tj
4.3086 0 Td
(MSC)Tj
2.2507 0 Td
(that)Tj
1.7745 0 Td
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(\(or)Tj
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(medium)Tj
3.6056 0 Td
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1.9786 0 Td
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4.5297 0 Td
(transwells\))Tj
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(the)Tj
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(the)Tj
-34.6391 -1.2982 Td
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(boyden)Tj
3.1861 0 Td
(chamber,)Tj
3.9572 0 Td
(and)Tj
1.7291 0 Td
(spheres)Tj
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(were)Tj
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2.2507 0 Td
(for)Tj
1.3833 0 Td
(6)Tj
0.6973 0 Td
(days.)Tj
2.2224 0 Td
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1.3379 0 Td
(assess)Tj
2.5229 0 Td
(sphere)Tj
2.8459 0 Td
(forma-)Tj
-34.0268 -1.2983 Td
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2.0636 0 Td
(bright-field)Tj
4.7621 0 Td
(pictures)Tj
3.4073 0 Td
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(well)Tj
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0.9525 0 Td
(4x)Tj
1.1622 0 Td
(magnification.)Tj
6.0094 0 Td
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1.4003 0 Td
(the)Tj
1.4683 0 Td
(spheres)Tj
3.2031 0 Td
(in)Tj
1.0375 0 Td
(a)Tj
-36.9238 -1.3039 Td
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(well)Tj
1.8199 0 Td
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(counted)Tj
3.4753 0 Td
(and)Tj
1.7291 0 Td
(the)Tj
1.4684 0 Td
(diameter)Tj
3.787 0 Td
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1.0148 0 Td
(each)Tj
2.0296 0 Td
(sphere)Tj
2.846 0 Td
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1.6951 0 Td
(measured)Tj
4.1102 0 Td
(with)Tj
1.9956 0 Td
(ImageJ)Tj
3.033 0 Td
(software.)Tj
-33.6583 -1.2983 Td
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1.3379 0 Td
(define)Tj
2.6986 0 Td
(a)Tj
0.6519 0 Td
(sphere,)Tj
3.0728 0 Td
(only)Tj
1.9729 0 Td
(cell)Tj
1.559 0 Td
(aggregates)Tj
4.3653 0 Td
(with)Tj
1.9956 0 Td
(more)Tj
2.3414 0 Td
(than)Tj
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(50)Tj
/F12 1 Tf
1.1679 0 Td
[()]TJ
/F6 1 Tf
(m)Tj
1.5477 0 Td
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(diameter)Tj
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(\(roughly)Tj
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(corre-)Tj
-33.2218 -1.2982 Td
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1.0261 0 Td
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1.0148 0 Td
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(10)Tj
1.1679 0 Td
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1.0374 0 Td
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1.559 0 Td
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2.8857 0 Td
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(analysis.)Tj
1.1962 -1.2983 Td
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1.3379 0 Td
(evaluate)Tj
3.4639 0 Td
(sphere)Tj
2.846 0 Td
(growth)Tj
3.0727 0 Td
(dependency)Tj
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(IL-6,)Tj
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(neutralizing)Tj
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(anti-IL-6)Tj
3.8268 0 Td
(monoclonal)Tj
5.0059 0 Td
(anti-)Tj
-35.172 -1.2983 Td
(body)Tj
2.2166 0 Td
(\(Tocilizumab,)Tj
5.8053 0 Td
(Roche\))Tj
3.0671 0 Td
(was)Tj
1.6951 0 Td
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2.6646 0 Td
(to)Tj
1.0204 0 Td
(MSC)Tj
2.2564 0 Td
(medium)Tj
3.6056 0 Td
(at)Tj
0.9525 0 Td
(the)Tj
1.4683 0 Td
(final)Tj
2.0013 0 Td
(concentration)Tj
5.8223 0 Td
(of)Tj
1.0148 0 Td
(100)Tj
/F12 1 Tf
1.6441 0 Td
[()]TJ
/F6 1 Tf
(g/)Tj
-35.2344 -1.3039 Td
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4.3426 0 Td
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3.4469 0 Td
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1.474 0 Td
(antibody)Tj
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(every)Tj
2.347 0 Td
(24)Tj
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(hours.)Tj
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(repli-)Tj
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ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 260.4472 cm
BT
/F0 1 Tf
1 TL
-0.0033 Tc
0 0 Td
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4.5118 0 Td
(profiler)Tj
3.2315 0 Td
(array)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 243.4393 cm
BT
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1 TL
-0.004 Tc
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1.7802 0 Td
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-34.4237 -1.2982 Td
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1.9956 0 Td
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1.559 0 Td
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1.9332 0 Td
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(quanti-)Tj
-34.4634 -1.2983 Td
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1.7234 0 Td
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2.0013 0 Td
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4.1272 0 Td
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3.2541 0 Td
(Human)Tj
3.3165 0 Td
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4.7849 0 Td
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2.2393 0 Td
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1.7972 0 Td
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2.5342 0 Td
(Kit)Tj
1.4626 0 Td
(\(R&D\))Tj
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1.9105 0 Td
(buffer.)Tj
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1.0148 0 Td
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(\(200)Tj
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1.9899 0 Td
[()]TJ
/F6 1 Tf
(g\))Tj
1.5307 0 Td
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2.1203 0 Td
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1.2756 0 Td
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-33.7774 -1.304 Td
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4.7905 0 Td
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4.439 0 Td
(to)Tj
1.0261 0 Td
(manufacturer's)Tj
6.2646 0 Td
(instructions,)Tj
5.2213 0 Td
(each)Tj
2.0296 0 Td
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3.0218 0 Td
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1.7801 0 Td
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1.0148 0 Td
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4.3086 0 Td
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1.1792 0 Td
(dedicated)Tj
4.0762 0 Td
(software)Tj
3.583 0 Td
(for)Tj
1.3889 0 Td
(densitometric)Tj
5.7543 0 Td
(evaluation)Tj
-31.2602 -1.2983 Td
(\(VisionWorksLS)Tj
6.9164 0 Td
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3.5773 0 Td
(Software,)Tj
3.9288 0 Td
(Biospectrum,)Tj
5.5446 0 Td
(UVP\).)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
(Immunofluorescenc)Tj
8.6315 0 Td
(e)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 122.0031 cm
BT
/F6 1 Tf
1 TL
-0.004 Tc
0 0 Td
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1.627 0 Td
(Ki67)Tj
2.109 0 Td
(staining,)Tj
3.634 0 Td
(HOS-CSC)Tj
4.3653 0 Td
(spheres)Tj
3.2031 0 Td
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2.126 0 Td
(trypsinized)Tj
4.6658 0 Td
(and)Tj
1.7348 0 Td
(counted.)Tj
3.6964 0 Td
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2.6475 0 Td
(cells)Tj
1.9219 0 Td
(were)Tj
2.1203 0 Td
(centri-)Tj
-33.8511 -1.2982 Td
(fuged)Tj
2.4491 0 Td
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0.9524 0 Td
(8,000)Tj
2.3471 0 Td
(rpm)Tj
1.9332 0 Td
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1.3833 0 Td
(8)Tj
0.6973 0 Td
(min)Tj
1.8539 0 Td
(with)Tj
1.9955 0 Td
(Cyto-Tek)Tj
4.0252 0 Td
(centrifuge)Tj
4.2576 0 Td
(\(Electron)Tj
3.9515 0 Td
(Microscopy)Tj
4.9436 0 Td
(Science\))Tj
3.5433 0 Td
(and,)Tj
-34.333 -1.3039 Td
(next,)Tj
2.1769 0 Td
(fixed)Tj
2.194 0 Td
(in)Tj
1.0375 0 Td
(3.7%)Tj
2.1543 0 Td
(paraformaldehyde)Tj
7.5231 0 Td
(20)Tj
1.1736 0 Td
(min.)Tj
2.0806 0 Td
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2.16 0 Td
(were)Tj
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