%PDF-1.4 % 8 0 obj <> endobj 56 0 obj <>stream /Artifact <>BDC q 1 i 0 841.89 595.276 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n 0 0 0 0 k /GS1 gs 0 466.055 595.2 375.985 re f EMC /Figure <>BDC Q q 1 i -8.504 850.394 603.78 -384.339 re W* n 0 841.89 595.276 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n /GS1 gs q 600.212 0 0 378.07 -2.5121 466.476 cm /Im1 Do Q EMC /Artifact <>BDC Q q 1 i 0 841.89 595.276 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n 0 .68 .9 0 k /GS1 gs 0 463.221 595.2 5.669 re f EMC /P <>BDC /GS2 gs BT /F3 1 Tf 7 0 0 7 42.5196 28.6398 Tm 0 0 0 1 k 0 Tc 0 Tw (26)Tj ET EMC /P <>BDC BT /F4 1 Tf 7 0 0 7 50.8077 28.6398 Tm ( )Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 42.5196 439.3701 Tm ( )Tj 0 0 TD [(T)71.8(oxicology testing is per)-12(formed to understand the adverse effects of)]TJ ET EMC /P <>BDC BT 8 0 0 8 283.0091 439.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 427.3701 Tm (drugs and chemical substances on humans and other living)Tj ET EMC /P <>BDC BT 8 0 0 8 252.5436 427.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 415.3701 Tm (organisms. In the EU, the Registration, Evaluation, Authorisation and)Tj ET EMC /P <>BDC BT 8 0 0 8 284.1516 415.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 403.3701 Tm (Restriction of Chemicals \(REACH\) regulation \(EC 1907/2006\) applies)Tj ET EMC /P <>BDC BT 8 0 0 8 284.0237 403.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 391.3701 Tm (a no data, no market rule, and responsibility has been placed on)Tj ET EMC /P <>BDC BT 8 0 0 8 277.8877 391.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 379.3701 Tm [(industr)-12(y to manage the risks from new chemicals and to provide)]TJ ET EMC /P <>BDC BT 8 0 0 8 270.2473 379.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 367.3701 Tm (safety information on them.)Tj ET EMC /P <>BDC BT 8 0 0 8 141.2717 367.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 355.3702 Tm ( )Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 9 0 0 9 42.5196 343.3702 Tm 0 .68 .9 0 k -.015 Tc [(W)23.9(h)0(a)23.9(t)0( is 3D cell culture?)]TJ ET EMC /P <>BDC BT 9 0 0 9 145.0222 343.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC /GS2 gs BT /F2 1 Tf 8 0 0 8 42.5196 331.3702 Tm 0 0 0 1 k (3D cell culture \()Tj ET EMC /P <>BDC BT /F5 1 Tf 8 0 0 8 98.9037 331.3702 Tm (Figure 1)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 129.3117 331.3702 Tm (\) describes a number of techniques to grow)Tj ET EMC /P <>BDC BT 8 0 0 8 283.6876 331.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 319.3702 Tm (cells in three dimensions, such as in a spheroid, using an artificially)Tj ET EMC /P <>BDC BT 8 0 0 8 277.5597 319.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 307.3702 Tm (created microenvironment. Cells in 3D cell culture have)Tj ET EMC /P <>BDC BT 8 0 0 8 239.0477 307.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 295.3702 Tm (physiological cell-cell interactions and cell-extracellular matrix)Tj ET EMC /P <>BDC BT 8 0 0 8 260.3678 295.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 283.3702 Tm (component interactions, which allow cells to grow )Tj ET EMC /P <>BDC BT /F6 1 Tf 8 0 0 8 223.5757 283.3702 Tm (in vitro)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 247.0317 283.3702 Tm ( in an)Tj ET EMC /P <>BDC BT 8 0 0 8 266.8877 283.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 271.3702 Tm (environment that closely resembles )Tj ET EMC /P <>BDC BT /F6 1 Tf 8 0 0 8 169.5196 271.3702 Tm (in vivo)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 191.3517 271.3702 Tm ( conditions. For example,)Tj ET EMC /P <>BDC BT 8 0 0 8 280.9597 271.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 259.3702 Tm (spheres possess a hypoxic \(oxygen-deprived\) core resembling solid)Tj ET EMC /P <>BDC BT 8 0 0 8 279.1437 259.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 247.3702 Tm (tumours with cells at the centre of the sphere having extremely low)Tj ET EMC /P <>BDC BT 8 0 0 8 281.5996 247.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 235.3702 Tm (oxygen and nutrient concentrations. Spheres also show complex)Tj ET EMC /P <>BDC BT 8 0 0 8 271.2957 235.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 223.3702 Tm (transport dynamics by creating diffusion gradients of drugs, oxygen,)Tj ET EMC /P <>BDC BT 8 0 0 8 284.8396 223.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 211.3702 Tm (nutrients and waste, and also show resistance and drugs/chemical)Tj ET EMC /P <>BDC BT 8 0 0 8 275.9756 211.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 199.3702 Tm (substances show low potency as an )Tj ET EMC /P <>BDC BT /F6 1 Tf 8 0 0 8 170.0876 199.3702 Tm (in vivo)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 191.9197 199.3702 Tm ( condition. The neighbour)Tj ET EMC /P <>BDC BT 8 0 0 8 284.7676 199.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 187.3702 Tm [(geometr)-12(y and cellular support found in 3D cell cultures can improve)]TJ ET EMC /P <>BDC BT 8 0 0 8 284.4954 187.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 175.3702 Tm (gene expression, cellular communication, migration, differentiation,)Tj ET EMC /P <>BDC BT 8 0 0 8 282.6797 175.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 163.3702 Tm [(sur)-12(vival and growth similar to )]TJ ET EMC /P <>BDC BT /F6 1 Tf 8 0 0 8 149.5514 163.3702 Tm (in vivo)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 171.3834 163.3702 Tm ( tissues, which provide better)Tj ET EMC /P <>BDC BT 8 0 0 8 274.7914 163.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 151.3702 Tm (representation for toxicological testing. Mixed-cell populations can)Tj ET EMC /P <>BDC BT 8 0 0 8 278.9357 151.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 139.3702 Tm (also be cultured in 3D to closely model human tissues. Hence, 3D)Tj ET EMC /P <>BDC BT 8 0 0 8 274.7197 139.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 127.3702 Tm (cell culture plays a vital role in measuring biological responses to)Tj ET EMC /P <>BDC BT 8 0 0 8 270.6157 127.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 115.3702 Tm (new chemicals.)Tj ET EMC /P <>BDC BT 8 0 0 8 96.5836 115.3702 Tm ( )Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 9 0 0 9 42.5196 103.3702 Tm 0 .68 .9 0 k ( )Tj ET EMC /P <>BDC BT 9 0 0 9 42.5196 91.3702 Tm -.015 Tc [(W)23.9(h)36.9(y should I use it?)]TJ ET EMC /P <>BDC BT 9 0 0 9 131.1267 91.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC /GS2 gs BT /F2 1 Tf 8 0 0 8 42.5196 79.3702 Tm 0 0 0 1 k -.005 Tc [(Use of animals in toxicological resear)17.8(ch has been reduced due to the)]TJ ET EMC /P <>BDC BT 8 0 0 8 282.3466 79.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5196 67.3702 Tm -.005 Tc (ethical concerns, expense, time consumption, and misleading results)Tj ET EMC /P <>BDC BT 8 0 0 8 282.9211 67.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 439.3701 Tm -.005 Tc [(due to differences between human and animal physiology)74(. The)]TJ ET EMC /P <>BDC BT 8 0 0 8 529.5779 439.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 427.3701 Tm -.005 Tc [(REACH regulation)54.9(s stated aim is: to ensure a high level of protection)]TJ ET EMC /P <>BDC BT 8 0 0 8 552.6825 427.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 415.3701 Tm -.005 Tc (of human health and the environment from effects of hazardous)Tj ET EMC /P <>BDC BT 8 0 0 8 534.0016 415.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 403.3701 Tm -.005 Tc (chemicals. It strives for a balance: to increase our understanding of)Tj ET EMC /P <>BDC BT 8 0 0 8 541.9619 403.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 391.3701 Tm -.005 Tc (the possible hazards of chemicals, while at the same time avoiding)Tj ET EMC /P <>BDC BT 8 0 0 8 541.4258 391.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 379.3701 Tm -.005 Tc [(unnecessar)-12(y testing on animals \(European Chemicals Agency)74(,)]TJ ET EMC /P <>BDC BT 8 0 0 8 527.6655 379.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 367.3701 Tm -.005 Tc (2020\). 3D cell culture is a much better technique, which supports)Tj ET EMC /P <>BDC BT 8 0 0 8 539.3778 367.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 355.3702 Tm -.005 Tc [(the 3Rs of animal resear)17.8(ch \(replacement, reduction and refinement\))]TJ ET EMC /P <>BDC BT 8 0 0 8 550.3712 355.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 343.3702 Tm -.005 Tc [(and the REACH recommendation to per)-12(form humane animal)]TJ ET EMC /P <>BDC BT 8 0 0 8 522.0091 343.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 331.3702 Tm -.005 Tc [(toxicology resear)17.8(ch.)]TJ ET EMC /P <>BDC BT 8 0 0 8 378.9863 331.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 319.3702 Tm -.015 Tc (Conventional 2D cell cultures \()Tj ET EMC /P <>BDC BT /F5 1 Tf 8 0 0 8 414.2725 319.3702 Tm (Figure 1)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 443.7206 319.3702 Tm (\) are unable to detect organ-)Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 307.3702 Tm (specific toxicity and have inadequate representation of cell migration,)Tj ET EMC /P <>BDC BT 8 0 0 8 546.6487 307.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 295.3702 Tm -.015 Tc [(differentiation, signal transduction, sur)-12(vival and growth. A 2D cell)]TJ ET EMC /P <>BDC BT 8 0 0 8 532.9764 295.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 283.3702 Tm -.015 Tc (culture does not reveal toxicological resistance \()Tj ET EMC /P <>BDC BT /F5 1 Tf 8 0 0 8 472.3606 283.3702 Tm (Figure 2)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 501.8087 283.3702 Tm [(\), ar)17.8(chitecture)]TJ ET EMC /P <>BDC BT 8 0 0 8 549.3221 283.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 271.3702 Tm -.015 Tc (as )Tj ET EMC /P <>BDC BT /F6 1 Tf 8 0 0 8 318.0244 271.3702 Tm (in vivo)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 339.0164 271.3702 Tm ( tissues, accurate depiction of cell polarisation or gene)Tj ET EMC /P <>BDC BT 8 0 0 8 523.1768 271.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 259.3702 Tm -.015 Tc (expression. It also provides unreliable predictions of )Tj ET EMC /P <>BDC BT /F6 1 Tf 8 0 0 8 487.0726 259.3702 Tm (in vivo)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 508.0647 259.3702 Tm ( drug)Tj ET EMC /P <>BDC BT 8 0 0 8 526.6328 259.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 247.3702 Tm -.015 Tc [(efficiency and toxicity)74(, which leads to low success rates in clinical trials.)]TJ ET EMC /P <>BDC BT 8 0 0 8 549.9451 247.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 235.3702 Tm -.015 Tc [(3D cell culture can over)17.8(come the disadvantages of )]TJ ET EMC /P <>BDC BT /F6 1 Tf 8 0 0 8 483.666 235.3702 Tm (in vivo)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 504.6581 235.3702 Tm ( animal)Tj ET EMC /P <>BDC BT 8 0 0 8 529.7542 235.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 223.3702 Tm -.015 Tc (testing and conventional 2D cell culture by providing a more accurate)Tj ET EMC /P <>BDC BT 8 0 0 8 548.6407 223.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 211.3702 Tm -.015 Tc (platform for short- and long-term studies, demonstrating the long-)Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 199.3702 Tm (term effects of the drugs.)Tj ET EMC /P <>BDC BT 8 0 0 8 395.0885 199.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 187.3702 Tm ( )Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 9 0 0 9 308.9764 175.3702 Tm 0 .68 .9 0 k -.015 Tc [(Ho)23.9(w is it done?)]TJ ET EMC /P <>BDC BT 9 0 0 9 375.0821 175.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC /GS2 gs BT /F2 1 Tf 8 0 0 8 308.9764 163.3702 Tm 0 0 0 1 k (Different 3D culture techniques such as anchorage independent,)Tj ET EMC /P <>BDC BT 8 0 0 8 537.4963 163.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 151.3702 Tm (anchorage dependent, hydrogels based and specialised culture)Tj ET EMC /P <>BDC BT 8 0 0 8 531.6724 151.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 139.3702 Tm (platforms can be used.)Tj ET EMC /P <>BDC BT 8 0 0 8 389.4404 139.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 127.3702 Tm (Anchorage-independent/scaffold-free techniques rely on non-)Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 115.3702 Tm (adherent cell-to-cell aggregation to form spheroids, which show)Tj ET EMC /P <>BDC BT 8 0 0 8 536.2163 115.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 103.3702 Tm (cell-cell interactions and secrete their own extracellular matrix. )Tj ET EMC /P <>BDC BT 8 0 0 8 532.0645 103.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 91.3702 Tm (These spheroids are formed without a physical support resulting )Tj ET EMC /P <>BDC BT 8 0 0 8 537.8643 91.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 79.3702 Tm (in consistency of shape and size, which provides better)Tj ET EMC /P <>BDC BT 8 0 0 8 502.8724 79.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9764 67.3702 Tm [(understanding about cellular cytotoxicity)74(. Low-adhesion plates )]TJ ET EMC /P <>BDC BT 8 0 0 8 532.3366 67.3702 Tm ( )Tj ET EMC /Artifact <>BDC .06 .07 .09 .15 K 0 J 0 j .5 w 10 M [] 0 d /GS1 gs q 1 0 0 1 297.638 452.37 cm 0 0 m 0 -384.339 l S Q EMC /P <>BDC BT /F4 1 Tf 16 0 0 16 42.5294 784.3701 Tm 0 0 0 0 k -.015 Tc (FOOD)Tj ET EMC /P <>BDC BT /F7 1 Tf 20 0 0 20 86.8654 784.3701 Tm .06 .07 .09 .15 k 0 Tc ( )Tj ET EMC /P <>BDC /GS2 gs BT /F4 1 Tf 7 0 0 7 68.0315 28.6398 Tm 0 0 0 1 k (TRESEARCH)Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 7 0 0 7 109.1284 28.6398 Tm .78 .02 .98 .09 k ( | )Tj ET EMC /P <>BDC /GS2 gs BT /F4 1 Tf 7 0 0 7 114.8265 28.6398 Tm 0 0 0 1 k (SPRING 2020)Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 7 0 0 7 157.7505 28.6398 Tm .78 .02 .98 .09 k ( | )Tj ET EMC /P <>BDC /GS2 gs BT /F4 1 Tf 7 0 0 7 163.4485 28.6398 Tm 0 0 0 1 k [(V)36.9(OLUME 15 : NUMBER 1)]TJ ET EMC /Artifact <>BDC 0 .68 .9 0 k /GS1 gs 42.52 777.161 42.519 1.418 re f EMC /P <>BDC BT /F1 1 Tf 35 0 0 35 42.5196 708.2772 Tm (3D mammalian)Tj ET EMC /P <>BDC BT 35 0 0 35 308.8697 708.2772 Tm ( )Tj ET EMC /P <>BDC BT 35 0 0 35 42.5196 665.4793 Tm [(cell cultur)12(e)]TJ ET EMC /P <>BDC BT 35 0 0 35 233.936 665.4793 Tm ( )Tj ET EMC /P <>BDC BT 35 0 0 35 42.5196 622.6814 Tm (models in)Tj ET EMC /P <>BDC BT 35 0 0 35 212.3747 622.6814 Tm ( )Tj ET EMC /P <>BDC BT 35 0 0 35 42.5196 579.8835 Tm [(to)5.9(xicolo)5.9(gy testing)]TJ ET EMC /P <>BDC BT 35 0 0 35 346.2596 579.8835 Tm ( )Tj ET EMC /P <>BDC /GS2 gs BT 12.87 0 0 13 42.5161 562.8835 Tm 0 0 0 1 k ( )Tj ET EMC /P <>BDC /GS1 gs BT 12.87 0 0 13 42.5161 545.8836 Tm 0 0 0 0 k -.015 Tc [(3D cell cultur)11.9(e can be successfull)11.9(y used as an)]TJ ET EMC /P <>BDC BT 12.87 0 0 13 319.8411 545.8836 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 12.87 0 0 13 42.5161 528.8836 Tm -.015 Tc [(alternati)11.9(v)11.9(e)0( to la)11.9(bor)17.8(ator)11.9(y animals,)36.8( and as a cost-)]TJ ET EMC /P <>BDC BT 12.87 0 0 13 42.5161 511.8836 Tm [(effecti)11.9(v)11.9(e)0( and time-sa)29.7(ving tissue cultur)11.9(e tec)17.8(hnique)17.8(,)]TJ ET EMC /P <>BDC BT 12.87 0 0 13 351.9643 511.8836 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 12.87 0 0 13 42.5161 494.8837 Tm -.015 Tc [(whic)17.8(h also r)11.9(educes the trial period for drug testing.)]TJ ET EMC Q endstream endobj 57 0 obj <>/XObject<>/ExtGState<>>> endobj 136 0 obj <> endobj 140 0 obj <> endobj 144 0 obj <> endobj 148 0 obj <> endobj 152 0 obj <> endobj 188 0 obj <>stream /P <>BDC q 1 i 0 841.89 595.275 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n /GS2 gs BT /F4 1 Tf 7 0 0 7 542.5214 28.6398 Tm 0 0 0 1 k 0 Tc 0 Tw ( )Tj ET EMC /P <>BDC BT /F3 1 Tf 7 0 0 7 544.4675 28.6398 Tm (27)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 42.5283 571.3701 Tm (are specialised culture plates with ultra-low attachment coating)Tj ET EMC /P <>BDC BT 8 0 0 8 266.1764 571.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 559.3701 Tm [(\(polyhydroxyethylmethacr)-12(ylate/agarose\), which promote cell)]TJ ET EMC /P <>BDC BT 8 0 0 8 259.024 559.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 547.3701 Tm (aggregation to form spheroids. Hanging drop plates are open)Tj ET EMC /P <>BDC BT 8 0 0 8 261.1043 547.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 535.3701 Tm (bottomless wells, which promote the formation of droplets of media)Tj ET EMC /P <>BDC BT 8 0 0 8 284.3443 535.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 523.3701 Tm (that provide space to form spheroids. In magnetic levitation, cells)Tj ET EMC /P <>BDC BT 8 0 0 8 273.8163 523.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 511.3701 Tm (are preloaded with magnetic nanoparticles and external magnetic)Tj ET EMC /P <>BDC BT 8 0 0 8 275.9362 511.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 499.3701 Tm (fields to provide non-adhesion, plate-like properties to form uniform)Tj ET EMC /P <>BDC BT 8 0 0 8 283.5123 499.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 487.3701 Tm (tumourspheres. A rotational stirrer has a container to hold cells )Tj ET EMC /P <>BDC BT 8 0 0 8 267.0004 487.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 475.3701 Tm [(and impeller)49.8(, stirring continuously)74(. Liquid flow prevents cell)]TJ ET EMC /P <>BDC BT 8 0 0 8 252.7221 475.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 463.3701 Tm (adhesion and distributes nutrition and oxygen uniformly to )Tj ET EMC /P <>BDC BT 8 0 0 8 254.0963 463.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 451.3701 Tm (form tumourspheres.)Tj ET EMC /P <>BDC BT 8 0 0 8 117.0004 451.3701 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 439.3701 Tm .01 Tc (The anchorage-dependent technique provides physical )Tj ET EMC /P <>BDC BT 8 0 0 8 243.3981 439.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 427.3701 Tm .01 Tc (support by using engineered porous membranes, polymeric fabric)Tj ET EMC /P <>BDC BT 8 0 0 8 281.5416 427.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 415.3701 Tm .01 Tc (meshes called scaffolds; these scaffolds can be made of natural or)Tj ET EMC /P <>BDC BT 8 0 0 8 283.4133 415.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 403.3701 Tm .01 Tc (synthetic components. This physical support can provide structures)Tj ET EMC /P <>BDC BT 8 0 0 8 285.5174 403.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 391.3701 Tm .01 Tc (from simple mechanical up to extracellular matrix-like structures.)Tj ET EMC /P <>BDC BT 8 0 0 8 276.8934 391.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 379.3701 Tm .01 Tc (Cells are embedded in extracellular components and are able to)Tj ET EMC /P <>BDC BT 8 0 0 8 273.6776 379.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 367.3701 Tm .01 Tc (initiate cell-cell and cell-matrix interactions, and physical support)Tj ET EMC /P <>BDC BT 8 0 0 8 277.0934 367.3701 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 355.3702 Tm .01 Tc (for cell growth, adhesion and proliferation. Natural scaffolds have)Tj ET EMC /P <>BDC BT 8 0 0 8 279.3734 355.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 343.3702 Tm .01 Tc (higher biocompatibility and lower toxicity when compared to)Tj ET EMC /P <>BDC BT 8 0 0 8 265.4697 343.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 331.3702 Tm .01 Tc (synthetic polymers.)Tj ET EMC /P <>BDC BT 8 0 0 8 112.7275 331.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 319.3702 Tm -.01 Tc [(Hydrogels can be defined as water-swollen networks of a polymer)49.8(,)]TJ ET EMC /P <>BDC BT 8 0 0 8 271.7246 319.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 307.3702 Tm -.01 Tc (which is a liquid at room temperature and forms a gel-like structure at)Tj ET EMC /P <>BDC BT 8 0 0 8 284.6994 307.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 295.3702 Tm -.01 Tc (37C. Cells can be embedded inside hydrogels and provide a similar)Tj ET EMC /P <>BDC BT 8 0 0 8 278.2752 295.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 283.3702 Tm -.01 Tc (microenvironment to an extracellular matrix. Animal- and plant-)Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 271.3702 Tm (derived hydrogels can be used in 3D cell culture. Specialised 3D cell)Tj ET EMC /P <>BDC BT 8 0 0 8 277.1794 271.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 259.3702 Tm -.01 Tc (culture platforms, such as microfluidic devices, can equally distribute)Tj ET EMC /P <>BDC BT 8 0 0 8 279.3075 259.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 247.3702 Tm -.01 Tc (oxygen and nutrients, while removing waste to facilitate spheroid)Tj ET EMC /P <>BDC BT 8 0 0 8 269.2111 247.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 235.3702 Tm -.01 Tc (formation. Micro-patterned plates are micro-space, low-adhesion)Tj ET EMC /P <>BDC BT 8 0 0 8 267.5551 235.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 223.3702 Tm -.01 Tc (plates that promote the formation of spheres.)Tj ET EMC /P <>BDC BT 8 0 0 8 200.3463 223.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 211.3702 Tm ( )Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 9 0 0 9 42.5283 199.3702 Tm 0 .68 .9 0 k -.015 Tc (Conclusion)Tj ET EMC /P <>BDC BT 9 0 0 9 90.8674 199.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC /GS2 gs BT /F2 1 Tf 8 0 0 8 42.5283 187.3702 Tm 0 0 0 1 k (3D cell culture can be successfully used as an alternative to)Tj ET EMC /P <>BDC BT 8 0 0 8 250.3283 187.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 175.3702 Tm [(laborator)-12(y animals, as a cost-effective and time-saving tissue culture)]TJ ET EMC /P <>BDC BT 8 0 0 8 281.928 175.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 163.3702 Tm (technique, and also to reduce trial periods for drug testing.)Tj ET EMC /P <>BDC BT 8 0 0 8 251.5363 163.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 151.3702 Tm [(Moreover)49.8(, they can be used as an alternative to 2D cell culture, since)]TJ ET EMC /P <>BDC BT 8 0 0 8 286.002 151.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 139.3702 Tm (they give an accurate outcome for toxicological testing. 3D cell)Tj ET EMC /P <>BDC BT 8 0 0 8 266.3763 139.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 127.3702 Tm (culture reduces the gap between )Tj ET EMC /P <>BDC BT /F6 1 Tf 8 0 0 8 161.3123 127.3702 Tm (in vitro)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 184.7682 127.3702 Tm ( and )Tj ET EMC /P <>BDC BT /F6 1 Tf 8 0 0 8 202.8002 127.3702 Tm (in vivo)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 224.6323 127.3702 Tm ( drug testing and)Tj ET EMC /P <>BDC BT 8 0 0 8 285.8643 127.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 115.3702 Tm (its effects at the clinical level.)Tj ET EMC /P <>BDC BT 8 0 0 8 144.7364 115.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 103.3702 Tm ( )Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 9 0 0 9 42.5283 91.3702 Tm 0 .68 .9 0 k -.015 Tc [(Ac)17.8(kno)23.9(wledgment)]TJ ET EMC /P <>BDC BT 9 0 0 9 118.4456 91.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC /GS2 gs BT /F2 1 Tf 8 0 0 8 42.5283 79.3702 Tm 0 0 0 1 k -.015 Tc (This study has been supported by Science Foundation Ireland \(SFI\))Tj ET EMC /P <>BDC BT 8 0 0 8 269.9207 79.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 42.5283 67.3702 Tm -.015 Tc (under grant number 17/CDA/4653.)Tj ET EMC /P <>BDC BT 8 0 0 8 165.8564 67.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 379.3701 Tm ( )Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 9 0 0 9 308.9851 367.3701 Tm 0 .68 .9 0 k ( )Tj ET EMC /P <>BDC BT 9 0 0 9 308.9851 355.3702 Tm -.015 Tc (Reference)Tj ET EMC /P <>BDC BT 9 0 0 9 351.1232 355.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC /GS2 gs BT /F2 1 Tf 8 0 0 8 308.9851 343.3702 Tm 0 0 0 1 k [(European Chemicals Agency)74(. \(2020\). Animal testing under REACH )]TJ ET EMC /P <>BDC BT 8 0 0 8 551.1692 343.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 331.3702 Tm [(ECHA\). [Accessed Februar)-12(y 23, 2020]. A)23.9(vailable from:)]TJ ET EMC /P <>BDC BT 8 0 0 8 500.4254 331.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 319.3702 Tm (https://echa.europa.eu/animal-testing-under-reach.)Tj ET EMC /P <>BDC BT 8 0 0 8 491.449 319.3702 Tm ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 307.3702 Tm ( )Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 9 0 0 9 308.9851 295.3702 Tm 0 .68 .9 0 k -.015 Tc [(A)29.8(uthors)]TJ ET EMC /P <>BDC BT 9 0 0 9 343.781 295.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC /GS2 gs BT /F5 1 Tf 8 0 0 8 308.9851 283.3702 Tm 0 0 0 1 k -.015 Tc [(Janith W)39.8(anigasekara)]TJ ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 382.9467 283.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 271.3702 Tm -.015 Tc [(Doctoral resear)17.8(cher)49.8(, T)71.8(eagasc and School of Food Science and)]TJ ET EMC /P <>BDC BT 8 0 0 8 518.1101 271.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 259.3702 Tm -.015 Tc [(Environmental Health, T)71.8(echnological University Dublin)]TJ ET EMC /P <>BDC BT 8 0 0 8 495.1471 259.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 247.3702 Tm ( )Tj ET EMC /P <>BDC BT /F5 1 Tf 8 0 0 8 308.9851 235.3702 Tm -.015 Tc [(Brijesh T)19.8(iwari)]TJ ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 357.7868 235.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 223.3702 Tm -.015 Tc [(Resear)17.8(ch Officer)49.8(, T)71.8(eagasc Food Resear)17.8(ch Centre, Ashtown, Dublin 15)]TJ ET EMC /P <>BDC BT 8 0 0 8 544.3675 223.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8.5 0 0 8.5 308.9851 211.3702 Tm (Correspondence: )Tj ET EMC /P <>BDC BT 8 0 0 8 375.7355 211.3702 Tm -.015 Tc (brijesh.tiwari@teagasc.ie)Tj ET EMC /P <>BDC BT 8 0 0 8 458.9756 211.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 199.3702 Tm ( )Tj ET EMC /P <>BDC BT /F5 1 Tf 8 0 0 8 308.9851 187.3702 Tm -.015 Tc (James Curtin)Tj ET EMC /P <>BDC BT /F2 1 Tf 8 0 0 8 355.2171 187.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 175.3702 Tm -.015 Tc (Head of the School of Food Science and Environmental Health,)Tj ET EMC /P <>BDC BT 8 0 0 8 524.2334 175.3702 Tm 0 Tc ( )Tj ET EMC /P <>BDC BT 8 0 0 8 308.9851 163.3702 Tm -.015 Tc [(T)71.8(echnological University Dublin)]TJ ET EMC /P <>BDC BT 8 0 0 8 416.787 163.3702 Tm 0 Tc ( )Tj ET EMC /Artifact <>BDC .06 .07 .09 .15 K 0 J 0 j .5 w 10 M [] 0 d /GS1 gs q 1 0 0 1 297.594 573.242 cm 0 0 m 0 -505.194 l S Q EMC /P <>BDC BT /F4 1 Tf 16 0 0 16 508.206 787.0821 Tm .06 .07 .09 .15 k -.015 Tc (FOOD)Tj ET EMC /P <>BDC /GS2 gs BT 7 0 0 7 350.057 28.6398 Tm 0 0 0 1 k 0 Tc (TRESEARCH)Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 7 0 0 7 391.154 28.6398 Tm .78 .02 .98 .09 k ( | )Tj ET EMC /P <>BDC /GS2 gs BT /F4 1 Tf 7 0 0 7 396.8518 28.6398 Tm 0 0 0 1 k (SPRING 2020)Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 7 0 0 7 439.776 28.6398 Tm .78 .02 .98 .09 k ( | )Tj ET EMC /P <>BDC /GS2 gs BT /F4 1 Tf 7 0 0 7 445.474 28.6398 Tm 0 0 0 1 k [(V)36.9(OLUME 15 : NUMBER 1)]TJ ET EMC /Artifact <>BDC 0 .68 .9 0 k /GS1 gs 510.244 777.161 42.519 1.418 re f EMC /P <>BDC /GS2 gs BT /F8 1 Tf 7 0 0 7 42.5193 595.0492 Tm 0 0 0 1 k [(FIGURE 1: 2D \(left\) and 3D \(right\) cell cultur)17.8(es of U373MG cells)17.8(.)]TJ ET EMC /P <>BDC BT 7 0 0 7 242.0707 595.0492 Tm ( )Tj ET EMC /P <>BDC BT 7 0 0 7 308.724 383.5138 Tm [(FIGURE 2: Differ)17.8(ent concentr)29.8(ations of dimeth)17.8(yl sulf)17.8(oxide \(DMSO\) c)17.8(ytotoxicity)]TJ ET EMC /P <>BDC BT 7 0 0 7 547.9834 383.5138 Tm ( )Tj ET EMC /P <>BDC BT 7 0 0 7 308.724 372.5138 Tm [(tow)12(ar)29.8(ds 3D and 2D cell cultur)17.8(e)17.8(.)]TJ ET EMC /Artifact <>BDC 0 .136 .18 0 k /GS1 gs 308.976 68.032 51.024 62.362 re f EMC /Figure <>BDC Q q 1 i 308.976 130.394 51.024 -62.362 re W* n 0 841.89 595.275 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n /GS1 gs q 53.757 0 0 65.138 307.627 66.56 cm /Im3 Do Q EMC /Figure <>BDC Q q 1 i 365.257 130.394 51.024 -62.362 re W* n 0 841.89 595.275 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n /GS1 gs q 54.223 0 0 64.933 363.764 66.528 cm /Im5 Do Q EMC /Artifact <>BDC Q q 1 i 0 841.89 595.275 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n 0 .136 .18 0 k /GS1 gs 421.539 68.032 51.023 62.362 re f EMC /Figure <>BDC Q q 1 i 421.539 130.394 51.023 -62.362 re W* n 0 841.89 595.275 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n /GS1 gs q 54.228 0 0 65.141 420.221 66.559 cm /Im7 Do Q EMC /Artifact <>BDC Q q 1 i 0 841.89 595.275 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n 0 0 0 0 k /GS1 gs 42.519 607.69 243.78 150.4 re f EMC /Figure <>BDC Q q 1 i 42.519 758.09 243.78 -150.4 re W* n 0 841.89 595.275 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n /GS1 gs q 246.802 0 0 152.274 41.232 607.295 cm /Im9 Do Q EMC /Artifact <>BDC Q q 1 i 0 841.89 595.275 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n 0 0 0 0 k /GS1 gs 308.976 607.69 243.779 150.4 re f EMC /Figure <>BDC Q q 1 i 308.976 758.09 243.779 -150.4 re W* n 0 841.89 595.275 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n /GS1 gs q 243.937 0 0 153.026 308.841 606.511 cm /Im11 Do Q EMC /P <>BDC Q q 1 i 0 841.89 595.275 -841.89 re 297.6 421.08 m W n 0 842.04 595.2 -841.92 re W n /GS2 gs BT /F2 1 Tf 8 0 0 8 410.5536 399.3421 Tm 0 0 0 1 k 0 Tc 0 Tw (Log \(concentration\))Tj ET EMC /Artifact <>BDC .78 .02 .98 .09 K 0 J 0 j .5 w 10 M [] 0 d /GS1 gs q 1 0 0 1 338.671 420.39 cm 0 0 m 0 137.422 l S Q EMC /P <>BDC /GS2 gs BT 0 8 -8 0 314.5209 458.6646 Tm (Cell viability %)Tj ET EMC /P <>BDC BT 7 0 0 7 321.4961 553.674 Tm (150)Tj ET EMC /P <>BDC BT 7 0 0 7 333.5502 553.674 Tm ( )Tj ET EMC /P <>BDC BT 7 0 0 7 321.4961 509.174 Tm (100)Tj ET EMC /P <>BDC BT 7 0 0 7 333.5502 509.174 Tm ( )Tj ET EMC /P <>BDC BT 7 0 0 7 325.5141 464.674 Tm (50)Tj ET EMC /P <>BDC BT 7 0 0 7 333.5502 464.674 Tm ( )Tj ET EMC /P <>BDC BT 7 0 0 7 329.5321 420.174 Tm (0)Tj ET EMC /P <>BDC /GS1 gs BT /F3 1 Tf 9 0 0 9 308.6488 571.3701 Tm 0 .68 .9 0 k [(DMSO cytoto)12(xicity assa)29.8(y for U373MG)]TJ ET EMC /Artifact <>BDC q 1 0 0 1 335.55 557.89 cm 0 0 m 217.205 0 l S Q EMC /Artifact <>BDC q 1 0 0 1 552.671 420.89 cm 0 0 m 0 136.922 l S Q EMC /Artifact <>BDC q 1 0 0 1 335.55 420.89 cm 0 0 m 217.205 0 l S Q EMC /P <>BDC /GS2 gs BT /F2 1 Tf 7 0 0 7 331.0501 410.0761 Tm 0 0 0 1 k (-1.0)Tj ET EMC /P <>BDC BT 7 0 0 7 344.7271 410.0761 Tm ( )Tj ET EMC /P <>BDC BT 7 0 0 7 374.8621 410.0761 Tm (-0.5)Tj ET EMC /P <>BDC BT 7 0 0 7 387.4926 410.0761 Tm ( )Tj ET EMC /P <>BDC BT 7 0 0 7 417.6276 410.0761 Tm (0.0)Tj ET EMC /P <>BDC BT 7 0 0 7 429.1208 410.0761 Tm ( )Tj ET EMC /P <>BDC BT 7 0 0 7 459.256 410.0761 Tm (0.5)Tj ET EMC /P <>BDC BT 7 0 0 7 470.2233 410.0761 Tm ( )Tj ET EMC /P <>BDC BT 7 0 0 7 500.3583 410.0761 Tm (1.0)Tj ET EMC /P <>BDC BT 7 0 0 7 511.3255 410.0761 Tm ( )Tj ET EMC /P <>BDC BT 7 0 0 7 541.4605 410.0761 Tm (1.5)Tj ET EMC /Artifact <>BDC /GS1 gs q 1 0 0 1 380.324 420.89 cm 0 0 m 0 -2.838 l S Q EMC /Artifact <>BDC q 1 0 0 1 421.924 420.89 cm 0 0 m 0 -2.8381 l S Q EMC /Artifact <>BDC q 1 0 0 1 463.524 420.89 cm 0 0 m 0 -2.838 l S Q EMC /Artifact <>BDC q 1 0 0 1 505.124 420.89 cm 0 0 m 0 -2.838 l S Q EMC /Artifact <>BDC q 1 0 0 1 546.724 420.89 cm 0 0 m 0 -2.838 l S Q EMC /Artifact <>BDC q 1 0 0 1 338.724 420.89 cm 0 0 m 0 -2.838 l S Q EMC /Artifact <>BDC q 1 0 0 1 335.55 467.046 cm 0 0 m 2.8379 0 l S Q EMC /Artifact <>BDC q 1 0 0 1 335.55 511.046 cm 0 0 m 2.8379 0 l S Q EMC /Artifact <>BDC 0 .68 .9 0 k q 1 0 0 1 357.434 511.08 cm 0 0 m 0 -1.5646 -1.2698 -2.8345 -2.8345 -2.8345 c -4.3991 -2.8345 -5.6689 -1.5646 -5.6689 0 c -5.6689 1.5646 -4.3991 2.8345 -2.8345 2.8345 c -1.2698 2.8345 0 1.5646 0 0 c h f* Q EMC /Artifact <>BDC q 1 0 0 1 434.354 495.12 cm 0 0 m 0 -1.5647 -1.2698 -2.8345 -2.8345 -2.8345 c -4.3991 -2.8345 -5.6689 -1.5647 -5.6689 0 c -5.6689 1.5646 -4.3991 2.8345 -2.8345 2.8345 c -1.2698 2.8345 0 1.5646 0 0 c f* Q EMC /Artifact <>BDC q 1 0 0 1 459.434 486 cm 0 0 m 0 -1.5647 -1.2698 -2.8345 -2.8345 -2.8345 c -4.3991 -2.8345 -5.6689 -1.5647 -5.6689 0 c -5.6689 1.5646 -4.3991 2.8345 -2.8345 2.8345 c -1.2698 2.8345 0 1.5646 0 0 c f* Q EMC /Artifact <>BDC q 1 0 0 1 487.394 469.08 cm 0 0 m 0 -1.5646 -1.2698 -2.8345 -2.8345 -2.8345 c -4.3991 -2.8345 -5.6689 -1.5646 -5.6689 0 c -5.6689 1.5646 -4.3991 2.8345 -2.8345 2.8345 c -1.2698 2.8345 0 1.5646 0 0 c f* Q EMC /Artifact <>BDC q 1 0 0 1 537.435 442.08 cm 0 0 m 0 -1.5646 -1.2698 -2.8345 -2.8345 -2.8345 c -4.3991 -2.8345 -5.6689 -1.5646 -5.6689 0 c -5.6689 1.5646 -4.3991 2.8345 -2.8345 2.8345 c -1.2698 2.8345 0 1.5646 0 0 c f* Q EMC /Artifact <>BDC .88 .45 .04 0 k q 1 0 0 1 534.921 427.78 cm 0 0 m 2.8339 -5.669 l -2.835 -5.67 l 0 0 l f* Q EMC /Artifact <>BDC q 1 0 0 1 483.921 439.78 cm 0 0 m 2.834 -5.669 l -2.835 -5.67 l 0 0 l f* Q EMC /Artifact <>BDC q 1 0 0 1 457.921 461.78 cm 0 0 m 2.834 -5.669 l -2.8349 -5.67 l 0 0 l f* Q EMC /Artifact <>BDC q 1 0 0 1 430.921 489.78 cm 0 0 m 2.834 -5.669 l -2.8349 -5.67 l 0 0 l f* Q EMC /Artifact <>BDC q 1 0 0 1 405.921 502.78 cm 0 0 m 2.834 -5.669 l -2.835 -5.67 l 0 0 l f* Q EMC /Artifact <>BDC 0 .68 .9 0 k q 1 0 0 1 409.394 505.08 cm 0 0 m 0 -1.5646 -1.2698 -2.8345 -2.8345 -2.8345 c -4.3991 -2.8345 -5.6689 -1.5646 -5.6689 0 c -5.6689 1.5646 -4.3991 2.8345 -2.8345 2.8345 c -1.2698 2.8345 0 1.5646 0 0 c f* Q EMC /Artifact <>BDC .88 .45 .04 0 k q 1 0 0 1 380.921 508.78 cm 0 0 m 2.834 -5.669 l -2.835 -5.67 l 0 0 l f* Q EMC /Artifact <>BDC 0 .68 .9 0 k q 1 0 0 1 383.354 510 cm 0 0 m 0 -1.5647 -1.2698 -2.8345 -2.8345 -2.8345 c -4.3991 -2.8345 -5.6689 -1.5647 -5.6689 0 c -5.6689 1.5646 -4.3991 2.8345 -2.8345 2.8345 c -1.2698 2.8345 0 1.5646 0 0 c f* Q EMC /Artifact <>BDC 0 .68 .9 0 K 1 w q 1 0 0 1 354.33 511.121 cm 0 0 m 39.625 2.769 77.779 -16 v S Q EMC /Artifact <>BDC q 1 0 0 1 431.493 494.428 cm 0 0 m 21.231 -2.461 53.846 -25.846 v S Q EMC /Artifact <>BDC q 1 0 0 1 484.109 468.89 cm 0 0 m 36.923 -24.615 50.461 -27.077 v S Q EMC /Artifact <>BDC .88 .45 .04 0 K q 1 0 0 1 354.57 509.15 cm 0 0 m 49.846 -2.106 76.923 -24.26 v S Q EMC /Artifact <>BDC q 1 0 0 1 431.186 485.505 cm 0 0 m 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