Pore size, shape, wall morphology, porosity, and interconnectivity are important characteristics of the scaffolds. Lithography is a manufacturing technique that allows the production of tridimensional scaffolds with a controllable and reproducible inner architecture. The aim of this study was to use lithography to create a poly-L-co-D,L lactide (PLDLA) scaffold with symmetrical pore size and distribution, and to evaluate its biocompatibility with osteoblasts in vitro. Lithographic moulds were used to produce porous PLDLA membranes by a casting procedure. Osteoblasts were removed from calvarial bones and seeded onto porous and smooth PLDLA membranes after which cell viability and adhesion assays, cytochemical analysis and scanning electron microscopy were used to characterize the cells. Cell viability and adhesion assays, cytochemical analysis, and scanning electron microscopy were carried out. Cell viability was similar on porous and smooth PLDLA membranes but higher than on a polystyrene substrate (positive control). Although osteoblasts adhered to the surface of all the materials tested, cell adhesion to lithographed PLDLA was greater than to smooth PLDLA membranes. In conclusion, osteoblasts interacted well with PLDLA membranes, as shown by the viability and adhesion assays and by the enhanced collagen production.171715Beresford, J.N., Graves, S.E., Smoothy, C.A., Formation of mineralized nodules by bone derived cells in vitro: A model of bone formation? (1993) American Journal of Medical Genetics, 45 (2), pp. 163-178. , http://dx.doi.org/10.1002/ajmg.1320450205, PMid:8456798Elgendy, H.M., Norman, M.E., Keaton, A.R., Laurencin, C.T., Osteoblast-like cell (MC3T3-E1) proliferation on bioerodible polymers: An approach towards the development of a bone-bioerodible polymer composite material (1993) Biomaterials, 14 (4), pp. 263-269. , http://dx.doi.org/10.1016/0142-9612(93)90116-JTang, Z.G., Hunt, J.A., The effect of PLGA doping of polycaprolactone films on the control of osteoblast adhesion and proliferation in vitro (2006) Biomaterials., 27 (25), pp. 4409-4418. , http://dx.doi.org/10.1016/j.biomaterials.2006.04.009, PMid:16677705Service, R.F., Tissue engineers build new bone (2000) Science., 289 (5484), pp. 1498-1500. , http://dx.doi.org/10.1126/science.289.5484.1498, PMid:10991738Wang, H., Li, Y., Zuo, Y., Li, J., Ma, S., Cheng, L., Biocompatibility and osteogenesis of biomimetic nano-hydroxyapatite/ polyamide composite scaffolds for bone tissue engineering (2007) Biomaterials., 28 (22), pp. 3338-3348. , http://dx.doi.org/10.1016/j.biomaterials.2007.04.014, PMid:17481726Wan, Y., Wang, Y., Liu, Z., Qu, X., Han, B., Bei, J., Adhesion and proliferation of OCT-1 osteoblast-like cells on micro-and nano-scale topography structured poly(L-lactide) (2005) Biomaterials., 26 (21), pp. 4453-4459. , http://dx.doi.org/10.1016/j.biomaterials.2004.11.016, PMid:15701374Sarazin, P., Roy, X., Favis, B.D., Controlled preparation and properties of porous poly(L-lactide) obtained from a co-continuous blend of two biodegradable polymers (2004) Biomaterials., 25 (28), pp. 5965-5978. , http://dx.doi.org/10.1016/j.biomaterials.2004.01.065, PMid:15183611Lee, S.J., Kang, H.W., Park, J.K., Rhie, J.W., Hahn, S.K., Cho, D.W., Application of microstereolithography in the development of three-dimensional cartilage regeneration scaffolds (2008) Biomedical Microdevices., 10 (2), pp. 233-241. , http://dx.doi.org/10.1007/s10544-007-9129-4, PMid:17885804Mendes, A.C., Smith, K.H., Tejeda-Montes, E., Engel, E., Reis, R.L., Azevedo, H.S., Co-assembled and microfabricated bioactive membranes (2013) Advanced Functional Materials, 23 (4), pp. 430-438. , http://dx.doi.org/10.1002/adfm.201201065Detsch, R., Guillon, O., Wondraczek, L., Boccaccini, A.R., Initial attachment of rMSC and MG-63 cells on Patterned Bioglass® substrates (2012) Advanced Engineering Materials, 14 (3), pp. B38-B44. , http://dx.doi.org/10.1002/adem.201180068Pelaez-Vargas, A., Gallego-Perez, D., Carvalho, A., Fernandes, M.H., Hansford, D.J., Monteiro, F.J., Effects of density of anisotropic microstamped silica thin films on guided bone tissue regeneration - In vitro study (2013) Journal of Biomedical Materials Research - Part B Applied Biomaterials, 101 (5), pp. 762-769. , http://dx.doi.org/10.1002/jbm.b.32879, PMid:23359600Wang, P.-Y., Li, W.T., Yu, J., Tsai, W.B., Modulation of osteogenic, adipogenic and myogenic differentiation of mesenchymal stem cells by submicron grooved topography (2012) Journal of Materials Science: Materials in Medicine, 23 (12), pp. 3015-3028. , http://dx.doi.org/10.1007/s10856-012-4748-6, PMid:22903603Prodanov, L., Lamers, E., Domanski, M., Luttge, R., Jansen, J.A., Walboomers, X.F., The effect of nanometric surface texture on bone contact to titanium implants in rabbit tibia (2013) Biomaterials., 34 (12), pp. 2920-2927. , http://dx.doi.org/10.1016/j.biomaterials.2013.01.027, PMid:23380354Cai, Y.-Z., Zhang, G.-R., Wang, L.-L., Jiang, Y.-Z., Ouyang, H.-W., Zou, X.-H., Novel biodegradable three-dimensional macroporous scaffold using aligned electrospun nanofibrous yarns for bone tissue engineering (2012) Journal of Biomedical Materials Research Part A., 100 (5), pp. 1187-1194. , http://dx.doi.org/10.1002/jbm.a.34063, PMid:22345081Ciapetti, G., Granchi, D., Devescovi, V., Baglio, S.R., Leonardi, E., Martini, D., Enhancing osteoconduction of PLLA-based nanocomposite scaffolds for bone regeneration using different biomimetic signals to MSCs (2012) International Journal of Molecular Sciences., 13 (2), pp. 2439-2458. , http://dx.doi.org/10.3390/ijms13022439, PMid:22408463 PMCid:PMC3292032Motta, A., Duek, E., Síntese, caracterização e degradação in vitro do poli(L-ácido latico) (2006) Polímeros., 16 (1), pp. 26-32. , http://dx.doi.org/10.1590/S0104-14282006000100008Agrawal, C.M., Ray, R.B., Biodegradable polymeric scaffolds for musculoskeletal tissue engineering (2001) Journal of Biomedical Materials Research, 55 (2), pp. 141-150. , http://dx.doi.org/10.1002/1097-4636(200105)552141::AID-JBM100030CO2-JPeters, M., Mooney, D., Synthetic extracellular matrices for cell transplantation (1997) Materials Science Forum, 250, pp. 43-52. , In: Liu D and Dixit V. Porous materials for tissue engineering,Einfield: Trans Tech PublicationMotta, A., Duek, E., Síntese e caracterização do copolímero poli(L-co-D,L-ácido lático) (2007) Polímeros., 17 (2), pp. 123-129. , http://dx.doi.org/10.1590/S0104-14282007000200011Yamamoto, N., Furuya, K., Hanada, K., Progressive development of the osteoblast phenotype during differentiation of osteoprogenitor cells derived from fetal rat calvaria: Model for in vitro bone formation (2002) Biological and Pharmaceutical Bulletin, 25 (4), pp. 509-515. , http://dx.doi.org/10.1248/bpb.25.509Moreira, P.L., An, Y.H., Santos, Jr.A.R., Genari, S.C., Vitro analysis of anionic collagen scaffolds for bone repair (2004) Journal of Biomedical Materials Research: Part B, Applied Biomaterials, 71 (2), pp. 229-237. , http://dx.doi.org/10.1002/jbm.b.30026, PMid:15386402Whiston, S.W., Whitson, M.A., Bowers Jr., D.E., Falk, M.C., Factors influencing synthesis and mineralization of bone matrix from fetal bovine bone cells grown in vitro (1992) Journal of Bone and Mineral Research, 7 (7), pp. 727-741Mosmann, T., Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays (1983) Journal of Immunological Methods, 65 (1-2), pp. 55-63. , http://dx.doi.org/10.1016/0022-1759(83)90303-4Lucchesi, C., Ferreira, B., Duek, E., Santos, A., Joazeiro, P., Increased response of Vero cells to PHBV matrices treated by plasma (2008) Journal of Materials Science: Materials in Medicine, 19 (2), pp. 635-643. , http://dx.doi.org/10.1007/s10856-007-0169-3, PMid:17619989Uzumaki, E.T., Lambert, C.S., Santos, Jr.A.R., Zavaglia, C.A.C., Surface properties and cell behaviour of diamond-like carbon coatings produced by plasma immersion (2006) Thin Solid Films., 515 (1), pp. 293-300. , http://dx.doi.org/10.1016/j.tsf.2005.12.081(1992) Biological Evaluation of Medical Devices, , International Organization for Standardization - ISO,Part 5: Tests for cytotoxicity: in vitro methods. ISOIgnatius, A.A., Claes, L.E., In vitro biocompatibility of bioresorbable polymers: Poly(L, DL-lactide) and poly(L-lactide-co-glycolide) (1996) Biomaterials, 17 (8), pp. 831-839. , http://dx.doi.org/10.1016/0142-9612(96)81421-9Marques, A.P., Cruz, H.R., Coutinho, O.P., Reis, R.L., Effect of starch-based biomaterials on the in vitro proliferation and viability of osteoblast-like cells (2005) Journal of Materials Science: Materials in Medicine, 16 (9), pp. 833-842. , http://dx.doi.org/10.1007/s10856-005-3580-7, PMid:16167112Coraça, D.C., Duek, E.A., Padovani, C.A., Camilli, J.A., Osteointegration of poly(L: -lactic acid)PLLA and poly(L: -lactic acid)PLLA/poly(ethylene oxide)PEO implants in rat tibiae (2008) Journal of Materials Science: Materials in Medicine, 19 (7), pp. 2699-2704. , http://dx.doi.org/10.1007/s10856-008-3397-2, PMid:18283533Coraça-Huber, D.C., Duek, E.A., Etchebehere, M., Magna, L.A., Amstalden, E.M., The use of vancomycin-loaded poly-l-lactic acid and poly-ethylene oxide microspheres for bone repair: An in vivo study (2012) Clinics, 67 (7), pp. 793-798. , http://dx.doi.org/10.6061/clinics/2012(07)15Ku, Y., Shim, I.K., Lee, J.Y., Park, Y.J., Rhee, S.H., Nam, S.H., Chitosan/poly(L-lactic acid) multilayered membrane for guided tissue regeneration (2009) Journal of Biomedical Materials Research: Part A., 90 (3), pp. 766-772. , http://dx.doi.org/10.1002/jbm.a.31846, PMid:18615563Pierucci, A., De Duek, E.A., De Oliveira, A.L., Peripheral nerve regeneration through biodegradable conduits prepared using solvent evaporation (2008) Tissue Engineering: Part A., 14 (5), pp. 595-606. , http://dx.doi.org/10.1089/tea.2007.0271, PMid:18399734Gong, Y., Ma, Z., Zhou, Q., Li, J., Gao, C., Shen, J., Poly(lactic acid) scaffold fabricated by gelatin particle leaching has good biocompatibility for chondrogenesis (2008) Journal of Biomaterials Science: Polymer Edition, 19 (2), pp. 207-221. , http://dx.doi.org/10.1163/156856208783432453, PMid:18237493Barauna, G., Coraça-Huber, D.C., Duek, E.A.R., Vitro degradation of Poly-L-co-D, L-lactic acid membranes (2013) Materials Research, 16 (1), pp. 221-226. , http://dx.doi.org/10.1590/S1516-14392012005000154Coimbra, M.E., Elias, C.N., Coelho, P.G., Vitro degradation of poly-L-D-lactic acid (PLDLA) pellets and powder used as synthetic alloplasts for bone grafting (2008) Journal of Materials Science: Materials in Medicine, 19 (10), pp. 3227-3234. , http://dx.doi.org/10.1007/s10856-008-3425-2, PMid:18454304Ikavalko, M., Skytta, E.T., Belt, E.A., One-year results of use of poly-L/D-lactic acid joint scaffolds and bone packing in revision metacarpophalangeal arthroplasty (2007) The Journal of Hand Surgery, European Volume, 32 (4), pp. 427-433. , http://dx.doi.org/10.1016/j.jhse.2007.03.006, PMid:17950198Stares, S.L., Boehs, L., Fredel, M.C., Aragones, A., Duek, E.A.R., Self-reinforced bioresorbable polymer P (L/DL) LA 70:30 for the manufacture of craniofacial implant (2012) Polímeros, 22 (4), pp. 378-383. , http://dx.doi.org/10.1590/S0104-14282012005000056Assaf, K., Duek, E.A.R., Oliveira, N.M., Efficacy of a combination of simvastatin and poly(DL-lactic-co-glycolic acid) in stimulating the regeneration of bone defects (2013) Materials Research., 16 (1), pp. 215-220. , http://dx.doi.org/10.1590/S1516-14392012005000159Pulliainen, O., Vasara, A.I., Hyttinen, M.M., Tiitu, V., Valonen, P., Kellomaki, M., Poly-L-D-lactic acid scaffold in the repair of porcine knee cartilage lesions (2007) Tissue Engineering., 13 (6), pp. 1347-1355. , http://dx.doi.org/10.1089/ten.2006.0347, PMid:17518746Esposito, A.R., Bonadio, A.C., Pereira, N.O., Cardoso, T.P., Barbo, M.L.P., Duek, E.A.R., The use of PLDLA/PCL-T scaffold to repair osteochondral defects in vivo (2013) Materials Research, 16 (1), pp. 105-115. , http://dx.doi.org/10.1590/S1516-14392012005000155Kangas, J., Pajala, A., Leppilahti, J., Ryhanen, J., Lansman, S., Tormala, P., Histomorphometric analysis of poly-L/D-lactide 96/4 sutures in the gastrocnemius tendon of rabbits (2006) The International Journal of Artificial Organs., 29 (9), pp. 893-899. , PMid:17033997Barauna, G.S., Pierucci, A., De Oliveira, A., Duarte, M.A.T., Duek, E., Estudo da degradação in vivo de poli(L-co-D,L-ácido lático) aplicado como prótese para regeneração nervosa periférica (2007) Revista Matéria., 12 (2), pp. 298-306Langer, R., Vacanti, J.P., Tissue engineering (1993) Science., 260 (5110), pp. 920-926. , http://dx.doi.org/10.1126/science.8493529, PMid:8493529Santos, Jr.A.R., Ferreira, B.M., Duek, E.A., Dolder, H., Wada, R.S., Wada, M.L., Differentiation pattern of Vero cells cultured on poly(L-lactic acid)/poly(hydroxybutyrate-co-hydroxyvalerate) blends (2004) Artificial Organs., 28 (4), pp. 381-389. , http://dx.doi.org/10.1111/j.1525-1594.2004.47199.x, PMid:15084200Wu, Y.C., Shaw, S.Y., Lin, H.R., Lee, T.M., Yang, C.Y., Bone tissue engineering evaluation based on rat calvaria stromal cells cultured on modified PLGA scaffolds (2006) Biomaterials., 27 (6), pp. 896-904. , http://dx.doi.org/10.1016/j.biomaterials.2005.07.002, PMid:16125224Gugala, Z., Gogolewski, S., Differentiation, growth and activity of rat bone marrow stromal cells on resorbable poly(L/DL-lactide) membranes (2004) Biomaterials., 25 (12), pp. 2299-2307. , http://dx.doi.org/10.1016/j.biomaterials.2003.09.009, PMid:14741595Bet, M.R., Goissis, G., Vargas, S., Selistre-De-Araujo, H.S., Cell adhesion and cytotoxicity studies over polyanionic collagen surfaces with variable negative charge and wettability (2003) Biomaterials., 24 (1), pp. 131-137. , http://dx.doi.org/10.1016/S0142-9612(02)00270-00273Ishaug-Riley, S.L., Crane-Kruger, G.M., Yaszemski, M.J., Mikos, A.G., Three-dimensional culture of rat calvarial osteoblasts in porous biodegradable polymers (1998) Biomaterials, 19 (15), pp. 1405-1412. , http://dx.doi.org/10.1016/S0142-9612(98)00021-0Barbanti, S.H., Santos, Jr.A.R., Zavaglia, C.A., Duek, E.A., Porous and dense poly(L-lactic acid) and poly(D,L-lactic acid-co-glycolic acid) scaffolds: In vitro degradation in culture medium and osteoblasts culture (2004) Journal of Materials Science: Materials in Medicine, 15 (12), pp. 1315-1321. , http://dx.doi.org/10.1007/s10856-004-5740-6, PMid:15747184Shi, X., Sitharaman, B., Pham, Q.P., Liang, F., Wu, K., Edward Billups, W., Fabrication of porous ultra-short single-walled carbon nanotube nanocomposite scaffolds for bone tissue engineering (2007) Biomaterials., 28 (28), pp. 4078-4090. , http://dx.doi.org/10.1016/j.biomaterials.2007.05.033, PMid:17576009 PMCid:PMC3163100Helen, W., Merry, C.L., Blaker, J.J., Gough, J.E., Three-dimensional culture of annulus fibrosus cells within PDLLA/Bioglass composite foam scaffolds: Assessment of cell attachment, proliferation and extracellular matrix production (2007) Biomaterials, 28 (11), pp. 2010-2020. , http://dx.doi.org/10.1016/j.biomaterials.2007.01.011, PMid:17250887Oliveira, A.L., Malafaya, P.B., Costa, S.A., Sousa, R.A., Reis, R.L., Micro-computed tomography (micro-CT) as a potential tool to assess the effect of dynamic coating routes on the formation of biomimetic apatite layers on 3D-plotted biodegradable polymeric scaffolds (2007) Journal of Materials Science: Materials in Medicine., 18 (2), pp. 211-223. , http://dx.doi.org/10.1007/s10856-006-0683-8, PMid:17323152Salgado, A.J., Figueiredo, J.E., Coutinho, O.P., Reis, R.L., Biological response to pre-mineralized starch based scaffolds for bone tissue engineering (2005) Journal of Materials Science: Materials in Medicine, 16 (13), pp. 267-275. , http://dx.doi.org/10.1007/s10856-005-6689-9, PMid:15744619Hall, B.K., Miyake, T., The membranous skeleton: The role of cell condensations in vertebrate skeletogenesis (1992) Anatomy and Embryology., 186 (2), pp. 107-124. , http://dx.doi.org/10.1007/BF00174948, PMid:1510240Lombello, C.B., Santos, Jr.A.R., Malmonge, S.M., Barbanti, S.H., Wada, M.L., Duek, E.A., Adhesion and morphology of fibroblastic cells cultured on different polymeric biomaterials (2002) Journal of Materials Science: Materials in Medicine, 13 (9), pp. 867-874. , http://dx.doi.org/10.1023/A:1016552413295, PMid:15348552Pelaez-Vargas, A., Gallego-Perez, D., Magallanes-Perdomo, M., Fernandes, M.H., Hansford, D.J., De Aza, A.H., Isotropic micropatterned silica coatings on zirconia induce guided cell growth for dental implants (2011) Dental Materials., 27 (6), pp. 581-589. , http://dx.doi.org/10.1016/j.dental.2011.02.014, PMid:21459429Pelaez-Vargas, A., Gallego-Perez, D., Ferrell, N., Fernandes, M.H., Hansford, D., Monteiro, F.J., Early spreading and propagation of human bone marrow stem cells on isotropic and anisotropic topographies of silica thin films produced via microstamping (2010) Microscopy and Microanalysis., 16 (6), pp. 670-676. , http://dx.doi.org/10.1017/S1431927610094158, PMid:20964878Carvalho, A., Pelaez-Vargas, A., Gallego-Perez, D., Grenho, L., Fernandes, M.H., De Aza, A.H., Micropatterned silica thin films with nanohydroxyapatite micro-aggregates for guided tissue regeneration (2012) Dental Materials., 28 (12), pp. 1250-1260. , http://dx.doi.org/10.1016/j.dental.2012.09.002, PMid:23026648Mata, A., Kim, E.J., Boehm, C.A., Fleischman, A.J., Muschler, G.F., Roy, S., A three-dimensional scaffold with precise micro-architecture and surface micro-textures (2009) Biomaterials, 30 (27), pp. 4610-4617. , http://dx.doi.org/10.1016/j.biomaterials.2009.05.0232014;17(1)15, PMid:19524292 PMCid:PMC3677580