Effects of calcium phosphates incorporation on structural, thermal and drug-delivery properties of collagen:chitosan scaffolds

In this study, we evaluated how different procedures of calcium phosphate synthesis and its incorporation in collagen:chitosan scaffolds could affect their structural and thermal properties, aiming the obtention of homogeneous scaffolds which can act as drug delivery vehicles in bone tissue engineering. Therefore, three different scaffold preparation procedures were developed, changing the order of addition of the components: in CC-CNPM1 and CC-CNPM2, calcium phosphate synthesis was performed in situ in the chitosan gel (1%, w/w) followed by mixture with collagen (1%, w/w), with changes in the reagents used for calcium phosphate formation; in CC-CNPM3 procedure, calcium phosphate was synthesized ex situ and then incorporated into the collagen gel, in which chitosan in powder was mixed. In all procedures, 5% (in dry mass) of ciprofloxacin was incorporated. FTIR analysis confirmed the presence of calcium phosphate in all scaffolds. DSC curves showed that collagen denaturation temperature (Td) increased with calcium incorporation. SEM photomicrographs of scaffolds cross-section revealed porous scaffolds with calcium phosphate grains internally distributed in the polymeric matrix. XRD diffractograms indicated that the calcium phosphates obtained are hydroxyapatite. The pore size distribution was more homogeneous for CC-CNPM3, which also stood out for its smaller porosity and lower absorption in PBS. These results indicate that the in situ or ex situ phosphate incorporation in the scaffolds had a great influence on its structural properties, which also had consequences for ciprofloxacin release. CC-CNPM3 released a smaller amount of antibiotic (30%), but its release profile was better described by all the tested models

Avaliação clínica e estudo da vascularização corneal induzida pelas membranas de colágeno nativo e aniônico em córneas de coelhos

PURPOSE: To evaluate the corneal vascularization (CV) and the clinical aspects induced by interlamellar graft with native (NCM) and anionic (ACM) collagen membranes in rabbits corneas. METHODS: An interlamellar graft with a 0.25 x 0.25 cm NCM (group 1) or ACM (group 2) fragment was performed in the right eye (treated eye). In the left eye, an estromal tunnel was done (control eye). Sixteen rabbits were used, and they were subdivided into two experimental groups of eight animals each. The clinical evaluation was performed at the 1st, 3rd, 7th, 15th and 30th postoperative days. Corneal vascularization analysis was performed after 30 days by the Images Analizator System Leica Qwin-550®. RESULTS: After 7 days, corneal vascularization was observed at about 2.25 ± 0.71 mm (NCM) and at about 1.0 ± 1.69 mm (ACM), respectively, from the limbus in direction to the central cornea. After 15 days, CV increased in both groups (5.25 ± 1.03 mm - NCM; 2.0 ± 2.39 mm - ACM) and then progressively decreased until day 30 (2.25 ± 2.10 mm - NCM; 0.75 ± 2.12 mm - ACM). The statistical analysis indicated that the averages of the distances from the limb vessels to the grafts observed after 7 and 15 days had not differed statistically (p=0.17), and after 15 and 30 postoperative days had a tendency to differ statistically (p=0.09). The control eyes did not present any changes. CONCLUSION: The interlamellar graft with native and anionic collagen membranes induced corneal vascularization when applied to rabbit corneas, but anionic collagen membrane induced a smaller corneal vascularization when compared to native collagen membrane. Although further studies are required, the results found in this study demonstrated the usefulness of interlamellar graft with native and anionic collagen membranes in keratoplasties. These membranes consists in one more graft option for the surgical treatment of corneal repair in rabbits and others animals, when other forms of medical and surgical treatment are not effective.OBJETIVO: Avaliar os aspectos clínicos e vascularização corneal (VC) induzida pelo enxerto interlamelar das membranas de colágeno nativo (MCN) e de colágeno aniônico (MCA) em córneas de coelhos. MÉTODOS: Um fragmento com 0,25 x 0,25 cm de MCN (grupo 1) e MCA (grupo 2) foi realizado no olho direito (olho tratado) por enxertia interlamelar. No olho esquerdo (olho controle) foi realizado apenas um túnel estromal. No olho direito (olho controle) foi realizado apenas um túnel estromal. Dezesseis coelhos foram utilizados e foram divididos em dois grupos experimentais composto por oito animais cada. A avaliação clínica foi realizada aos 1, 3, 7, 15 e 30 dias de pós-operatório. A análise da vascularização corneal foi realizada após 30 dias pelo Sistema de analisador de imagens Leica Qwin-550®. RESULTADOS: Após 7 dias, a vascularização corneal do limbo em direção central da córnea observada foi de 2,25 ± 0,71 mm (MCN) e 1,0 ± 1,69 mm (ACM), respectivamente. Após 15 dias a vascularização corneal aumentou em ambos os grupos (5,25 ± 1,03 mm - MCN; 2,0 ± 2,39 mm - MCA), diminuindo até o 30º dia (2,25 ± 2,10 mm - MCN; 0,75 ± 2,12 mm - MCA). A análise estatística indicou que as médias das distâncias dos vasos do limbo em direção ao enxerto observadas após 7 e 15 dias não diferiram estatisticamente (p=0,17), e 15 e 30 dias de pós-operatório houve tendência a diferir estatisticamente (p=0,09). Os olhos controles não apresentaram nenhuma alteração. CONCLUSÃO: As membranas de colágeno nativo e de colágeno aniônico induzem a vascularização corneal quando aplicadas na córnea de coelhos por meio de enxertia interlamelar, mas membrana de colágeno ativo induz a pequena vascularização corneal quando comparada à membrana de colágeno aniônico. Embora estudos adicionais sejam necessários, os resultados encontrados no presente estudo demonstraram que as membranas de CN e CA possam ser úteis em ceratoplastias. Estas membranas consistem em mais uma opção de enxerto para o tratamento cirúrgico de reparo da córnea em coelhos e outros animais, quando não há resolução com outras formas de tratamento médico e cirúrgico.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

In vitro evaluation of hydroxyapatite, chitosan, and carbon nanotube composite biomaterial to support bone healing

O compósito à base de hidroxiapatita, quitosana e nanotubo de carbono foi desenvolvido com o intuito de auxiliar na consolidação óssea. Estudos anteriores sugerem que a combinação de substitutos ósseos e células-tronco mesenquimais (CTM) podem auxiliar a potencializar e promover a regeneração óssea. No presente estudo, o biomaterial foi desenvolvido e a viabilidade e a citotoxicidade de células Vero (ATCC CCL-81) e CTM obtidas de medula óssea provenientes de ovinos utilizando ensaios metil-tiazol-tetrazólio, MTT e iodeto de propídeo (PI) foram avaliadas em diferentes concentrações de extrato desse compósito. O compósito demonstrou ausência de citotoxicidade com comportamento semelhante ao grupo controle. Amostras com 50% e 10% de concentração de extrato do compósito mostraram resultados maiores comparados ao grupo controle (ensaio MTT). Esses resultados também sugerem que a presença do biomaterial pode ser utilizada em associação a CTM. Assim, esse estudo conclui que o compósito apresentado de hidroxiapatita, quitosana e nanotubo de cabono não foi considerado citotóxico e pode ser utilizado em teste in vivo.Hydroxyapatite, chitosan, and carbon nanotube composite biomaterial were developed to improve bone healing. Previous studies suggested that a combination of biomaterials and mesenchymal stem cells (MSCs) can potentially help promote bone regeneration. In the present study, we first developed hydroxyapatite, chitosan, and carbon nanotube composite biomaterial. Then, the effect of different concentrations of the extract on the viability of Vero cells (ATCC CCL-81) and MSCs obtained from sheep bone marrow using methylthiazol tetrazolium (MTT) and propidium iodide (PI) assays were evaluated. The biomaterial group demonstrated an absence of cytotoxicity, similar to the control group. Samples with 50% and 10% biomaterial extract concentrations showed higher cell viability compared to samples from the control group (MTT assay). These results suggest that the presence of this composite biomaterial can be used with MSCs. This study also concluded that hydroxyapatite, chitosan, and carbon nanotube composite biomaterial were not cytotoxic. Therefore, these could be used for performing in vivo tests

Properties of experimental resins based on synthesized propoxylated bis-GMA with different propionaldehyde ratios

The effect of different propionaldehyde ratios on the properties of bis-GMA-based comonomers and copolymers diluted with propoxylated bis-GMA (CH3bis-GMA) was evaluated. Five experimental comonomers were prepared combining bis-GMA with CH3bis-GMA and propionaldehyde at 0, 2, 8, 16, 24 mol%. Light polymerization was effected with the use of 0.2 wt. (%) each of camphorquinone and N,N-dimethyl-p-toluidine. Resin degrees of conversion (%DC) were evaluated by FT-IR spectrophotometry and Tg by Differential Scanning Calorimeter. Complex viscosity (η*), the effect of temperature on η*, and Microhardness (H) for dry and wet samples were also determined. Data were analyzed by Student's t-test, one-way ANOVA and Tukey-Kramer test (α = 0.05). The group with 24 mol% additive had a significant increase in %DC and H, and the lowest comonomer Tg and η*. No remarkable variation was noted in copolymers Tg s. All resins presented Newtonian behavior of viscosity, which linearly decreased with increased temperature. The η* decreased sigmoidally as the additive ratio increased

Preparation And Characterization Of Collagen-chitosan Blends [obtenção E Caracterização De Blendas Colágeno-quitosana]

Biodegradable polymer blends were obtained using collagen and chitosan. Membranes of collagen and chitosan in different proportions (3:1, 1:1 and 1:3) were prepared by mixing their acetate solutions (pH 3.5) at room temperature. The blends were characterized by differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), Fourier Transform infrared (FTIR) spectroscopy, specific viscosity, water absorption and stress-strain assays. The results showed that chitosan did not interfere in the structural arrangement of the collagen triple helix and the properties of the blends can be controlled by varing the proportion of the collagen and the chitosan.256 A943948Vidik, A., Vuust, J., (1980) Biology of Collagen, , Academic Press: New YorkChamberlain, L.J., Yannas, I.V., Arrizabalaga, A., Hsu, H.P., Norregard, T.V., Spector, M., (1998) Biomaterials, 19, p. 1393Yao, K., Peng, T., Yin, Y., Xu, M.X., (1995) Macromol. Chem. Phys., C35, p. 158Muzzarelli, R.A.A., Mattioli-Belmonte, M., Tietz, C., Biagini, R., Ferioli, G., Brunelli, M.A., Fini, M., Biagini, G., (1994) Biomaterials, 15, p. 1075Miyata, T., Taira, T., Noishiki, Y., (1992) Clin. Mater., 9, p. 139Biagini, G., Bertani, A., Muzzarelli, R., Damatei, A., Dibenedetto, G., Belligolli, A., Ricotil, G., (1991) Biomaterials, 12, p. 281Thomas, D.A., Sperling, L.H., (1978) Polymer Blends, p. 3. , Paul, D. R.Newman, S., eds.Academic Press: New YorkUtracki, L.A., (1990) Polymer Alloys and Blends: Thermodynamics and Rheology, pp. 1-27. , Hanser Publishers: New YorkThacharodi, D., Rao, P.K., (1995) Int. J. Pharm., 120, p. 115Izume, M., Tairat, T., Kimura, J., Miyata, T., 4th International Conference on Chitin and Chitosan, p. 653. , Elsevier: LondonTaravel, M.N., Domard, A., (1993) Biomaterials, 14, p. 930Goissis, G., Plepis, A.M.G., Rocha, J.L., Br PI 9.405.043-0 A, 1996Laemmli, U.K., (1970) Nature, 227, p. 680George, A., Veis, A., (1991) Biochemistry, 30, p. 2372Hirai, A., Odani, H., Nakajima, A., (1991) Polym. Bull., 26, p. 87Yomota, C., Miyazaki, T., Okada, S., (1993) Colloid. Polym. Sci., 271, p. 76Wang, W., Bo, S., Li, S., Qin, W., (1991) Int. J. Biol. Macromol., 13, p. 281Rinaudo, M., Milas, D.P.L., (1993) Int. J. Biol. Macromol., 15, p. 281Veis, A., (1982) Conn. Tiss. Res., 10, p. 11Ramachandran, G.N., (1967) Treatise on Collagen, p. 103. , Academic Press: LondonPayne, K.J., Veis, A., (1988) Biopolymers, 27, p. 1749Shigemasa, Y., Matsuura, H., Sashiwa, H., Saimoto, H., (1996) Int. J. Biol. Macromol., 18, p. 237Privalov, P.I., Tiktopulo, E.I., (1970) Biopolymers, 9, p. 127Cárdenas, T.G., Ernal, A.L., Tagle, D.L.H., (1992) Thermochim. Acta, 195, p. 33Nieto, J.M., Peniche, A.C.C., (1991) Thermochim. Acta, 176, p. 63Sakurai, K., Maegawa, T., Takahashi, T., (2000) Polymer, 41, p. 705