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    A novel method for constructing an acellular 3D biomatrix from bovine spinal cord for neural tissue engineering applications
    (Wiley, 2019) Arslan, Yavuz Emre; Efe, Burcu; Arslan, Tugba Sezgin
    In this study, we aimed at generating 3-dimensional (3D) decellularized bovine spinal cord extracellular matrix-based scaffolds (3D-dCBS) for neural tissue engineering applications. Within this scope, bovine spinal cord tissue pieces were homogenized in 0.1 M NaOH and this viscous mixture was molded to attain 3D bioscaffolds. After resultant bioscaffolds were chemically crosslinked, the decellularization process was conducted with detergent, buffer, and enzyme solutions. Nuclear remnants in the native tissue and 3D-dCBS were determined with DNA content analysis and agarose gel electrophoresis. Afterward, 3D-dCBS were biochemically characterized in depth via glycosaminoglycan (GAG) content, hydroxyproline (HYP) assay, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Cellular survival of human adipose-derived mesenchymal stem cells (hAMSCs) on the 3D-dCBS for 3rd, 7th, and 10th days was assessed via MTT assay. Scaffold and cell/scaffold constructs were also evaluated with scanning electron microscopy and histochemical studies. DNA contents for native and 3D-dCBS were respectively found to be 520.76 +/- 18.11 and 28.80 +/- 0.20 ng/mg dry weight (n = 3), indicating a successful decellularization process. GAG content, HYP assay, and SDS-PAGE results proved that the extracellular matrix was substantially preserved during the decellularization process. In conclusion, it is believed that the novel decellularization method may allow fabricating 3D bioscaffolds with desired geometry from soft nervous system tissues.
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    Öğe
    A Novel Protocol to Generate Decellularized Bovine Spinal Cord Extracellular Matrix-based Scaffolds (3D-dCBS)
    (Bio-Protocol, 2019) Arslan, Yavuz E.; Efe, Burcu; Arslan, Tugba Sezgin
    Extracellular matrix (ECM)-based tissue engineering scaffolds have an essential role in promoting tissue regeneration. Nerve tissue engineering aims at facilitating the repair of permanent damage to the peripheral and central nervous systems, which are difficult to heal. For this purpose, a variety of biomaterials are being developed consisting of numerous synthetic and/or natural polymers to provide axonal reinnervation and to direct the growth of axons. Here, we present a novel protocol that enables to fabricate a 3-dimensional (3D) decellularized scaffold derived from the bovine spinal cord (BSC) ECM (3D-dCBS) for neural tissue engineering applications. In this protocol, a viscous ECMderived gel from BSC is prepared, molded, and chemically crosslinked with EDC/NHS (3D-CBS) before decellularization process. Decellularization of 3D-CBS is performed with 1% SDS to attain 3D-dCBS. As compared with other available methods, our protocol is a novel decellularization method that preserves a more significant part of the ECM. We believe that the mentioned protocol has the potential to produce a bioengineered scaffold from spinal cord tissue with desired geometry for regenerative medicine applications related to neural tissue engineering.
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    Assessment of the Cytotoxicity of Melia azedarach L. Extracts on Human Adipose-derived Mesenchymal Stem Cells
    (2018) Efe, Burcu; Galata, Yusuf Furkan; Arslan, Yavuz Emre
    In this study, aqueous extracts of Melia azedarach L. green fruit and leaves were obtained using two different extraction methods. The extraction yields of the green fruit and leaves were found as 24.11% and 37.98% for the infusion method; 17.76% and 27.00% for the rotating method, respectively. The total phenolic content, related to the infusion method, was ascertained for green fruit extract 173.67±10.84 mg Gallic Acid Equivalent (GAE)/g dry weight and leaf extract 312.33±9.81 mg GAE/g dry weight. In other respects, antioxidant activity related to the infusion method was determined for green fruit extract 172.51±13.23 mg Trolox/L and leaf extract 569.16±10.41 mg Trolox/L. Gas chromatography-mass spectrometry (GC-MS) analysis was performed to identify the chemical composition of the extracts. The cytotoxicity levels of the extracts were assessed on human adipose-derived mesenchymal stem cells (hAMSCs) using commercially available XTT assay. Consequently, it has been found that the green fruit extract has more cytotoxic activity than the leaf extract on hAMSCs.
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    Öğe
    Sinir doku mühendisliği için hücrelerinden arındırılmış sığır omuriliğinin insan adipoz mezenkimal kök hücrelerle etkileşimlerinin incelenmesi
    (Çanakkale Onsekiz Mart Üniversitesi, 2018) Efe, Burcu; Arslan, Yavuz Emre
    Bu çalışmada, Balıkesir'de sağlıklı hayvanların kesildiği yerel bir kesimhaneden sığır spinal kordu alınmıştır. Spinal kord kaynaklı iskeleler, sinir doku mühendisliği için en uygun yöntemle deselülerize (hücrelerinden arındırma) edilmiştir. Spinal kord temelli iskeleler Sodyum Dodesil Sülfat (SDS) Deoksiribonükleaz (DNaz) - Ribonükleaz (RNaz), etanol ve perasetik asit gibi kimyasal ajanlar kullanılarak deselülerize edilmiştir. Hücrelerinden arındırılmış iskeleler; DNA içerik analizi, glikozaminoglikan (GAG) testi, hidroksiprolin analizi, agaroz jel elektroforezi ve Sodyum Dodesil Sülfat Poliakrilamid Jel Elektroforezi (SDS-PAGE) analizleri ile karakterize edilmiştir. Satın alınan insan adipoz kaynaklı mezenkimal kök hücreler (iAMKH'ler) desellülerize iskele üzerine ekilmiştir. Desellülerize iskele üzerinde iAMKH'lerin hücre canlılığı 3., 7. ve 10. günler için 3-(4,5-dimetiltiazol-2yl) -2,5-difenil tetrazolyum bromür (MTT) testi kullanılarak değerlendirilmiştir. MTT testi sonuçlarına göre deselülerize iskelelerin hücre canlılığı üzerinde herhangi bir sitotoksik etkisi görülmemiştir. Ham doku, deselülerize iskele ve hücreli iskele hakkında niteliksel ve niceliksel bilgi elde etmek için, Taramalı Elektron Mikroskopisi (SEM), Enerji Dağılımlı X-Işını Spektroskopisi (EDX) ve histolojik analizler yapılmıştır. Analiz sonuçları, hücrelerin deselülerize iskele üzerinde kümelenmeye doğru gittiğini göstermiştir. Bunun yanı sıra, iskele üzerinde hücre yoğunluğu, hücre-hücre etkileşimleri ve hücre bağlantılarının olduğu SEM ve histolojik analizler ile kanıtlanmıştır. Sonuç olarak, hücrelerinden arındırılmış spinal kord temelli iskelelerin sinir doku mühendisliği uygulamalarında kullanım potansiyelinin olabileceği düşünülmektedir.