Skeletal myotube formation enhanced through fibrillated collagen nanofibers coated on a 3D-printed polycaprolactone surface

Chae, SooJung,Lee, JiUn,Kim, GeunHyung Elsevier 2019 Colloids and Surfaces B Vol.181 No.-

<P><B>Abstract</B></P> <P>This work focused on considering the cellular responses of the growth and differentiation of myoblasts, C2C12, on fibrillated collagen-coated poly(ε-caprolactone) (PCL) surfaces. Through a fibrillation processing window using NaCl and collagen weight fractions, collagen fibril coating density can be controlled. Three different collagen-fibril densities coated on PCL strut were used to investigate the effects of the collagen fibril on the myoblast activities. After physical and cellular analyses of the scaffolds, such as surface morphology, fibronectin absorption, wettability, and mechanical properties, the rate of cell growth and the proficiency of the myoblasts to develop skeletal myotubes were evaluated. Based on the results, although the coated collagen nanofibers were randomly distributed, the fibrillated collagen layer with the appropriate density on the PCL surface promoted a greater myotube formation than that of the control, which had no fibrillated collagen. In particular, relatively higher densities of collagen fibril showed significantly greater myotube formation than those of the control (not-fibrillated collagen-coated on the PCL surface) and lower density of collagen fibril.</P> <P><B>Highlights</B></P> <P> <UL> <LI> A fibrillated collagen-coated polycaprolactone surface was prepared. </LI> <LI> The proper fibrillation condition of the coated collagen was selected through a processing window. </LI> <LI> Faster myotube formation on fibrous collagen surface was achieved. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>

Fibrillar assembly and stability of collagen coating on titanium for improved osteoblast responses

Kim, Hae-Won,Li, Long-Hao,Lee, Eun-Jung,Lee, Su-Hee,Kim, Hyoun-Ee Wiley Subscription Services, Inc., A Wiley Company 2005 Journal of biomedical materials research. Part A Vol.a75 No.3

<P>Collagen, as a major constituent of human connective tissues, has been regarded as one of the most important biomaterials. As a coating moiety on Ti hard-tissue implants, the collagen has recently attracted a great deal of attention. This article reports the effects of fibrillar assembly and crosslinking of collagen on its chemical stability and the subsequent osteoblastic responses. The fibrillar self-assembly of collagen was carried out by incubating acid-dissolved collagen in an ionic-buffered medium at 37°C. The degree of assembly was varied with the incubation time and monitored by the turbidity change. The differently assembled collagen was coated on the Ti and crosslinked with a carbodiimide derivative. The partially assembled collagen contained fibrils with varying diameters as well as nonfibrillar aggregates. On the other hand, the fully assembled collagen showed the complete formation of fibrils with uniform diameters of ∼100–200 nm with periodic stain patterns within the fibrils, which are typical of native collagen fibers. Through this fibrillar assembly, the collagen coating had significantly improved chemical stability in both the saline and collagenase media. The subsequent crosslinking step also improved the stability of the collagen coating, particularly in the unassembled collagen. The fibrillar assembly and the crosslinking of collagen significantly influenced the osteoblastic cell responses. Without the assembly, the collagen layer on Ti adversely affected the cell attachment and proliferation. However, those cellular responses were improved significantly when the collagen was assembled to fibrils and the assembly degree was increased. After crosslinking the collagen coating, these cellular responses were significantly enhanced in the case of the unassembled collagen but were not altered much in the assembled collagen. Based on these observations, it is suggested that the fibrillar assembly and the crosslinking of collagen require careful considerations in the collagen administration as a coating moiety. © 2005 Wiley Periodicals, Inc. J Biomed Mater Res, 2005</P>

Reconstituted Fibril from Heterogenic Collagens-A New Method to Regulate Properties of Collagen Gels

Jian Yang,Haibo Wang,Lang He,Benmei Wei,Chengzhi Xu,Yuling Xu,Juntao Zhang,Sheng Li 한국고분자학회 2019 Macromolecular Research Vol.27 No.11

Heterotypic collagen fibril has long been found in the tissues of organisms, which plays an important role in the formation and function of complex structures of organisms. Inspiring by the phenomenon, scholars tried to incubate collagens from different sources into novel collagen materials in vitro, and the forming of heterogenic reconstituted collagen fibrils (RF) was often demonstrated by differential scanning calorimetry (DSC) and fluorescence quenching analysis. In this work, we used two type-I collagens from different species (bovine tendon and grasscarp fish skin) to coassemble in vitro, and verified the formation of RF from a new rheological perspective. In addition, we also investigated the assembly behavior, surface hydrophilicity and hydrophobicity, microscopic morphology and cell proliferation ability of the RF. The results showed that the assembly rate, surface properties, fibril size, viscoelastic properties of RF can be delicately regulated by the method of heterogenic collagen reconstitution. This study provides new experimental evidence for the reconstitution of heterogenic collagens, and also offers a new means for the regulation of collagen gel performance, which would help to expand the application range of collagen gel materials.

가령에 따른 흰쥐 하악과두 섬유층의 미세구조 및 교원원섬유의 변화

변기정,Byeon, Ki-Jeong 대한악안면성형재건외과학회 1998 Maxillofacial Plastic Reconstructive Surgery Vol.20 No.4

흰쥐에 있어서 가령에 따라 하악과두에 가해지는 기계적 부하의 변화에 따라 하악과두 섬유층의 미세구조적 변화를 관찰, 분석한 결과를 요약하면 다음과 같다. 하악과두 섬유층의 세포는 생후 초기에는 미분화간엽세포 및 섬유모세포로 구성되어 있었다. 가령에 따라 생후 14일, 27일군 등에서는 생후 초기에 비해 교원원섬유를 활발하게 합성, 분비하는 것으로 생각되는 섬유모세포가 발달하였는데 이들 세포는 과립형 물질로 채워져 있는 내강이 대단히 확장되어 있으며, 리보솜이 치밀하게 존재하는 과립세포질세망과 골지장치가 발달되어 있었다. 또한 이들 세포내에는 세포소기관 사이에 세사가 점차로 발달하는 양상을 보였다. 생후 55일군 이후에서는, 가늘고 긴 세포돌기를 가지며, 과립세포질세망 및 골지장치가 발달되어 있어 교원섬유의 합성이 왕성한 것으로 추측되는 세포, 약간 둥글거나 다각형의 모양을 보이며, 세포질내에 사립체, 용해소체 및 골지장치가 발달하였으며, 손상된 교원섬유 혹은 변성된 세포의 흡수에 관여하는 것으로 추측되는 세포, 세포질의 상당부분이 세사로 채워져 있으며, 사립체, 용해소체 및 과립세포질내세망과 같은 세포질내 소기관은 잘 발달되어 있지 않아 가해진 기계적 자극에 대해 적응하였거나 활동이 정지된 세포 등 3종류의 세포가 관찰되었다. 세포간질의 교원원섬유는 생후 초기부터 가령에 따라 점차로 치밀하게 배열되는 양상을 보였다. 교원원섬유의 직경은 생후 1일, 7일군에서는 $38.48{\pm}3.81nm$, $38.06{\pm}3.86nm$로서 유사하였으나 생후 14일군에서는 $50.21{\pm}3.93nm$로 가장 굵은 양상을 보였으며(p<0.05), 그후 생후 27일, 55일군에서는 각기 $40.05{\pm}2.52nm$, $43.63{\pm}1.20nm$로서 점차로 가늘어져 생후 1년군에서는 $37.38{\pm}2.17nm$로서 가장 가늘어 졌다(p<0.05). 그 분포에 있어서는 생후 초기에는 직경이 30-60인 교원원섬유가 대다수를 차지하는 unimordal한 분포를 보였으나, 생후 27일 이후에는 직경이 작은 교원원섬유가 점차로 증가하는 양상을 보였다. 이상의 가령에 따른 결과로 미루어보아 하악과두 섬유층의 미세구조, 교원원섬유의 배열 및 직경의 분포에서의 변화는 가해지는 기계적 부하에 대한 적응한 결과라고 생각된다. The fibrous layer of mandibular condyle of the neonatal, 1-, 7-, 14-, 27-, 55-days and 1 year old rats were examined in the electron microscope with particular attention to the ultrastructure and diameter of collagen fibrils. In the 1-day rats, most of the cells of the fibrous layer were undifferentiated mesenchymal cells and fibroblasts with rough a little developed rough endoplasmic reticulum(RER) and golgi apparatus(GA). In 7-, 17 and 27-days old rats, most of the fibroblast showed well developed GA and RER with widely distended cisternae containing granular materials. In many of these cells contained intracytoplasmic filaments among the cytoplamic organelle. In 55-day and 1-year old rats, three types of cells were observed, ie, cells containing well developed cytoplasmic organelle presumed to be involved in the collagen fibril synthesis, cells showing well developed lysosomes, golgi apparatus, mitochondria and short cytoplasmic process presumed to be involved in the active resorption of the injured collagen fibrils or cellular debris, cells containing many intracytoplasmic filaments and a little organelle presumed to be cells of inactive state. The average diameters of collagen fibrils were similar in 1- and 7-day old rats as $38.48{\pm}3.81nm$, $38.06{\pm}3.86nm$. That was thickest in 14 days old rats as $50.21{\pm}3.93nm$ among experimental groups. They were gradually thinner in 27-, 55-day rats as $40.05{\pm}2.52nm$, $43.63{\pm}1.20nm$ and thinnest in 1-year old rats as $37.38{\pm}2.17nm$. The distribution pattern of diameters of collagen fibrils were unimordal with peak of 30-60nm in rats from 1-day to 17-day old. With aging from 27-day to 1 year old rats, collagen fibril diameters showed wide distribution pattern and percentage of thin collagen fibrils increased. These results may show the functional adaptation of fibrous layer of mandibular condyle to the increased mechanical forces with aging.

Effect of Ionic Liquids on the Fibril-Formation and Gel Properties of Grass Carp (Ctenopharyngodon idellus) Skin Collagen

Zhongwei Zhai,Haibo Wang,Benmei Wei,Peiwen Yu,Chengzhi Xu,Lang He,Juntao Zhang,Yuling Xu 한국고분자학회 2018 Macromolecular Research Vol.26 No.7

Self-assembled environment of collagen is one of the important factors for improving and regulating the properties of collagen-based biomaterials. This study aimed to investigate the effect of ionic liquids (ILs) on the fibril-formation and gel properties of grass carp (Ctenopharyngodon idellus) skin collagen. Fibrillogenic kinetics analysis showed that the collagen self-assembly can be suppressed by the introduction of ILs, and the inhibitory effect is influenced by concentration and types of ILs. Scanning electron microscopy test indicated that the assembled collagen fibrils in the presence of ILs had bigger diameters than that in the conventional buffer. Differential scanning calorimetry analysis revealed that the thermal stability of collagen fibrils can be significantly increased when self-assembly is performed in the presence of ILs. Moreover, the introduction of ILs enhanced the mechanical strength of collagen gels. Finding from this work provides a new idea for improving the performance of fish-sourced collagen biomaterials.

The Packing Nature of Apatite Crystals in Calcified Collagen Fibrils of Fish Bone

Kim, Hyun Man The Official Publication of Korean Academy of Oral 1993 International Journal of Oral Biology Vol.17 No.1

The packing nature of bone apatite crystals in the calcified collagen fibrils was studied on the calcified collagen fibrils isolated from fish rib bone with the treatment of detergent and ultrasonication. Stereo-electron micrograms and pattern of selected area electron diffraction with tilting of sample were obtained under transmission electron microscope. This study showed pattern of electron diffraction which cannot be explained by previously proposed packing model of crystals in collagen fibrils. The arrangement of crystals is interpreted as followings: [0001] of flat crystals run along the longitudinal axis of the collagen fibril with a deviation from general direction: tilting up to 30 degree up and down and sperading up to 45 degree right and left against longitudinal axis of collagen fibrils. Crystals in group also tilt around the longitudinal axis of collagen fibrils along the longitudinal axis of collagen fibril. Most of crystals are in parallel each other, but a few crystals interpose between parallel crystals with a limited angle.

원자현미경을 이용한 마이토마이신 노출 시간에 따른 공막 표면 콜라젠의 변화 관찰

이희재,최삼진,정유진,정경복,진경현,박헌국,이승준,Hui-Jae Lee,MD,Samjin Choi,PhD,Youjin Cheong,MD,PhD Candidate,Gyeong Bok Jung,PhD,Kyung-Hyun Jin,MD,Hun-Kuk Park,MD,PhD,Seung Jun Lee,MD 대한안과학회 2011 대한안과학회지 Vol.52 No.6

Purpose: To investigate the effects of mitomycin C on the scleral collagen surfaces using atomic force microscopy (AFM). Methods: Two non-contact mode AFM machines were used to observe changes in the morphological characteristics of human scleral surfaces before and after one, three, and five minutes of 0.02% mitomycin C application. Based on AFM topography and deflection images of the collagen fibril, the morphological characteristics of scleral fibrils including the fibril diameter and D-period were measured using the line profile. Results: The sclera collagen fibril treated with 0.02% mitomycin C for one minute did not show any significant increases in mean fibril diameter (155.04 ± 17.46 nm) or mean D-periodicity (70.02 ± 3.33 nm), compared to those of the control group. However, the scleral collagen fibrils treated with 0.02% mitomycin C for three and five minutes showed significant increases in mean fibril diameter (182.33 ± 16.33 nm, 199.20 ± 12.40 nm, respectively) and mean D-periodicity (70.27 ± 13.66 nm, 72.75 ± 19.32 nm, respectively), compared to those of the control group. Conclusions: The present study examined the structural changes in the scleral collagen fibrils before and after mitomycin C application according to atomic force microscopy. The results indirectly suggest that three or more minutes of 0.02% mitomycin C application affects the morphology of scleral collagen. J Korean Ophthalmol Soc 2011;52(6):671-678

Lateral Force Microscopy를 이용한 Collagen Fibrils의 운동 특성에 관한 연구

정구현(Koo-Hyun Chung) 한국트라이볼로지학회 2012 한국트라이볼로지학회 학술대회 Vol.2012 No.4

Optical Characteristics of Corneal Nanostructure According to the Angle of Collagen-fiber-layer Arrangement

이명희,김영철 한국광학회 2022 Current Optics and Photonics Vol.6 No.2

Collagen fibers tens of nanometers in size, which constituting most of the corneal volume of the hu-man eye, are layered in a uniform direction, and adjacent fiber layers are arranged at an angle of 90° toeach other. According to the results of this study, the transmittance at 45° of interlayer rotation angle is highest, and higher than that of the 90° body structure. The transmittance is examined, concerning the polarization state of the incident light; circularly polarized light case shows higher transmittance than linearly polarized. Through this, a simulation to confirm the deformed structure of collagen fibers, which show higher transmittance than the anatomical structure of the cornea, is attempted.

Molecular architecture of collagen fibrils: A critical length scale for tough fibrils

Markus J. Buehler 한국물리학회 2008 Current Applied Physics Vol.8 No.3,4

Protein materials constitute Nature’s most intriguing material concepts, leading to multi-functional and stimuli responsive materials. Such materials often feature a characteristic hierarchical design, which is characterized by structural features starting at nanoscale. Here we review recent studies of deformation of collagen, Nature’s most abundant structural protein material forming the basis of bone, tendon and skin. We have discovered that a specific nanostructural design with molecular lengths of 200 nm leads to the strongest possible ultra-structure that is still capable of dissipating large amounts of energy before fracture occurs, maximizing the toughness of the material [M.J. Buehler, Proceedings of the National Academy of Sciences USA 103 (2006) 12285]. The analysis explains prevalent molecular length scales observed in tendon, bone and the eye’s cornea, and explains how molecular properties influence the deformation and fracture mechanics of tissues. Protein materials constitute Nature’s most intriguing material concepts, leading to multi-functional and stimuli responsive materials. Such materials often feature a characteristic hierarchical design, which is characterized by structural features starting at nanoscale. Here we review recent studies of deformation of collagen, Nature’s most abundant structural protein material forming the basis of bone, tendon and skin. We have discovered that a specific nanostructural design with molecular lengths of 200 nm leads to the strongest possible ultra-structure that is still capable of dissipating large amounts of energy before fracture occurs, maximizing the toughness of the material [M.J. Buehler, Proceedings of the National Academy of Sciences USA 103 (2006) 12285]. The analysis explains prevalent molecular length scales observed in tendon, bone and the eye’s cornea, and explains how molecular properties influence the deformation and fracture mechanics of tissues.