Name: development of amelogenin-chitosan hydrogel for in vitro enamel regrowth with a dense interface
Uploaded: 2014-07-10T15:00:00
Description: Watch this Scientific Journal Video about Development of Amelogenin-chitosan Hydrogel for In Vitro Enamel Regrowth with a Dense Interface at JoVE.com

The authors would like to thank Prof. Steven Nutt and Mr. Yuzheng Zhang for assistant with the Focus Ion Beam, and the Center for Electron Microscopy and Microanalysis (CEMMA) at USC for electron microscopy. Research was supported by NIH-NIDCR grants; DE-13414 and DE-020099 to J.M.O.

인간의 어금니는 남부 캘리포니아 대학의 치과의 스트로우 학교에서 추출을위한 표준 절차에 따라 추출 및 기관 검토위원회의 승인을 처리했다. 1. 산 - 에칭 치아의 준비 조각 <ol><li> 복원 된 모든 충치가없는 사람의 세 번째 어금니를 선택합니다. </li><li> 어금니의 뿌리 부분을 제거하고, 길이 방향으로 두께 수냉식 저속 다이아몬드 톱을 사용하여 조각 2mm에 몰의 왕관을 ?...

<ol>
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에나멜의 미네랄 함량이 높은 그것을 인간의 몸에서 가장 단단한 광물 조직을 만드는 동안,이 바이오 세라믹은 종종 충치 또는 부식으로 발생하는 탈염 공정에 영향을 받기 쉽다. 유전자 돌연변이는 Amelogenesis 부전증 (20)라는 에나멜 기형의 상속 질병의 시리즈로 이어지는 얇은 또는 연약한 사기질을 일으킬 수 있습니다. 불소 또는 CPP-ACP를 포함하는 구강 건강 관리 제품은 몇 년 초...

Dental enamel is the hard mineralized surface of human teeth. It is composed of numerous needle-like apatite crystals, which are bundled in organized, parallel prisms to ensure the unique mechanical strength and biological protection that enamel provides1-2. Unlike other mineralized tissues, such as bone and dentin, mature enamel is acellular and cannot regenerate itself after substantial mineral loss1-2, which often occurs as dental caries or erosion. Commercially available products such as fluoride containing varnishes, tooth pastes and mouthwashes are effective in re-mineralizing enamel but none of them have the potential to promote the formation of organized apatite crystals. Clinically, the conventional treatment for enamel repair involves a filling procedure with artificial materials such as amalgam, ceramics, or composite resin 3. However, these materials usually do not interface well with the natural tissue surrounding the lesion, because their structures, components and properties are different from the natural enamel. As a result, secondary caries frequently develops overtime at the interface between the tooth and foreign materials. Therefore, in situ regrowth of enamel with a dense interface is an attractive target for the fields of materials science and stomatology. One particularly promising way to achieve this purpose is biomimetic synthesis of enamel-like material on the enamel surface. Recently, numerous in vitro attempts have been made to prepare enamel-like materials using biomimetic systems that contain nano-apatites or different organic additives 4-12. However, developing the optimal biomimetic strategy to promote remineralization crystals to achieve a perfect dense interface is still a challenge. 
During enamel mineralization, the oriented growth and elongation of apatite crystals is regulated by an amelogenin-rich matrix 1,13-14. To mimic the organic matrix in developing enamel, herein we describe a detailed protocol for fabricating an amelogenin-chitosan (CS-AMEL) hydrogel for in situ enamel regrowth on an acid-etched enamel surface used as a model for erosive lesions. As &#8220;the most versatile growth media&#8221; for crystals 15, hydrogel matrices have an advantage over a solution system since clinically they are easier to handle. Moreover, CS-AMEL is biocompatible, biodegradable, and has unique antimicrobial and adhesion properties that compare favorably with other biomimetic systems for dental applications 16. Importantly, the in situ mineralization of apatite crystals on the enamel surface provides a dense interface between the repaired layer and the natural enamel, which can potentially improve the durability of restorations and prevent the formation of new caries at the margin of the restoration.

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			<td height="21" style="height:21px;width:331px;">Name</td>
			<td style="width:209px;">Company</td>
			<td style="width:164px;">Catalog Number</td>
			<td style="width:516px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Material/ Reagent</td>
			<td style="width:209px;"></td>
			<td style="width:164px;"></td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:331px;">Human Third Molar&#160;</td>
			<td style="width:209px;">Ostrow School of Dentistry of the University of Southern California&#160;</td>
			<td style="width:164px;">N/A</td>
			<td style="width:516px;">The human molars were extracted following the standard procedures for extraction at the Ostrow School of Dentistry of the University of Southern California and handled with the approval of the Institutional Review Board.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:331px;">Recombinant Pocine Amelogenin</td>
			<td style="width:209px;">Expression and purification&#160; in lab</td>
			<td style="width:164px;">N/A</td>
			<td style="width:516px;">rP172, full-length&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Chitosan&#160;</td>
			<td style="width:209px;">Sigma-Aldrich</td>
			<td style="width:164px;">448877</td>
			<td style="width:516px;">medium molecular weight, 75-85% deacetylated</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Phosphoric Acid&#160;</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">AA033266</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Acetic Acid Glacial</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">A036289</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Sodium Hydroxide</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">BDH9292</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Calcium Chloride&#160;</td>
			<td style="width:209px;">Sigma-Aldrich</td>
			<td style="width:164px;">223506</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Dibasic Sodium Phosphate Anhydrous</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">BDH0316</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">BL21-CodonPlus (DE3)-RP&#160;</td>
			<td style="width:209px;">Agilent Technologies Inc.</td>
			<td style="width:164px;">230255</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Ammonium Sulfate</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">BDH8001</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Trifluoroacetic Acid</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">AAAL06374</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Acetonitrile</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">BDH1103</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Magnesium Chloride&#160;</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">BDH0244</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Potassium Dihydrogen Phosphate&#160;</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">BDH9268</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Potassium Chloride&#160;</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">BDH0258</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Ammonium Chloride&#160;</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">AAAA15000</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:331px;">HEPES (4-(2-Hydroxyethyl)piperazine-1-ethane-sulfonic acid)</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">AAA14777</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Sodium Fluoride&#160;</td>
			<td style="width:209px;">VWR</td>
			<td style="width:164px;">AA11561</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Tris-Buffered Saline</td>
			<td style="width:209px;">Bio-Rad</td>
			<td style="width:164px;">170-6435</td>
			<td style="width:516px;">10&#215; TBS</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Bovine Serum Albumin</td>
			<td style="width:209px;">EMD Millipore&#160;</td>
			<td style="width:164px;">12659</td>
			<td style="width:516px;">CalBioChem, Albumin, Bovine Serum, Fraction V, Low Heavy Metals&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Triton X-100</td>
			<td style="width:209px;">EMD Millipore&#160;</td>
			<td style="width:164px;">TX1568-1</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Chicken Anti-Amelogenin</td>
			<td style="width:209px;">N/A</td>
			<td style="width:164px;">N/A</td>
			<td style="width:516px;">A gift from Prof. Malcolm Snead, University of Southern California</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Bovine Anti-Chicken IgY-FITC</td>
			<td style="width:209px;">Santa Cruz Biotechnology</td>
			<td style="width:164px;">Sc-2700</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="32">
			<td height="32" style="height:32px;width:331px;">Equipments</td>
			<td style="width:209px;"></td>
			<td style="width:164px;"></td>
			<td></td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;width:331px;">High Performance Liquid Chromatography System</td>
			<td style="width:209px;">Agilent Technologies Inc.</td>
			<td style="width:164px;">Varian Prostar 210</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">C4 column</td>
			<td style="width:209px;">Phenomenx&#160;</td>
			<td style="width:164px;">Jupiter 5&#956; 300A</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Scanning Electron Microscopy&#160;</td>
			<td style="width:209px;">JEOL&#160;</td>
			<td style="width:164px;">JSM-7001</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">FIB-SEM&#160;</td>
			<td style="width:209px;">JEOL&#160;</td>
			<td style="width:164px;">JIB-4500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Transmission Electron Microscopy&#160;</td>
			<td style="width:209px;">JEOL</td>
			<td style="width:164px;">JEM-2100F</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Digital Low Speed Diamond Saw</td>
			<td style="width:209px;">MTI Corporation</td>
			<td style="width:164px;">SYJ-150</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Fluorescence Microscopy</td>
			<td style="width:209px;">Leica</td>
			<td style="width:164px;">DMI3000 B</td>
			<td style="width:516px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Ultrasonic Cleaner&#160;</td>
			<td style="width:209px;">Branson&#160;</td>
			<td style="width:164px;">2510</td>
			<td style="width:516px;">42 kHz, 100 W</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:331px;">Nano-indenter&#160;</td>
			<td style="width:209px;">Agilent Technologies Inc.</td>
			<td style="width:164px;">MTS XP</td>
			<td style="width:516px;"></td>
		</tr>
	</tbody>
</table>

development of amelogenin-chitosan hydrogel for in vitro enamel regrowth with a dense interface

Biomimetic enamel reconstruction is a significant topic in material science and dentistry as a novel approach for the treatment of dental caries or erosion. Amelogenin has been proven to be a critical protein for controlling the organized growth of apatite crystals. In this paper, we present a detailed protocol for superficial enamel reconstruction by using a novel amelogenin-chitosan hydrogel. Compared to other conventional treatments, such as topical fluoride and mouthwash, this method not only has the potential to prevent the development of dental caries but also promotes significant and durable enamel restoration. The organized enamel-like microstructure regulated by amelogenin assemblies can significantly improve the mechanical properties of etched enamel, while the dense enamel-restoration interface formed by an in situ regrowth of apatite crystals can improve the effectiveness and durability of restorations. Furthermore, chitosan hydrogel is easy to use and can suppress bacterial infection, which is the major risk factor for the occurrence of dental caries. Therefore, this biocompatible and biodegradable amelogenin-chitosan hydrogel shows promise as a biomaterial for the prevention, restoration, and treatment of defective enamel.

여기에 설명 된 프로토콜의 효과는 전자 현미경 (SEM), 선택 영역 전자 회절 (SAED) 및 X 선 회절 (XRD) 분석을 스캔에 의해 입증된다. 칠일위한 유전자 성별 감식-키토산 (CS-AMEL) 하이드로 겔에 의한 수리 후, 15 ㎛의 두께를 에나멜 조의 층이 에칭 된 에나멜 표면에 형성 하였다. 새로 성장 된 층 (도 3a의 화살표) 표면에 수직 인 C 축을 따라 우선적으로 성장하고, 50 ~ nm의 직경이 결정의 고도?...

Watch this Scientific Journal Video about Development of Amelogenin-chitosan Hydrogel for In Vitro Enamel Regrowth with a Dense Interface at JoVE.com

Development of Amelogenin-chitosan Hydrogel for In Vitro Enamel Regrowth with a Dense Interface

에 대한 성별 감정 - 키토산 하이드로 겔의 개발<em&gt; 체외</em고밀도 인터페이스&gt; 에나멜 재성장

이 문서에서는, 우리는 표면의 에나멜 재건을위한 유전자 성별 감식 - 키토산 하이드로 겔을 제조하는 프로토콜을 설명합니다. 하이드로 겔의 인회석 결정의 현장 성장 조직은 수복물의 효율성과 내구성을 향상하는 조밀 한 에나멜 복원 인터페이스를 형성했다.

Biomimetic enamel reconstruction is a significant topic in material science and dentistry as a novel approach for the treatment of dental caries or ...

Research

JoVE Journal

Bioengineering

Development of Amelogenin-chitosan Hydrogel for In Vitro Enamel Regrowth with a Dense Interface

Encapsulation of Cardiomyocytes in a Fibrin Hydrogel for Cardiac Tissue Engineering

심장 조직 공학을위한 섬유소 히드로겔에 Cardiomyocytes의 캡슐

Culturing cells in a three dimensional hydrogel environment is an important technique for developing constructs for tissue engineering as well as studying ...

연구

생체공학

Cultivation of Human Neural Progenitor Cells in a 3-dimensional Self-assembling Peptide Hydrogel

3 차원 자기 조립 펩티드 히드로겔에서 인간 신경 전구 세포의 배양

The  influence of 3-dimensional (3D) scaffolds on growth, proliferation and finally  neuronal differentiation is of great interest in order to find new ...

Bridging the Bio-Electronic Interface with Biofabrication

Biofabrication와 바이오 전자 인터페이스 브리지

Advancements in lab-on-a-chip technology promise to revolutionize both research and medicine through lower costs, better sensitivity, portability, and ...

Synthesis of an Intein-mediated Artificial Protein Hydrogel

테인 매개 인공 단백질 히드로 겔의 합성

We present the synthesis of a highly stable protein hydrogel mediated by a split-intein-catalyzed protein trans-splicing reaction. The building blocks of ...

Hydrogel Nanoparticle Harvesting of Plasma or Urine for Detecting Low Abundance Proteins

낮은 풍부 단백질을 검출하기위한 플라즈마 또는 소변의 하이드로 겔 나노 입자 수확

Novel biomarker discovery plays a crucial role in providing more sensitive and specific disease detection. Unfortunately many low-abundance biomarkers ...

Eye Irritation Test (EIT) for Hazard Identification of Eye Irritating Chemicals using Reconstructed Human Cornea-like Epithelial (RhCE) Tissue Model

Eye Irritation Test EIT for Hazard Identification of Eye Irritating Chemicals using Reconstructed Human Cornea-like Epithelial RhCE Tissue Model

복원 된 인간의 각막 모양의 상피 (RHCE) 조직 모델을 사용하여 눈에 자극성 화학 물질의 위험 식별을위한 눈 자극 시험 (EIT)

To comply with the Seventh Amendment to the EU Cosmetics Directive and EU REACH legislation, validated non-animal alternative methods for reliable and ...

In Vitro Model of Physiological and Pathological Blood Flow with Application to Investigations of Vascular Cell Remodeling

In Vitro Model of Physiological and Pathological Blood Flow with Application to Investigations of Vascular Cell Remodeling

혈관 세포 리모델링의 조사에 생리의 체외 모델 및 응용 프로그램과 함께 병리학 혈액 흐름

Vascular disease is a common cause of death within the United States. Herein, we present a method to examine the contribution of flow dynamics towards ...

Development of an In Vitro Ocular Platform to Test Contact Lenses

Development of an In Vitro Ocular Platform to Test Contact Lenses

개발<em&gt; 체외</em&gt; 안구 플랫폼은 콘택트 렌즈를 테스트하는

Currently, in vitro evaluations of contact lenses (CLs) for drug delivery are typically performed in large volume vials,1-6 which fail to mimic ...

Improved 3D Hydrogel Cultures of Primary Glial Cells for In Vitro Modelling of Neuroinflammation

Improved 3D Hydrogel Cultures of Primary Glial Cells for In Vitro Modelling of Neuroinflammation

생체 외에서 Neuroinflammation의 모델링에 대 한 기본 Glial 세포의 3 차원 하이드로 겔 문화 개선

In the central nervous system, numerous acute injuries and neurodegenerative disorders, as well as implanted devices or biomaterials engineered to enhance ...

Bioprintable Alginate/Gelatin Hydrogel 3D In Vitro Model Systems Induce Cell Spheroid Formation

Bioprintable Alginate/Gelatin Hydrogel 3D In Vitro Model Systems Induce Cell Spheroid Formation

셀 회전 타원 체 형성을 유발 하는 모델 시스템에 3D 생체 외에서 Bioprintable Alginate/젤라틴 하이드로 겔

The cellular, biochemical, and biophysical heterogeneity of the native tumor microenvironment is not recapitulated by growing immortalized cancer cell ...