この研究は、助成金番号1 - DP2 - OD004309 - 01を介して、NIHのディレクターの新イノベーター賞プログラム、医学研究のためのNIHロードマップの一部が部分的にサポートされていました。 SBS - N。大学院研究フェローシップのためにNSFを感謝したいと思います。

<ol>
	<li>Seif-Naraghi, S. B., Salvatore, M. A., Magoffin-Schup, P. J., Hu, D. P., Christman, K. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Design+and+characterization+of+an+injectable+pericardial+matrix+gel:+A+potentially+autologous+scaffold+for+cardiac+tissue+engineering.">Design and characterization of an injectable pericardial matrix gel: A potentially autologous scaffold for cardiac tissue engineering.</a> Tissue Engineering. , (2009).</li><li>Freytes, D. O., Martin, J., Velankar, S. S., Lee, A. S., Badylak, S. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preparation+and+rheological+characterization+of+a+gel+form+of+the+porcine+urinary+bladder+matrix.">Preparation and rheological characterization of a gel form of the porcine urinary bladder matrix.</a> Biomaterials. 29, 1630-1630 (2008).</li><li>Gilbert, T. W., Sellaro, T. L., Badylak, S. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decellularization+of+tissues+and+organs.">Decellularization of tissues and organs.</a> Biomaterials. 27, 3675-3675 (2006).</li><li>Liao, J., Joyce, E. M., Sacks, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+decellularization+on+the+mechanical+and+structural+properties+of+the+porcine+aortic+valve+leaflet.">Effects of decellularization on the mechanical and structural properties of the porcine aortic valve leaflet.</a> Biomaterials. 29, 1065-1065 (2008).</li><li>Singelyn, J. M., DeQuach, J. A., Seif-Naraghi, S. B., Littlefield, R. B., Schup-Magoffin, P. J., Christman, K. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Naturally+derived+myocardial+matrix+as+an+injectable+scaffold+for+cardiac+tissue+engineering.">Naturally derived myocardial matrix as an injectable scaffold for cardiac tissue engineering.</a> Biomaterials. 30, 5409-5409 (2009).</li><li>Christman, K. L., Vardanian, A. J., Fang, Q., Sievers, R. E., Fok, H. H., Lee, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Injectable+fibrin+scaffold+improves+cell+transplant+survival,+reduces+infarct+expansion,+and+induces+neovasculature+formation+in+ischemic+myocardium.">Injectable fibrin scaffold improves cell transplant survival, reduces infarct expansion, and induces neovasculature formation in ischemic myocardium.</a> J Am Coll Cardiol. 44, 654-654 (2004).</li><li>Christman, K. L., Fok, H. H., Sievers, R. E., Fang, Q., Lee, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fibrin+glue+alone+and+skeletal+myoblasts+in+a+fibrin+scaffold+preserve+cardiac+function+after+myocardial+infarction.">Fibrin glue alone and skeletal myoblasts in a fibrin scaffold preserve cardiac function after myocardial infarction.</a> Tissue Eng. 10, 403-410 (2004).</li><li>Huang, N. F., Sievers, R. E., Park, J. S., Fang, Q., Li, S., Lee, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+rodent+model+of+myocardial+infarction+for+testing+the+efficacy+of+cells+and+polymers+for+myocardial+reconstruction.">A rodent model of myocardial infarction for testing the efficacy of cells and polymers for myocardial reconstruction.</a> Nat Protoc. (1), 1596-1609 (2006).</li><li>Ott, H. C., Matthiesen, T. S., Goh, S. K., Black, L. D., Kren, S. M., Netoff, T. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Perfusion-decellularized+matrix:+using+nature's+platform+to+engineer+a+bioartificial+heart.">Perfusion-decellularized matrix: using nature's platform to engineer a bioartificial heart.</a> Nat Med. 14, 213-221 (2008).</li><li>Badylak, S. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+extracellular+matrix+as+a+biologic+scaffold+material.">The extracellular matrix as a biologic scaffold material.</a> Biomaterials. 28, 3587-3593 (2007).</li></ol>

動物の使用：本研究ではすべての実験は、カリフォルニア州、サンディエゴおよび実験動物のケアの認定のためのアメリカ連合の大学の動物実験使用の委員会が公開されているガイドラインに準拠して行われた。

このメソッドは、心筋組織工学のための生物学的に誘導された、注射足場の世代を可能にする。これらのメソッドが最初に製造するため、心筋マトリックスゲルの in vivo試験で開発され、心膜マトリックスのゲルをここに提示されたが、このプロトコルは、組織が ​​適切に脱細胞することができます提供されて、どんな組織で使用するために適応させることができる。細胞化?...

<table><tbody><tr><td>&lt;strong&gt;Reagents:&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Pepsin</td><td>Sigma-Aldrich</td><td>p6887-1G</td><td>Lyophilized </td></tr><tr><td>Biotin</td><td>Thermo Fisher Scientific, Inc.</td><td>21217</td><td></td></tr><tr><td>Neutravidin-HRP</td><td>Thomas Scientific</td><td>21130</td><td></td></tr><tr><td>&lt;strong&gt;Equipment:&lt;/strong&gt; </td><td></td><td></td><td></td></tr><tr><td>Wiley Mini Mill</td><td>Thomas Scientific</td><td>3383L10</td><td></td></tr><tr><td>Labconco Lyophilizer</td><td>Labconco Corp.</td><td>7670520</td><td></td></tr><tr><td>&lt;strong&gt;Surgical supplies:&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Betadine</td><td>Purdue Products L.P.</td><td>67618-154-16</td><td></td></tr><tr><td>Lactated Ringers Solution</td><td>MWI Veterinary Supply</td><td>003966</td><td></td></tr><tr><td>KY Jelly</td><td>MWI Veterinary Supply</td><td>28658</td><td></td></tr><tr><td>Lidocaine, 2%</td><td>MWI Veterinary Supply</td><td>17767</td><td></td></tr><tr><td>Buprenorphine hydrochloride</td><td>Reckitt Benckiser</td><td>12496-0757-1</td><td></td></tr><tr><td>Artificial tear ointment</td><td>Fisher Scientific</td><td>NC9860843</td><td></td></tr><tr><td>Triple antibiotic ointment</td><td>Fisher Scientific</td><td>19082795</td><td></td></tr><tr><td>Isoflurane</td><td>MWI Veterinary Supply</td><td>60307-120-25</td><td></td></tr><tr><td>Otoscope </td><td>MWI Veterinary Supply</td><td>008699</td><td></td></tr><tr><td>Stop cock</td><td>MWI Veterinary Supply</td><td>006245</td><td></td></tr><tr><td>3-0 Vicrile suture</td><td>MWI Veterinary Supply</td><td>J327H</td><td></td></tr><tr><td>5-0 Proline suture</td><td>MWI Veterinary Supply</td><td>s-1173</td><td></td></tr><tr><td>Reverse cutting (RC) needle</td><td>Ethicon Inc.</td><td>8684G</td><td></td></tr><tr><td>Microhemostats </td><td>Fine Science Tools</td><td>13013-14</td><td></td></tr><tr><td>Rat tooth microforceps</td><td>Fine Science Tools</td><td>11084-07</td><td></td></tr><tr><td>No. 10 scalpel</td><td>Fine Science Tools</td><td>10110-01</td><td></td></tr><tr><td>Blunt scissors</td><td>Fine Science Tools</td><td>14108-09</td><td></td></tr><tr><td>Sharp, curved scissors</td><td>Fine Science Tools</td><td>14085-08</td><td></td></tr><tr><td>Large, serrated forceps</td><td>Fine Science Tools</td><td>1106-12</td><td></td></tr><tr><td>PE160 suction tubing</td><td>BD Biosciences</td><td>427430</td><td></td></tr><tr><td>Clippers </td><td>MWI Veterinary Supply</td><td>21608</td><td></td></tr><tr><td>Skin staples/stapler </td><td>Ethicon Inc.</td><td>PRR35</td><td></td></tr><tr><td>&lt;strong&gt;General supplies:&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Stir plates</td><td></td><td></td><td></td></tr><tr><td>0.1 M HCl</td><td></td><td></td><td></td></tr><tr><td>1 M NaOH</td><td></td><td></td><td></td></tr><tr><td>10x PBS</td><td></td><td></td><td></td></tr><tr><td>1x PBS</td><td></td><td></td><td></td></tr><tr><td>70% Ethanol</td><td></td><td></td><td></td></tr><tr><td>0.1 mL syringes</td><td></td><td></td><td></td></tr><tr><td>10 mL syringe</td><td></td><td></td><td></td></tr><tr><td>Q-tips</td><td></td><td></td><td></td></tr><tr><td>Surgical glue</td><td></td><td></td><td></td></tr><tr><td>Surgical drape</td><td></td><td></td><td></td></tr><tr><td>Towel clamps</td><td></td><td></td><td></td></tr><tr><td>Small hand-held vacuum </td><td></td><td></td><td></td></tr></tbody></table>

fabrication of biologically derived injectable materials for myocardial tissue engineering

このプロトコルは、心筋組織の工学的応用のための注射細胞外マトリックス（ECM）ゲルの調製のためのメソッドを提供します。簡単に言えば、脱細胞組織は、凍結粉砕、酵素的に消化し、生理的pHにもたらされる。凍結乾燥は、小さなミルで微粉末に粉砕することができる乾燥したECMで、その結果、組織からすべての水分を取り除きます。粉砕後、ECMの粉末を注射マトリックスを形成するために、ペプシンで消化する。 pH7.4に調整した後、液体マトリックス材料は、心筋内に注入することができます。前の特性評価試験の結果は、脱細胞心膜と心筋組織から生成行列のゲルは多様なタンパク質、ペプチドおよびグリコサミノグリカンを含むネイティブなECM成分を、保持することが示されている。組織工学のためのこの物質の使用を考えると、in vivoでの特性評価に特に有用であり、ここ、左心室に壁内注射を行う方法（LV）自由壁がで行列のゲルへのホストの応答を分析する手段として提示され小動物モデル。胸腔へのアクセスは、ダイヤフラムを介して得ていると注射は少しLV自由壁の頂部の上に作られています。生物由来スキャフォールドは、注射の前にビオチン標識によって視覚化し、ホースラディッシュペルオキシダーゼ結合ニュートラビジンで組織切片を染色し、ジアミノベンジジン（DAB）染色を介して可視化することができます。注入領域の解析も組織学的および免疫組織化学的染色を行うことができます。この方法では、以前に調べた心膜と心筋のマトリックスのゲルは、繊維状、多孔質ネットワークを形成し、注入の領域内の血管形成を促進することが示された。

Watch this Scientific Journal Video about Fabrication of Biologically Derived Injectable Materials for Myocardial Tissue Engineering at JoVE.com

Fabrication of Biologically Derived Injectable Materials for Myocardial Tissue Engineering

脱細胞組織から注射マトリックスのゲルを調製し、ラット心筋にそれを注入するための方法<em&gt; in vivoで</em&gt;説明して​​います。

心筋組織工学のための生物由来注射用材料の作製

This protocol provides methods for the preparation of an injectable extracellular matrix (ECM) gel for myocardial tissue engineering applications.  ...

fabrication-biologically-derived-injectable-materials-for-myocardial

Research

JoVE Journal

Bioengineering

15.0K ビュー.  University of California, San Diego. 脱細胞組織から注射マトリックスのゲルを調製し、ラット心筋にそれを注入するための方法 in vivoで説明して​​います。

ビデオ: Fabrication of Biologically Derived Injectable Materials for Myocardial Tissue Engineering

Coronary Artery Ligation and Intramyocardial Injection in a Murine Model of Infarction

Mouse models are a valuable tool for studying acute injury and chronic remodeling of the myocardium in vivo. With the advent of genetic modifications to the whole organism or the myocardium and an array of biological and/or synthetic materials, there is great potential for any combination of these to assuage the extent of acute ischemic injury and impede the onset of heart failure pursuant to myocardial remodeling. 
Here we present the methods and materials used to reliably perform this microsurgery and the modifications involved for temporary (with reperfusion) or permanent coronary artery occlusion studies as well as intramyocardial injections. The effects on the heart that can be seen during the procedure and at the termination of the experiment in addition to histological evaluation will verify efficacy. 
Briefly, surgical preparation involves anesthetizing the mice, removing the fur on the chest, and then disinfecting the surgical area. Intratracheal intubation is achieved by transesophageal illumination using a fiber optic light. The tubing is then connected to a ventilator. An incision made on the chest exposes the pectoral muscles which will be cut to view the ribs. For ischemia/reperfusion studies, a 1 cm piece of PE tubing placed over the heart is used to tie the ligature to so that occlusion/reperfusion can be customized. For intramyocardial injections, a Hamilton syringe with sterile 30gauge beveled needle is used. When the myocardial manipulations are complete, the rib cage, the pectoral muscles, and the skin are closed sequentially. Line block analgesia is effected by 0.25% marcaine in sterile saline which is applied to muscle layer prior to closure of the skin. The mice are given a subcutaneous injection of saline and placed in a warming chamber until they are sternally recumbent. They are then returned to the vivarium and housed under standard conditions until the time of tissue collection. At the time of sacrifice, the mice are anesthetized, the heart is arrested in diastole with KCl or BDM, rinsed with saline, and immersed in fixative. Subsequently, routine procedures for processing, embedding, sectioning, and histological staining are performed. 
Nonsurgical intubation of a mouse and the microsurgical manipulations described make this a technically challenging model to learn and achieve reproducibility. These procedures, combined with the difficulty in performing consistent manipulations of the ligature for timed occlusion(s) and reperfusion or intramyocardial injections, can also affect the survival rate so optimization and consistency are critical.

Numerous genetic manipulations and/or intramyocardial injections of genes, proteins, cells, and/or biomaterials are superimposed upon the dimension of time in studies of acute ischemia/ reperfusion injury and chronic remodeling in mice. This video illustrates the microsurgical procedures for ischemia/reperfusion, permanent coronary artery ligation, and intramyocardial injection studies.

心筋梗塞のマウスモデルにおける冠動脈結紮および心筋内注入

Mouse models are a valuable tool for studying acute injury and chronic remodeling of the myocardium in vivo.  With the advent of genetic modifications to ...

医学

Processing of Human Cardiac Tissue Toward Extracellular Matrix Self-assembling Hydrogel for In Vitro and In Vivo Applications

Acellular extracellular matrix preparations are useful for studying cell-matrix interactions and facilitate regenerative cell therapy applications. Several commercial extracellular matrix products are available as hydrogels or membranes, but these do not possess tissue-specific biological activity. Because perfusion decellularization is usually not possible with human heart tissue, we developed a 3-step immersion decellularization process. Human myocardial slices procured during surgery are first treated with detergent-free hyperosmolar lysis buffer, followed by incubation with the ionic detergent, sodium dodecyl sulfate, and the process is completed by exploiting the intrinsic DNase activity of fetal bovine serum. This technique results in cell-free sheets of cardiac extracellular matrix with largely preserved fibrous tissue architecture and biopolymer composition, which were shown to provide specific environmental cues to cardiac cell populations and pluripotent stem cells. Cardiac extracellular matrix sheets can then be further processed into a microparticle powder without further chemical modification, or, via short-term pepsin digestion, into a self-assembling cardiac extracellular matrix hydrogel with preserved bioactivity.

Processing of Human Cardiac Tissue Toward Extracellular Matrix Self-assembling Hydrogel for In Vitro and In Vivo Applications

This protocol describes the complete decellularization of human myocardium while preserving its extracellular matrix components. Further processing of the extracellular matrix results in the production of microparticles and a cytoprotective self-assembling hydrogel.

ひと心筋細胞外マトリックスの in VitroとIn Vivo応用のハイドロゲルを自己組織化に向けての処理

Acellular extracellular matrix preparations are useful for studying cell-matrix interactions and facilitate regenerative cell therapy applications. ...

発生生物学

An Injectable and Drug-loaded Supramolecular Hydrogel for Local Catheter Injection into the Pig Heart

Regeneration of lost myocardium is an important goal for future therapies because of the increasing occurrence of chronic ischemic heart failure and the limited access to donor hearts. An example of a treatment to recover the function of the heart consists of the local delivery of drugs and bioactives from a hydrogel. In this paper a method is introduced to formulate and inject a drug-loaded hydrogel non-invasively and side-specific into the pig heart using a long, flexible catheter. The use of 3-D electromechanical mapping and injection via a catheter allows side-specific treatment of the myocardium. To provide a hydrogel compatible with this catheter, a supramolecular hydrogel is used because of the convenient switching from a gel to a solution state using environmental triggers. At basic pH this ureido-pyrimidinone modified poly(ethylene glycol) acts as a Newtonian fluid which can be easily injected, but at physiological pH the solution rapidly switches into a gel. These mild switching conditions allow for the incorporation of bioactive drugs and bioactive species, such as growth factors and exosomes as we present here in both in vitro and in vivo experiments. The in vitro experiments give an on forehand indication of the gel stability and drug release, which allows for tuning of the gel and release properties before the subsequent application in vivo. This combination allows for the optimal tuning of the gel to the used bioactive compounds and species, and the injection system.

Supramolecular hydrogelators based on ureido-pyrimidinones allow full control over the macroscopic gel properties and the sol&#8211;gel switching behavior using pH. Here, we present a protocol for formulating and injecting such a supramolecular hydrogelator via a catheter delivery system for local delivery directly in relevant areas in the pig heart.

豚の心臓にローカルカテーテルを注入するための注射や薬物を装填した超分子ヒドロゲル

Regeneration of lost myocardium is an important goal for future therapies because of the increasing occurrence of chronic ischemic heart failure and the ...

生物工学

Ultrasound-guided Transthoracic Intramyocardial Injection in Mice

Murine models of cardiovascular disease are important for investigating pathophysiological mechanisms and exploring potential regenerative therapies. Experiments involving myocardial injection are currently performed by direct surgical access through a thoracotomy. While convenient when performed at the time of another experimental manipulation such as coronary artery ligation, the need for an invasive procedure for intramyocardial delivery limits potential experimental designs. With ever improving ultrasound resolution and advanced noninvasive imaging modalities, it is now feasible to routinely perform ultrasound-guided, percutaneous intramyocardial injection. This modality efficiently and reliably delivers agents to a targeted region of myocardium. Advantages of this technique include the avoidance of surgical morbidity, the facility to target regions of myocardium selectively under ultrasound guidance, and the opportunity to deliver injectate to the myocardium at multiple, predetermined time intervals. With practiced technique, complications from intramyocardial injection are rare, and mice quickly return to normal activity on recovery from anesthetic. Following the steps outlined in this protocol, the operator with basic echocardiography experience can quickly become competent in this versatile, minimally invasive technique.

Echocardiography-guided percutaneous intramyocardial injection represents an efficient, reliable, and targetable modality for the delivery of gene transfer agents or cells into the murine heart. Following the steps outlined in this protocol, the operator can quickly become competent in this versatile, minimally invasive technique.

マウスにおける超音波ガイド下経胸壁心筋内注射

Murine models of cardiovascular disease are important for investigating pathophysiological mechanisms and exploring potential regenerative therapies. ...

3D Microtissues for Injectable Regenerative Therapy and High-throughput Drug Screening

To upgrade traditional 2D cell culture to 3D cell culture, we have integrated microfabrication with cryogelation technology to produce macroporous microscale cryogels (microcryogels), which can be loaded with a variety of cell types to form 3D microtissues. Herein, we present the protocol to fabricate versatile 3D microtissues and their applications in regenerative therapy and drug screening. Size and shape-controllable microcryogels can be fabricated on an array chip, which can be harvested off-chip as individual cell-loaded carriers for injectable regenerative therapy or be further assembled on-chip into 3D microtissue arrays for high-throughput drug screening. Due to the high elastic nature of these microscale cryogels, the 3D microtissues exhibit great injectability for minimally invasive cell therapy by protecting cells from mechanical shear force during injection. This ensures enhanced cell survival and therapeutic effect in the mouse limb ischemia model. Meanwhile, assembly of 3D microtissue arrays in a standard 384-multi-well format facilitates the use of common laboratory facilities and equipment, enabling high-throughput drug screening on this versatile 3D cell culture platform.

This protocol describes the fabrication of elastic 3D macroporous microcryogels by integrating microfabrication with cryogelation technology. Upon loading with cells, 3D microtissues are generated, which can be readily injected in vivo to facilitate regenerative therapy or assembled into arrays for in vitro high-throughput drug screening.

注射の再生療法と高スループットの薬剤のスクリーニングのための 3 D への放射能

To upgrade traditional 2D cell culture to 3D cell culture, we have integrated microfabrication with cryogelation technology to produce macroporous ...

A Hydrogel Construct and Fibrin-based Glue Approach to Deliver Therapeutics in a Murine Myocardial Infarction Model.

The murine MI model is widely recognized in the field of cardiovascular disease, and has consistently been used as a first step to test the efficacy of treatments in vivo1. The traditional, established protocol has been further fine-tuned to minimize the damage to the animal. Notably, the pectoral muscle layers are teased away rather than simply cut, and the thoracotomy is approached intercostally as opposed to breaking the ribs in a sternotomy, preserving the integrity of the ribcage. With these changes, the overall stress on the animal is decreased. 
Stem cell therapies aimed to alleviate the damage caused by MIs have shown promise over the years for their pro-angiogenic and anti-apoptotic benefits. Current approaches of delivering cells to the heart surface typically involve the injection of the cells either near the damaged site, within a coronary artery, or into the peripheral blood stream2-4. While the cells have proven to home to the damaged myocardium, functionality is limited by their poor engraftment at the site of injury, resulting in diffusion into the blood stream5. This manuscript highlights a procedure that overcomes this obstacle with the use of a cell-encapsulated hydrogel patch. The patch is fabricated prior to the surgical procedure and is placed on the injured myocardium immediately following the occlusion of the left coronary artery. To adhere the patch in place, biocompatible external fibrin glue is placed directly on top of the patch, allowing for it to dry to both the patch and the heart surface. This approach provides a novel adhesion method for the application of a delicate cell-encapsulating therapeutic construct.

This protocol aims to alleviate the limitation of poor cell engraftment for stem cell treatment of myocardial infarctions through the use of a hydrogel system and a fibrin-based glue. With this approach, cell-to-tissue contact post-infarction can be maintained, increasing the therapeutic potential of beneficial agents at the site of injury.

ヒドロゲルは、構築し、マウス心筋梗塞モデルにおける治療薬を送達するためのフィブリンベースの接着剤のアプローチ。

The murine MI model is widely recognized in the field of cardiovascular disease, and has consistently been used as a first step to test the efficacy of ...

3D Whole-heart Myocardial Tissue Analysis

Cardiac regenerative therapies aim to protect and repair the injured heart in patients with ischemic heart disease. By injecting stem cells or other biologicals that enhance angio- or vasculogenesis into the infarct border zone (IBZ), tissue perfusion is improved, and the myocardium can be protected from further damage. For maximum therapeutic effect, it is hypothesized that the regenerative substance is best delivered to the IBZ. This requires accurate injections and has led to the development of new injection techniques. To validate these new techniques, we have designed a validation protocol based on myocardial tissue analysis. This protocol includes whole-heart myocardial tissue processing that enables detailed two-dimensional (2D) and three-dimensional (3D) analysis of the cardiac anatomy and intramyocardial injections. In a pig, myocardial infarction was created by a 90-min occlusion of the left anterior descending coronary artery. Four weeks later, a mixture of a hydrogel with superparamagnetic iron oxide particles (SPIOs) and fluorescent beads was injected in the IBZ using a minimally-invasive endocardial approach. 1 h after the injection procedure, the pig was euthanized, and the heart was excised and embedded in agarose (agar). After the solidification of the agar, magnetic resonance imaging (MRI), slicing of the heart, and fluorescence imaging were performed. After image post-processing, 3D analysis was performed to assess the IBZ targeting accuracy. This protocol provides a structured and reproducible method for the assessment of the targeting accuracy of intramyocardial injections into the IBZ. The protocol can be easily used when the processing of scar tissue and/or validation of the injection accuracy of the whole heart is desired.

This protocol describes a novel method for the 3D comparison of whole-heart myocardial tissue with MRI. This is designed for the accurate assessment of intramyocardial injections in the infarct border zone of a chronic porcine model of myocardial infarction.

3Dホール・ハート心筋組織の分析

Cardiac regenerative therapies aim to protect and repair the injured heart in patients with ischemic heart disease. By injecting stem cells or other ...

組織損傷を軽減するための新しい低侵襲スローインジェクション法

A Novel Cell Injection Method with Minimum Invasion

Directly injecting cells into tissues is a necessary process in cell administration and/or replacement therapy. The cell injection requires a sufficient amount of suspension solution to allow the cells to enter the tissue. The volume of the suspension solution affects the tissue, and this can cause major invasive injury as a result of the cell injection. This paper reports on a novel cell injection method, called slow injection, that aims to avoid this injury. However, pushing out the cells from the needle tip requires a sufficiently high injection speed according to Newton's law of shear force. To solve the above contradiction, a non-Newtonian fluid, such as gelatin solution, was used as the cell suspension solution in this work. Gelatins solution have temperature sensitivity, as their form changes from gel to sol at approximately 20 &#176;C. Therefore, to maintain the cell suspension solution in the gel form, the syringe was kept cooled in this protocol; however, once the solution was injected into the body, the body temperature converted it to a sol. The interstitial tissue fluid flow can absorb excess solution. In this work, the slow injection technique allowed cardiomyocyte balls to enter the host myocardium and engraft without surrounding fibrosis. This study employed a slow injection method to inject purified and ball-formed neonatal rat cardiomyocytes into a remote area of myocardial infarction in the adult rat heart. At 2 months following the injection, the hearts of the transplanted groups showed significantly improved contractile function. Furthermore, histological analyses of the slow-injected hearts revealed seamless connections between the host and graft cardiomyocytes via intercalated disks containing gap junction connections. This method could contribute to next-generation cell therapies, particularly in cardiac regenerative medicine.

著者スポットライト:最小限の侵襲で新しい細胞注入法  

This method eliminates any major invasion during cell injections caused by the cell suspension solution.

浸潤の少ない新規細胞注入法

Directly injecting cells into tissues is a necessary process in cell administration and/or replacement therapy. The cell injection requires a sufficient ...

心筋組織工学のための生物由来注射用材料の作製

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心筋梗塞のマウスモデルにおける冠動脈結紮および心筋内注入

ひと心筋細胞外マトリックスの in VitroとIn Vivo応用のハイドロゲルを自己組織化に向けての処理

豚の心臓にローカルカテーテルを注入するための注射や薬物を装填した超分子ヒドロゲル

マウスにおける超音波ガイド下経胸壁心筋内注射

注射の再生療法と高スループットの薬剤のスクリーニングのための 3 D への放射能

ヒドロゲルは、構築し、マウス心筋梗塞モデルにおける治療薬を送達するためのフィブリンベースの接着剤のアプローチ。

3Dホール・ハート心筋組織の分析

浸潤の少ない新規細胞注入法

浸潤の少ない新規細胞注入法

浸潤の少ない新規細胞注入法