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             <a href="http://openwetware.org/wiki/Biomod/2013/Sendai/sponsor" class="whiteSendai">Sponsor</a>
             <a href="http://openwetware.org/wiki/Biomod/2013/Sendai/sponsor" class="whiteSendai">Sponsor</a>
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             <a href="http://openwetware.org/wiki/Biomod/2013/Sendai"><h1 style="color:white;" ><b>Biomod<span>2013<br>&emsp; Team</span>Sendai</b></h1></a>  
             <a href="http://openwetware.org/wiki/Biomod/2013/Sendai"><h1 style="color:white;" ><b>Biomod<span>2013<br>&emsp;Team</span>Sendai</b></h1></a>  
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        <h2>Project</h2>
<h2>Project</h2>
<h3>Background</h3>
<Img Src="http://openwetware.org/images/1/16/Top%E7%B5%B5-ver2final_mini.jpg"><br>
Recently genomics and proteomics have well studied, and new drugs such as siRNA have been developed. As a further effective approach to cure diseases, drug delivery systems (DDS) in living bodies have been studied. Liposome and polymers are usually used in DDS as containers.<br>
<br>
<h3>Motivation</h3>
Controlling the quantity of drug release is very important to develop effective DDS. <br>
Delivering drugs at appropriate places which are usually very narrow spaces is another important point to avoid adverse effect. <br>
If initial releasing signals are released at a very limited place, and if the signals continuously transduce the initial signals with releasing drugs, are the two important points fulfilled? This is the motivation of our project.<br>
<br>
<h3>Project Goal</h3>
Our project aims to construct a chain-reactive molecule-releasing system in a spontaneous manner like organisms. The system requires two sub-systems: “egg-type initiator” and “chain-reactive burst”. Sequential working of these sub-systems is the goal of our project.<br>
<br>


<h4>1. Egg-type initiator
<h3>Background</h3>
<!--
<a href="http://openwetware.org/wiki/Biomod/2013/Sendai/design#egg">1. Egg-type initiator</a>
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</h4>
<div class="caption-right">
<img src="http://openwetware.org/images/2/21/Project-egg.png"><span>Fig1 Step1. Egg-type initiator</span>
</div>
Eggs are the origin of all animals’ life, and egg-type birth makes it possible to release concentrated chemicals at a narrow space. Such a concentrated release has an advantage to initiate subsequent reactions.<br>
Alginate hydrogel is chosen as a shell membrane of the egg-type. The gels encapsulate solution phase like salmon caviar. The solution phase contains temperature-sensitive liposomes that have some trigger DNAs and chelate compounds inside. Increasing temperature causes collapse of liposomes, and EGTA, a chelate to melt alginate gels, in liposomes is released. As a result, many trigger DNAs are released at a limited point.<br>
<div class="c-both"></div>


<h4>2. Chain-reactive burst
&nbsp;For all living organisms, sensing weak signals in environment and amplifying them are critically important to survive. Various types of signal sensing/ transduction systems have been highly developed in the course of evolution, e.g. immune and neuro-transmission systems.<br>&nbsp;These systems are not only astonishingly sensitive but very effective and efficient. If we could borrow principles from the living systems, we could create a system with novel functionalities.<br>&nbsp;In our project, we decided to create a sensing and signal transduction systems made of designed artificial biomolecules and chemicals.<br><br>
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<div class="caption-right">
<img src="http://openwetware.org/images/2/20/Project-chain.png"><span>Fig2 Step2. Chain-reactive burst</span>
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Chain-reaction is a way to achieve exponential signal amplification. If chain-reactive collapse of liposomes happens in a limited space, release of drugs inside liposomes is expected to be controlled around a limited space. Our chain-reactive burst system is a good one for doing this strategy.<br>
Each liposome encapsulates trigger DNAs and drugs, and has aptamer DNAs in their outer surface. Hybridizations of trigger DNAs and aptamer DNAs deform liposome rapidly, and consequently, the liposomes are destroyed. This destruction continuously occurs by releasing new trigger DNAs inside liposomes to disrupt neighbor liposomes. These processes achieve concentrated drug release. We call these processes as “Chain-reactive burst.” <br>
<div class="c-both"></div>
<br>
<h3>Methods</h3>
To develop the egg-type initiator, following experiments are needed. We plan to create the egg-type initiator by combining these sub-parts.<br>
<br>
(1) Collapse of liposomes by temperature shift<br>
(2) Construction of alginate hydrogel membranes that have many liposomes in their solution phase<br>
(3) Melting the alginate hydrogel membranes by EGTA released from temperature-sensitive liposomes<br>
<br>
To develop the chain-reactive burst, following experiments are needed. <br>
(1) Design and construction of trigger DNA and aptamer DNA to cause collapse of liposomes.<br>
(2) Confirm chain reaction of liposomal collapse by new triggers inside disrupt neighbor liposomes.<br>
 
<!--
 
 
 
 
<h3>Project goal</h3>
Our project goal is construction of egg-type-molecular-robot and chain of molecules-releasing systems by DNA trigger released from egg-type-molecular-robot.</br>
</br>
<ol>
<li>Construction of egg-type-molecular-robot: <br>
this is accomplished by dual structure that have liposome include structure of trigger DNA for chain breaking in alginic acid gel. It sense a stimulation from outside and release many of trigger DNA. For this aim, we achieve ① construction of liposome that sense a stimulation and breaked it. ② construction of alginic acid gel include buffer and many of liposome. ③ Voluntary dissolution of the alginic acid gel by a chelating agent included breaking-liposome. We achieve them for sub goal.</li><br>
 
<li>Construction of molecules-releasing systems: <br>
we break liposome like a chain reaction. Each liposome include many kinds of trigger DNA structures and causes voluntary and sequential chemical reaction by chain-reaction. For realizing this purpose, we achieve next two sub goal;①design and construction of DNA structure which breaks liposome, ② breaking liposome like chain reaction by the DNA that is released from another destruction of  liposome. </li>
 
</ol>
</br>
</p>
 
<p><h3>Merit</h3>
By achieving these subproject goals, we implement chain of molecules-releasing systems from one point. This system releases molecules by diffusion of chain reaction from releasing at the point on liposome-alginate hydro gel double membrane. Thereby a difference arises in the time of reaction, and the order of reactions can be controlable by using it. Therefore, we achieve the system which can control the order of reactions by chain reaction.
</br>
<ol>
<li>Merits of releasing by egg-type-molecular-robot<br>
            -It can start diffusion from one point by reaction of one trigger.<br>
            If there are a lot of triggers around eggs, cahain reactions happens somewhere.<br>
            -It can anneal and release triggers without any effect around environment.<br>
            Because of alginic acid protective membrane, triggers are protected. <br>
 
</li><br>
<li>Merits of liposome chain breaking reactions<br>
            -Using time difference of chain reaction, the order of reactions can be controlled.<br>
</li>
</ol>
</br>
</p>


<h3>Motivation</h3>


            <p><h3>Background</h3>
&nbsp;In order to realize a signal transduction system, we need to develop two types of system; a sensing system that detects external signal and a transmitting system that amplifies the signal and releases large amount of output molecules (payloard). We adopt liposomes as a container of the system. <br><br>
近年、ゲノミクス・プロテオミクス研究によりsiRNAなどの新しい薬が生み出されているが、それらの成否を握るのは、<a href="http://www.dojindo.co.jp/letterj/119/news119.pdf">生体へのデリバリー技術</a>である。DDSはこれに有効なアプローチである。DDSにはリポソームやポリマーが利用され、これらは薬の薬学的・治療の有効性を上げることに寄与しうる。</br>
Recently genomics and proteomics have well studied, and new drugs such as siRNA are created. But, Delivery technology to the living body is important so that those drugs show the true value. (Referred http://www.dojindo.co.jp/letterj/119/news119.pdf) DDS is the effective approach for this. Liposome and a polymer are used for DDS, These contribute to giving the effectiveness of drugs.</br>
</p>
<p><h3>Motivation</h3>
DDSでは、過剰に薬剤が投与されてしまうと、組織に損害を与えてしまう。そこで、制御された薬の放出系が必要となる(<a href="http://www.sciencemag.org/content/303/5665/1818.short">Drug delivery systems: entering the mainstream</a>)。</br>
生体分子のうち、人の手で自在にプログラムから生成を行える分子はDNAである。DNAは反応経路を正確にプログラムでき、論理計算も行えるので(何か論文リンク貼る)、
私たちは運び屋としてリポソームを、制御分子としてDNAを用いて、制御された薬剤放出系を実現しようと考えた。
In conventional DDS, the control of the timing to release drugs is difficult in already sent DDS. A large quantity of trigger is necessary to release a large quantity of drugs.</br>
To solve these problems, we build two systems.　First, the system which release trigger destroying liposome on a certain condition. Second, the system which release drugs exponentially by liposome being destroyed like a chain reaction by trigger. </br>


</p>
<p><h3>Method</h3>
薬の運び屋であるリポソームを、DNAをトリガーとして割る。割ったらリポソームの内容物が放出される。リポソームは有効成分と、DNA構造物を含んでいる。内部に含まれるDNA構造体を、トリガーと同じ構造にすれば、出てきた構造物は、新たなトリガーとなり、再び周囲のリポソームを破壊する。こうして、次々とリポソームの内容物が連鎖的に放出される。</br>
We destroy the liposome which is bagman of drugs with DNA structure with a role of the trigger. If liposome was destroyed, contents of liposome are released, liposome include active ingredient and DNA structure. This DNA structure is the same as trigger DNA structure. Then DNA structure released newly destroy other liposomes again. As such, contents of liposome are released like a chain reaction one after another.</br>
<h4>・The beginning of interaction</h4>
反応を開始するトリガーDNAはリポソーム・アルギン酸二重膜に閉じ込められている。また、リポソーム内部にはキレート剤、ステイプル、M13,尿素が含まれており、高濃度の尿素により、ステイプルとM13は別個に(DNAオリガミ構造体を作らずに)存在している。</br>
リポソーム表面には、ニッパム分子が修飾されており、37℃程度に上げることで、ニッパム分子が収縮しリポソームが破壊される。次に、リポソーム内部に含まれていたキレート剤の効果により、アルギン酸膜が破壊される。37℃条件下で両膜が破壊されると、尿素濃度が低下し、ステイプルとM13からDNAオリガミ構造体が生成され、トリガーとして外に出ていく。</br>
Trigger DNA structure starting reaction is confined in liposome-alginate hydro gel  double membrane In addition, chelating agent, staple, M13mp18, urea are included in the liposome. Because of high density urea, staple and M13mp18 exist separately(without becoming the DNA origami ). </br>
On the surface of liposome, NIPAM molecules are modified. NIPAM molecules shrink at around 37℃ and liposome are destroyed by this effect. Then urea (DNA denaturant) is diluted, and DNA origami is completed without annealing. Furthermore, an alginate hydro gel membrane is destroyed by chelating agent included in the liposome, and a trigger DNA structure is released.</br>


<h4>・The chain reaction</h4>
<h3>Project: Lipo-HANABI</h3>
リポソームをDNAをトリガーとして割るには、まず、リポソームにコレステロール修飾DNAを添加し、表面に一本鎖DNAが現れるようにする。これを、トリガーDNA構造体の一部と相補的な配列を持つ一本鎖DNAとすれば、トリガーDNAはリポソーム表面に多数ハイブリダイゼーションすることになる。</br>
&nbsp;In our system, stored molecules in liposome are released in chain-reaction triggered by environmental stimuli. The system adopts a two-stage mechanism as follows.<br><br>
一本鎖DNA付きリポソームの生成後、相補的な配列をもつトリガーDNA構造物を加える。トリガーDNAがリポソーム表面に付着すると、その物理的な特性により、リポソームが破壊される(影山先輩の数式へリンク貼る。込み入った数式を用いた説明はそこでする)。</br>
リポソームは有効成分と、新たなトリガーを含んでおり、放出された構造物は、連鎖反応的に周囲のリポソームを破壊する。</br>
To destroy liposome with DNA structure as trigger, we add cholesterol-labeled DNA in liposome. Then single strand DNA emerge on the surface of liposome. If we grow the strand which is complementary for this single strand DNA in Trigger DNA structure, a lot of Trigger DNA structure will hybridization on the  surface of liposome. After made liposome with the single strand DNA, we add the trigger DNA structure with a complementary strand. If trigger DNA bind to on the liposomal surface, liposome is destroyed by physical characteristic which come from trigger DNA structure. The liposome includes active ingredient and new trigger DNA structure, and the released structures destroy other liposome for a chain reaction.</br>
</br>


<h4>First stage: Sensing system</h4>
<Img Src="http://openwetware.org/images/1/1d/%3Basfinal.jpg" width="45%" height="45%" style="padding-left:10mm" align="right">


&nbsp;The First stage detects a certain signal in the environment, and release molecules to activate the Second stage. In this project, we chose temperature change as an environmental signal that initiates the whole process.<br>&nbsp;Namely, we use a temperature-sensitive liposome for the First stage. At a certain triggering temperature, they break and release key molecules for the Second stage. <br>&nbsp;The released key molecules attach on the Second stage liposomes and induce puncture of them.


-->
<h4>Second stage: Amplification system</h4>


<Img Src="http://openwetware.org/images/e/e9/Top%E7%B5%B5-final_mini.jpg" width="45%" height="45%" style="padding-left:10mm" align="right">


&nbsp;The Second stage liposomes contain both the payload molecules (e.g. drug) and the same key molecules released by First stage. Once some of them break, it releases more key molecules, and they break other liposomes in their neighborhood. <br>&nbsp;As a consequence, bursting of the liposomes propagates exponentially in a chain-reactive way and releases a lot of payload molecules
The advantage of adopting the two-stage strategy is that various types of signal in the environment can be detected by First stage design, without changing the Second stage.<br>&nbsp;The mechanism of this system is similar to that of HANABI (fireworks in Japanese), therefore, we termed the project "Lipo-HANABI".<br>


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<h2>Project</h2> <Img Src="http://openwetware.org/images/1/16/Top%E7%B5%B5-ver2final_mini.jpg"><br>

<h3>Background</h3>

&nbsp;For all living organisms, sensing weak signals in environment and amplifying them are critically important to survive. Various types of signal sensing/ transduction systems have been highly developed in the course of evolution, e.g. immune and neuro-transmission systems.<br>&nbsp;These systems are not only astonishingly sensitive but very effective and efficient. If we could borrow principles from the living systems, we could create a system with novel functionalities.<br>&nbsp;In our project, we decided to create a sensing and signal transduction systems made of designed artificial biomolecules and chemicals.<br><br>

<h3>Motivation</h3>

&nbsp;In order to realize a signal transduction system, we need to develop two types of system; a sensing system that detects external signal and a transmitting system that amplifies the signal and releases large amount of output molecules (payloard). We adopt liposomes as a container of the system. <br><br>


<h3>Project: Lipo-HANABI</h3> &nbsp;In our system, stored molecules in liposome are released in chain-reaction triggered by environmental stimuli. The system adopts a two-stage mechanism as follows.<br><br>

<h4>First stage: Sensing system</h4> <Img Src="http://openwetware.org/images/1/1d/%3Basfinal.jpg" width="45%" height="45%" style="padding-left:10mm" align="right">

&nbsp;The First stage detects a certain signal in the environment, and release molecules to activate the Second stage. In this project, we chose temperature change as an environmental signal that initiates the whole process.<br>&nbsp;Namely, we use a temperature-sensitive liposome for the First stage. At a certain triggering temperature, they break and release key molecules for the Second stage. <br>&nbsp;The released key molecules attach on the Second stage liposomes and induce puncture of them.

<h4>Second stage: Amplification system</h4>

<Img Src="http://openwetware.org/images/e/e9/Top%E7%B5%B5-final_mini.jpg" width="45%" height="45%" style="padding-left:10mm" align="right">

&nbsp;The Second stage liposomes contain both the payload molecules (e.g. drug) and the same key molecules released by First stage. Once some of them break, it releases more key molecules, and they break other liposomes in their neighborhood. <br>&nbsp;As a consequence, bursting of the liposomes propagates exponentially in a chain-reactive way and releases a lot of payload molecules The advantage of adopting the two-stage strategy is that various types of signal in the environment can be detected by First stage design, without changing the Second stage.<br>&nbsp;The mechanism of this system is similar to that of HANABI (fireworks in Japanese), therefore, we termed the project "Lipo-HANABI".<br> 


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