Name: comprehensive evaluation of the effectiveness and safety of placenta-targeted drug delivery using three complementary methods
Uploaded: 2018-09-10T14:30:00
Description: Watch this Scientific Journal Video about Comprehensive Evaluation of the Effectiveness and Safety of Placenta-Targeted Drug Delivery Using Three Complementary Methods at JoVE.com

Dette arbejde blev støttet af tilskud fra National Natural Sciences Foundation (81771617) og Natural Science Foundation i Guangdong provinsen (2016A030313178) tildeles X.F.; et tilskud fra de Shenzhen grundlæggende Research Fund (JCYJ20170413165233512) tildeles X.F; og Eunice Kennedy Shriver National Institute of Child Health & menneskelig udvikling af National Institutes of Health under Award antal R01HD088549 (indholdet er udelukkende ansvarlig for forfattere og repræsenterer ikke nødvendigvis officielle visninger af National Institutes of Health) til NN

Alle mus eksperimenter følges strengt protokoller (SIAT-IRB-160520-YYS-FXJ-A0232) godkendt af Animal Care og brug Udvalget af Shenzhen Institutes of Advanced Technology, kinesiske Academy of Sciences.
1. Sammenfatning af placenta Chondroitin sulfat A-målrettet Lipid-Polymer nanopartikler
<ol><li>Syntetisere MTX - og regeringskonferencen-loaded lipid-polymer nanopartikler (MNPs og INPs henholdsvis) og plCSA-BP-konjugeret nanopartikler (plCSA-MNPs og plCSA-INPs) som beskrevet i detaljer ande...

<ol>
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W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drug+delivery+to+the+human+and+mouse+uterus+using+immunoliposomes+targeted+to+the+oxytocin+receptor.">Drug delivery to the human and mouse uterus using immunoliposomes targeted to the oxytocin receptor.</a> American Journal of Obstetrics and Gynecology. 216 (3), e281-e283 (2017).</li><li>Zhang, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Placenta-specific+drug+delivery+by+trophoblast-targeted+nanoparticles+in+mice.">Placenta-specific drug delivery by trophoblast-targeted nanoparticles in mice.</a> Theranostics. 8 (10), 2765-2781 (2018).</li><li>Fan, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Noninvasive+monitoring+of+placenta-specific+transgene+expression+by+bioluminescence+imaging.">Noninvasive monitoring of placenta-specific transgene expression by bioluminescence imaging.</a> PloS One. 6 (1), e16348 (2011).</li><li>Murata, M., Tahara, K., Takeuchi, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Real-time+in+vivo+imaging+of+surface-modified+liposomes+to+evaluate+their+behavior+after+pulmonary+administration.">Real-time in vivo imaging of surface-modified liposomes to evaluate their behavior after pulmonary administration.</a> European Journal of Pharmaceutics and Biopharmaceutics. 86 (1), 115-119 (2014).</li><li>Ito, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+whole-body+multimodality+imaging+of+gastric+cancer+peritoneal+metastasis+combining+fluorescence+imaging+with+ICG-labeled+antibody+and+MRI+in+mice.">New whole-body multimodality imaging of gastric cancer peritoneal metastasis combining fluorescence imaging with ICG-labeled antibody and MRI in mice.</a> Gastric Cancer. 17 (3), 497-507 (2014).</li><li>Mazza, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Liposome-Indocyanine+Green+Nanoprobes+for+Optical+Labeling+and+Tracking+of+Human+Mesenchymal+Stem+Cells+Post-Transplantation+In+Vivo.">Liposome-Indocyanine Green Nanoprobes for Optical Labeling and Tracking of Human Mesenchymal Stem Cells Post-Transplantation In Vivo.</a> Advanced Healthcare Materials. 6 (21), (2017).</li><li>Greco, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+frequency+ultrasound+for+in+vivo+pregnancy+diagnosis+and+staging+of+placental+and+fetal+development+in+mice.">High frequency ultrasound for in vivo pregnancy diagnosis and staging of placental and fetal development in mice.</a> PloS One. 8 (10), e77205 (2013).</li><li>Spurney, C. 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I dette manuskript skitsere vi en tre-metode system til bestemmelse af, om plCSA-BP-styrede nanopartikler er et effektivt redskab til at målrette leveringen af narkotika til moderkagen. Brug af i vivo billeddannelse til at overvåge den infrarøde fluorescerende regeringskonferencen signal bekræftet placenta målretning specificiteten af plCSA-BP. Brug HFUS og HPLC, vi viste, at plCSA-BP-konjugeret nanopartikler effektivt kan levere MTX kun til den placenta celler, ikke for fosteret.
<p class="jove_content...

Moderkagen-medieret graviditetskomplikationer, herunder præeklampsi, graviditet tab, placenta abruption og små gestationsalder (SGA), er fælles og føre til betydelige føtal og maternel sygelighed og dødelighed1,2, 3, og meget få narkotika har vist sig for at være effektiv til behandling af graviditet lidelser4,5. Udvikling af strategier for mere selektive og sikrere moderkagen-målrettet medicinafgivelse under graviditet er stadig en udfordring i moderne medicinsk behandling.
I de seneste år, har flere rapporter fokuseret på målrettet levering af medicin til uteroplacental væv af belægning nanopartikler med peptider eller antistoffer som moderkagen-målrettede værktøjer. Disse omfatter en anti-epidermal vækstfaktor receptor (EGFR)6 antistof, tumor-homing peptider (CGKRK og iRGD)7, moderkagen-målrettet peptider8, placenta Vaskulaturen-målrettet peptider9 og antistoffer mod den oxytocin receptor10.
Vi viser her, at en syntetisk placenta chondroitin sulfat A bindende peptid (plCSA-BP) kan bruges til målrettet levering af nanopartikler og deres stof payloads til moderkagen11. PlCSA-BP-styrede nanopartikler er komplementær til den rapporterede uteroplacental målretningsmetoder, fordi de rettet mod de placenta trofoblast.
Som en ikke-invasiv metode, i vivo billeddannelse er blevet brugt til at overvåge moderkagen-specifikke genekspression i mus12og indocyanine grøn (regeringskonferencen) har været meget anvendt til at spore nanopartikler ved hjælp af fluorescens imaging systems13, 14,15. Dermed, vi injiceres intravenøst plCSA-BP-konjugeret nanopartikler fyldt med regeringskonferencen (plCSA-INPs) til at visualisere plCSA-INP distribution hos gravide mus med et fluorescens imager. Vi derefter intravenøst sprøjtet methotrexat (MTX)-påfyldt gravide mus plCSA-NPs. Høj-frekvens ultralyd (HFUS), en anden ikke-invasiv, real-time imaging værktøj16,17 blev brugt til at overvåge fostrets og placenta udvikling i mus. Endelig, vi brugte højtydende væskekromatografi (HPLC) for at kvantificere MTX distribution i placenta og fostre.
I denne protokol beskriver vi i detaljer de tre-metode system bruges til at vurdere effektiviteten af moderkagen-målrettet medicinafgivelse af plCSA-BP-styret nanocarriers.

<table border="0" cellpadding="0" cellspacing="0" style="width:785px;" width="786">
	<tbody>
		<tr>
			<td>CD-1 mice</td>
			<td>Beijing Vital River</td>
			<td>201</td>
			<td>Female (8-12 week)</td>
		</tr>
		<tr>
			<td>Insulin syringe</td>
			<td>BD</td>
			<td>328421</td>
			<td>for IV injection</td>
		</tr>
		<tr>
			<td>Ethanol absolute</td>
			<td>Sinopharm Chemical</td>
			<td>10009218</td>
			<td>for nanoparticles synthesis</td>
		</tr>
		<tr>
			<td>Soybean lecithin</td>
			<td>Avanti Polar Lipids</td>
			<td>441601</td>
			<td>for nanoparticles synthesis</td>
		</tr>
		<tr>
			<td>DSPE-PEG-COOH</td>
			<td>Avanti Polar Lipids</td>
			<td>880125</td>
			<td>for nanoparticles synthesis</td>
		</tr>
		<tr>
			<td>PLGA</td>
			<td>Sigma-Aldrich</td>
			<td>719897</td>
			<td>for nanoparticles synthesis</td>
		</tr>
		<tr>
			<td>Ultrasonic processor</td>
			<td>Sonics</td>
			<td>VCX130</td>
			<td>for nanoparticles synthesis</td>
		</tr>
		<tr>
			<td>Methotrexate (MTX)</td>
			<td>Sigma-Aldrich</td>
			<td>V900324</td>
			<td>for nanoparticles synthesis</td>
		</tr>
		<tr>
			<td>Indocyanine green (ICG)</td>
			<td>Sigma-Aldrich</td>
			<td>1340009</td>
			<td>for in vivo imaging</td>
		</tr>
		<tr>
			<td>phosphate-buffered saline (PBS)</td>
			<td>Hyclone</td>
			<td>SH30028.01</td>
			<td></td>
		</tr>
		<tr>
			<td>IVIS spectrum instrument</td>
			<td>Perkin Elmer</td>
			<td></td>
			<td>for in vivo imaging</td>
		</tr>
		<tr>
			<td>Ultrasound transmission gel</td>
			<td>Guanggong</td>
			<td>ZC4252418</td>
			<td>for ultrasound imaging</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Lunan Pharmaceutical</td>
			<td>I0040</td>
			<td>for maintain the anesthesia</td>
		</tr>
		<tr>
			<td>Depilatory cream</td>
			<td>Nair</td>
			<td>TMG001</td>
			<td>for removing fur</td>
		</tr>
		<tr>
			<td>40 MHz transducer</td>
			<td>VisualSonics</td>
			<td>MS550S</td>
			<td>for ultrasound imaging</td>
		</tr>
		<tr>
			<td>High-frequency ultrasound imaging system</td>
			<td>VisualSonics</td>
			<td>Vevo2100</td>
			<td>for ultrasound imaging</td>
		</tr>
		<tr>
			<td>Avertin</td>
			<td>Sigma-Aldrich</td>
			<td>T48402</td>
			<td>for anesthesia</td>
		</tr>
		<tr>
			<td>Syringe pump</td>
			<td>Mindray</td>
			<td>SK-500III</td>
			<td>forcardiac perfusion</td>
		</tr>
		<tr>
			<td>0.9% saline solution</td>
			<td>Meilunbio</td>
			<td>MA0083</td>
			<td>forcardiac perfusion</td>
		</tr>
		<tr>
			<td>1.5 mL Polypropylene tubes</td>
			<td>AXYGEN</td>
			<td>MCT-150-C</td>
			<td></td>
		</tr>
		<tr>
			<td>-80 &#176;C freezer</td>
			<td>Thermo Fisher Scientific</td>
			<td>88600V</td>
			<td></td>
		</tr>
		<tr>
			<td>Centriguge</td>
			<td>Cence</td>
			<td>H1650R</td>
			<td></td>
		</tr>
		<tr>
			<td>Perchloric acid</td>
			<td>Sigma-Aldrich</td>
			<td>311421</td>
			<td>for precipitating protein</td>
		</tr>
		<tr>
			<td>Homogenizer</td>
			<td>SCIENTZ</td>
			<td>SCIENTZ-48</td>
			<td>for homogenizing tissue</td>
		</tr>
		<tr>
			<td>Syringe filter (0.45 &#956;m)</td>
			<td>Millipore</td>
			<td>SLHV033RS01</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium hydroxide</td>
			<td>Sinopharm Chemical</td>
			<td>10019763</td>
			<td>for solving MTX</td>
		</tr>
		<tr>
			<td>HPLC vials</td>
			<td>Waters</td>
			<td>670650620</td>
			<td>for HPLC</td>
		</tr>
		<tr>
			<td>Potassium phosphate dibasic</td>
			<td>Sinopharm Chemical</td>
			<td>20032117</td>
			<td>for HPLC</td>
		</tr>
		<tr>
			<td>Acetonitrile</td>
			<td>JKchemical</td>
			<td>932537</td>
			<td>for HPLC</td>
		</tr>
		<tr>
			<td>C18 column</td>
			<td>Waters</td>
			<td>186003966</td>
			<td>for HPLC</td>
		</tr>
		<tr>
			<td>HPLC system</td>
			<td>Shimadzu</td>
			<td></td>
			<td>for HPLC</td>
		</tr>
	</tbody>
</table>

comprehensive evaluation of the effectiveness and safety of placenta-targeted drug delivery using three complementary methods

Ingen effektive behandlinger findes for moderkagen-associerede graviditetskomplikationer, og udvikle strategier for målrettet levering af lægemidler til moderkagen samtidig minimere føtal og maternel side effects er stadig en udfordring. Målrettet nanopartikel luftfartsselskaber giver nye muligheder for at behandle placenta lidelser. Vi har for nylig vist, at en syntetisk placenta chondroitin sulfat A bindende peptid (plCSA-BP) kan bruges til at guide nanopartikler til at levere lægemidler til moderkagen. I denne protokol, vi beskriver i detaljer et system til vurdering af effektiviteten af medicinafgivelse til moderkagen af plCSA-BP, der beskæftiger tre separate metoder anvendes i kombination: i vivo billeddannelse, høj frekvens ultralyd (HFUS) og høj ydeevne væskekromatografi (HPLC). Bruger i vivo blev billeddannelse, plCSA-BP-styrede nanopartikler visualiseret i placenta af levende dyr, mens HFUS og HPLC viste, at plCSA-BP-konjugeret nanopartikler effektivt og konkret leveret methotrexat til moderkagen. Således, en kombination af disse metoder kan bruges som et effektivt redskab til målrettet levering af lægemidler til moderkagen og udvikling af ny behandling strategier for flere graviditetskomplikationer.

PlCSA-BP-konjugeret nanopartikler lastet med MTX (plCSA-MNPs) eller regeringskonferencen (plCSA-INPs) var intravenøst injiceres gravide mus i dette håndskrift. In vivo billeddannelse afslørede stærke signaler om regeringskonferencen i regionen i livmoderen 30 min efter plCSA-INP injektion. INPs er primært lokaliseret i lever og milt regionen (figur 1A). På 48 timer efter plCSA-INP injektion, blev gravide mus ofret, afslørende regeringskonferen...

Watch this Scientific Journal Video about Comprehensive Evaluation of the Effectiveness and Safety of Placenta-Targeted Drug Delivery Using Three Complementary Methods at JoVE.com

Comprehensive Evaluation of the Effectiveness and Safety of Placenta-Targeted Drug Delivery Using Three Complementary Methods

Vi beskriver et system, der benytter tre metoder til at evaluere sikkerheden og effektiviteten af moderkagen-målrettet medicinafgivelse: i vivo billeddannelse til at overvåge nanopartikel ophobning, høj frekvens ultralyd til at overvåge placenta og fostrets udvikling , og HPLC at kvantificere medicinafgivelse til væv.

Omfattende evaluering af effektiviteten og sikkerheden af moderkagen-målrettet medicinafgivelse ved hjælp af tre komplementære metoder

No effective treatments currently exist for placenta-associated pregnancy complications, and developing strategies for the targeted delivery of drugs to ...

Research

JoVE Journal

Bioengineering

Self-reporting Scaffolds for 3-Dimensional Cell Culture

Selvrapportering Stilladser for 3-Dimensional Cell Culture

Culturing cells in 3D on appropriate scaffolds is thought to better mimic the in vivo microenvironment and increase cell-cell interactions. The resulting ...

Models and Methods to Evaluate Transport of Drug Delivery Systems Across Cellular Barriers

Modeller og metoder til evaluering Transport af Drug Delivery Systems over cellulære Barrierer

Sub-micrometer carriers (nanocarriers; NCs) enhance efficacy of drugs by improving solubility, stability, circulation time, targeting, and release. ...

Adapting the Electrospinning Process to Provide Three Unique Environments for a Tri-layered In Vitro Model of the Airway Wall

Adapting the Electrospinning Process to Provide Three Unique Environments for a Tri-layered In Vitro Model of the Airway Wall

Tilpasning af Electrospinning proces med at give tre unikke miljøer for en Tri-lags<em&gt; In vitro</em&gt; Model af Airway Wall

Electrospinning is a highly adaptable method producing porous 3D fibrous scaffolds that can be exploited in in vitro cell culture. Alterations to ...

Manufacture and Drug Delivery Applications of Silk Nanoparticles

Fremstilling and Drug Delivery Anvendelser af Silk Nanopartikler

Silk is a promising biopolymer for biomedical and pharmaceutical applications due to its outstanding mechanical properties, biocompatibility and ...

Automated Contraction Analysis of Human Engineered Heart Tissue for Cardiac Drug Safety Screening

Automatiseret Sammentrækning Analyse af humant Engineered Heart Væv til Cardiac Drug Safety Screening

Cardiac tissue engineering describes techniques to constitute three dimensional force-generating engineered tissues. For the implementation of these ...

Use of High-Throughput Automated Microbioreactor System for Production of Model IgG1 in CHO Cells

Brug af høj overførselshastighed automatiseret Microbioreactor System for produktion af Model IgG1 i CHO celler

Automated microscale bioreactors (15 mL) can be a useful tool for cell culture engineers. They facilitate the simultaneous execution of a wide variety of ...

Purification and Analytics of a Monoclonal Antibody from Chinese Hamster Ovary Cells Using an Automated Microbioreactor System

Rensning og analyse af et monoklonalt antistof fra ovarieceller fra kinesiske hamster ved hjælp af et automatiseret Mikrobioreaktor system

Monoclonal antibodies (mAbs) are one of the most popular and well-characterized biological products manufactured today. Most commonly produced using ...

Evaluation of Keratinocyte Proliferation on Two- and Three-dimensional Type I Collagen Substrates

Evaluering af keratinocytter spredning på to - og tre - dimensionelle Type I kollagen substrater

Type I collagen, useful as a substrate for cell culture, exists in two forms: the two-dimensional, non-fibrous form and three-dimensional, fibril form. ...

Preparing Protein Producing Synthetic Cells using Cell Free Bacterial Extracts, Liposomes and Emulsion Transfer

Forberedelse proteinproducerende syntetiske celler ved hjælp af cellefri bakterielle ekstrakter, liposomer og emulsionoverførsel

The bottom-up assembly approach for construction of synthetic cells is an effective tool for isolating and investigating cellular processes in a cell ...

Primary Clarification of CHO Harvested Cell Culture Fluid using an Acoustic Separator

Primær afklaring af CHO Harvested Cell Culture Fluid ved hjælp af en akustisk separator

Primary clarification is an essential step in a biomanufacturing process for the initial removal of cells from therapeutic products within the harvested ...