Name: generation of hook ischemia-reperfusion model using a three-day developing chick embryo
Uploaded: 2022-02-19T14:00:00
Description: Watch this Scientific Journal Video about Generation of Hook Ischemia-Reperfusion Model using a Three-Day Developing Chick Embryo at JoVE.com

We want to express our gratitude to Hari Shankar for his critical inputs during videography and editing, Mr. Baqer Hussain for voice-over, Mr. Asghar Rizvi for video editing, Mr. Mohammad Haider for video shoots, Mr. Mohammad Danish Siddiqui for assistance during the experiments.

The Institutional Animal Ethical Committee at Era&#39;s Lucknow Medical College and Hospital issued a written waiver stating that no formal approval was required to conduct these experiments in accordance with the Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA). However, Standard Operating Procedures were followed to minimize any potential for embryonic distress.
1. Buffer preparation (Table 1)
<ol>
	<li>Prepare Ringer&#39;s solution
	...

<ol>
	<li>Fauzia, E., 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=Chick+Embryo:+A+Preclinical+Model+for+Understanding+Ischemia-Reperfusion+Mechanism.">Chick Embryo: A Preclinical Model for Understanding Ischemia-Reperfusion Mechanism.</a> Frontiers in Pharmacology. 21 (9), 1034 (2018).</li><li>Eltzschig, H. K., Eckle, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ischemia+and+reperfusion--from+mechanism+to+translation.">Ischemia and reperfusion--from mechanism to translation.</a> Nature Medicine. 17 (11), 1391-1401 (2011).</li><li>Raza, S. S., 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=Neuroprotective+effect+of+naringenin+is+mediated+through+suppression+of+NF-&#954;B+signaling+pathway+in+experimental+stroke.">Neuroprotective effect of naringenin is mediated through suppression of NF-&#954;B signaling pathway in experimental stroke.</a> Neuroscience. 29 (230), 157-171 (2013).</li><li>Raza, S. S., 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=Hesperidin+ameliorates+functional+and+histological+outcome+and+reduces+neuroinflammation+in+experimental+stroke.">Hesperidin ameliorates functional and histological outcome and reduces neuroinflammation in experimental stroke.</a> Brain Research. 28 (1420), 93-105 (2011).</li><li>Raza, S. S., 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=Silymarin+protects+neurons+from+oxidative+stress+associated+damages+in+focal+cerebral+ischemia:+a+behavioral,+biochemical+and+immunohistological+study+in+Wistar+rats.">Silymarin protects neurons from oxidative stress associated damages in focal cerebral ischemia: a behavioral, biochemical and immunohistological study in Wistar rats.</a> Journal of the Neurological Sciences. 15 (1-2), 45-54 (2011).</li><li>Fan, L., Zhou, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=AG490+protects+cerebral+ischemia/reperfusion+injury+via+inhibiting+the+JAK2/3+signaling+pathway.">AG490 protects cerebral ischemia/reperfusion injury via inhibiting the JAK2/3 signaling pathway.</a> Brain and Behavior. 11 (1), 01911 (2021).</li><li>Wu, M. Y., 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=Current+Mechanistic+Concepts+in+Ischemia+and+Reperfusion+Injury.">Current Mechanistic Concepts in Ischemia and Reperfusion Injury.</a> Cellular Physiology and Biochemistry. 46 (4), 1650-1667 (2018).</li><li>Collard, C. D., Gelman, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pathophysiology,+clinical+manifestations,+and+prevention+of+ischemia-reperfusion+injury.">Pathophysiology, clinical manifestations, and prevention of ischemia-reperfusion injury.</a> Anesthesiology. 94 (6), 1133-1138 (2001).</li><li>Allen, D. D., 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=Cell+lines+as+in+vitro+models+for+drug+screening+and+toxicity+studies.">Cell lines as in vitro models for drug screening and toxicity studies.</a> Drug Development and Industrial Pharmacy. 31 (8), 757-768 (2005).</li><li>Schmeer, C., Gamez, A., Tausch, S., Witte, O. W., Isenmann, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Statins+modulate+heat+shock+protein+expression+and+enhance+retinal+ganglion+cell+survival+after+transient+retinal+ischemia/reperfusion+in+vivo.">Statins modulate heat shock protein expression and enhance retinal ganglion cell survival after transient retinal ischemia/reperfusion in vivo.</a> Investigative Ophthalmology and Visual Science. 49 (11), 4971-4981 (2008).</li><li>Huang, K. Y., 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=A+systematic+review+and+meta-analysis+of+acupuncture+for+improving+learning+and+memory+ability+in+animals.">A systematic review and meta-analysis of acupuncture for improving learning and memory ability in animals.</a> BMC Complementary and Alternative Medicine. 16 (1), 297 (2016).</li><li>Sommer, C. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ischemic+stroke:+Experimental+models+and+reality.">Ischemic stroke: Experimental models and reality.</a> Acta Neuropathologica. 133 (2), 245-261 (2017).</li><li>Yang, W., Chen, J., Meng, Y., Chen, Z., Yang, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+targets+for+treating+ischemia-reperfusion+injury+in+the+liver.">Novel targets for treating ischemia-reperfusion injury in the liver.</a> International Journal of Molecular Sciences. 19 (5), 1302 (2018).</li><li>Seabra, R., Bhogal, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+research+using+early+life+stage+models.">In vivo research using early life stage models.</a> In Vivo. 24 (4), 457-462 (2010).</li><li>Liu, H., 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=Adiponectin+peptide+alleviates+oxidative+stress+and+NLRP3+inflammasome+activation+after+cerebral+ischemia-reperfusion+injury+by+regulating+AMPK/GSK-3beta.">Adiponectin peptide alleviates oxidative stress and NLRP3 inflammasome activation after cerebral ischemia-reperfusion injury by regulating AMPK/GSK-3beta.</a> Experiments in Neurology. 329, 113302 (2020).</li><li>Aboutaleb, N., Jamali, H., Abolhasani, M., Pazoki Toroudi, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lavender+oil+(Lavandula+angustifolia)+attenuates+renal+ischemia/reperfusion+injury+in+rats+through+suppression+of+inflammation,+oxidative+stress+and+apoptosis.">Lavender oil (Lavandula angustifolia) attenuates renal ischemia/reperfusion injury in rats through suppression of inflammation, oxidative stress and apoptosis.</a> Biomedicine and Pharmacotherapy. 110, 9-19 (2019).</li><li>Wallert, 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=alpha-Tocopherol+preserves+cardiac+function+by+reducing+oxidative+stress+and+inflammation+in+ischemia/reperfusion+injury.">alpha-Tocopherol preserves cardiac function by reducing oxidative stress and inflammation in ischemia/reperfusion injury.</a> Redox Biology. 26, 101292 (2019).</li><li>Ashafaq, 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=Catechin+hydrate+ameliorates+redox+imbalance+and+limits+inflammatory+response+in+focal+cerebral+ischemia.">Catechin hydrate ameliorates redox imbalance and limits inflammatory response in focal cerebral ischemia.</a> Neurochemical Research. 37 (8), 1747-1760 (2012).</li><li>Gallagher, S., Chakavarti, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Immunoblot+analysis.">Immunoblot analysis.</a> Journal of Visualized Experiments. 20 (16), 759 (2008).</li><li>Abt, M. A., Grek, C. L., Ghatnekar, G. S., Yeh, E. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation+of+lung+metastasis+in+mouse+mammary+tumor+models+by+quantitative+real-time+PCR.">Evaluation of lung metastasis in mouse mammary tumor models by quantitative real-time PCR.</a> Journal of Visualized Experiments. (107), e53329 (2016).</li><li>Lee, P. Y., Costumbrado, J., Hsu, C. Y., Kim, Y. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Agarose+gel+electrophoresis+for+the+separation+of+DNA+fragments.">Agarose gel electrophoresis for the separation of DNA fragments.</a> Journal of Visualized Experiments. (62), e3923 (2012).</li><li>Wu, Y., 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=Cathelicidin+aggravates+myocardial+ischemia/reperfusion+injury+via+activating+TLR4+signaling+and+P2X(7)R/NLRP3+inflammasome.">Cathelicidin aggravates myocardial ischemia/reperfusion injury via activating TLR4 signaling and P2X(7)R/NLRP3 inflammasome.</a> Journal of Molecular and Cellular Cardiology. 139, 75 (2020).</li><li>Franke, 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=The+NLRP3+inflammasome+drives+inflammation+in+ischemia/reperfusion+injury+after+transient+middle+cerebral+artery+occlusion+in+mice.">The NLRP3 inflammasome drives inflammation in ischemia/reperfusion injury after transient middle cerebral artery occlusion in mice.</a> Brain Behaviour and Immunity. 92, 223 (2021).</li><li>Lawrence, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+nuclear+factor+NF-kappaB+pathway+in+inflammation.">The nuclear factor NF-kappaB pathway in inflammation.</a> Cold Spring Harbor Perspectives in Biology. 1 (6), 001651 (2009).</li><li>Liu, H., 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=Pterostilbene+attenuates+astrocytic+inflammation+and+neuronal+oxidative+injury+after+ischemia-reperfusion+by+inhibiting+NF-kappaB+phosphorylation.">Pterostilbene attenuates astrocytic inflammation and neuronal oxidative injury after ischemia-reperfusion by inhibiting NF-kappaB phosphorylation.</a> Frontiers in Immunology. 10, 2408 (2009).</li><li>Prakash, R., 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=Sivelestat-loaded+nanostructured+lipid+carriers+modulate+oxidative+and+inflammatory+stress+in+human+dental+pulp+and+mesenchymal+stem+cells+subjected+to+oxygen-glucose+deprivation.">Sivelestat-loaded nanostructured lipid carriers modulate oxidative and inflammatory stress in human dental pulp and mesenchymal stem cells subjected to oxygen-glucose deprivation.</a> Materials Science and Engineering: C Materials for Biological Applications. 120, 111700 (2021).</li><li>Prakash, R., 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=Oxidative+stress+enhances+autophagy+in+stem+cells+through+Erk1/2+signaling+pathway+-+implications+for+neurotransplantations.">Oxidative stress enhances autophagy in stem cells through Erk1/2 signaling pathway - implications for neurotransplantations.</a> Stem Cell Reviews and Reports. , (2021).</li><li>Ahmad, 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=Gelatin-coated+polycaprolactone+nanoparticle-mediated+naringenin+delivery+rescue+human+mesenchymal+stem+cells+from+oxygen+glucose+deprivation-induced+inflammatory+stress.">Gelatin-coated polycaprolactone nanoparticle-mediated naringenin delivery rescue human mesenchymal stem cells from oxygen glucose deprivation-induced inflammatory stress.</a> ACS Biomaterials Science and Engineering. 5 (2), 683-695 (2019).</li><li>Guan, 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=The+neuroprotective+effects+of+carvacrol+on+ischemia/reperfusion-induced+hippocampal+neuronal+impairment+by+ferroptosis+mitigation.">The neuroprotective effects of carvacrol on ischemia/reperfusion-induced hippocampal neuronal impairment by ferroptosis mitigation.</a> Life Science. 235, 116795 (2019).</li><li>Jin, Z., Guo, P., Li, X., Ke, J., Wang, Y., Wu, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuroprotective+effects+of+irisin+against+cerebral+ischemia/+reperfusion+injury+via+Notch+signaling+pathway.">Neuroprotective effects of irisin against cerebral ischemia/ reperfusion injury via Notch signaling pathway.</a> Biomedicine and Pharmacotherapy. 120, 109452 (2019).</li><li>Wainrach, S., Sotelo, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electron+microscope+study+of+the+developing+chick+embryo+heart.">Electron microscope study of the developing chick embryo heart.</a> Zeitschrift fur Zellforschung und mikroskopische Anatomie. 55, 622-634 (1961).</li><li>Joshi, V. C., Wilson, A. C., Wakil, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assay+for+the+terminal+enzyme+of+the+stearoyl+coenzyme+A+desaturase+system+using+microsomes.">Assay for the terminal enzyme of the stearoyl coenzyme A desaturase system using microsomes.</a> Journal of Lipid Research. 18 (1), 32-36 (1977).</li><li>Kain, K. H., 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=The+chick+embryo+as+an+expanding+experimental+model+for+cancer+and+cardiovascular+research.">The chick embryo as an expanding experimental model for cancer and cardiovascular research.</a> Development Dynamics. 243 (2), 216-228 (2014).</li><li>Mann, R. A., Moore, K. L., Persaud, T. V. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Limitations+in+the+u~e+of+the+early+chick+embryo+88+a+teratological+model.">Limitations in the u~e of the early chick embryo 88 a teratological model.</a> Teratology. 7, 22-23 (1973).</li><li>Chen, T., Vunjak-Novakovic, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+models+of+ischemia-reperfusion+injury.">In vitro models of ischemia-reperfusion injury.</a> Regenerative English and Translation Medicine. 4 (3), 142-153 (2018).</li><li>Ma, R., 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=Animal+models+of+cerebral+ischemia:+A+review.">Animal models of cerebral ischemia: A review.</a> Biomedicine and Pharmacotherapy. 131, 110686 (2020).</li><li>Bromage, D. I., 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=Remote+ischaemic+conditioning+reduces+infarct+size+in+animal+in+vivo+models+of+ischaemia-reperfusion+injury:+a+systematic+review+and+meta-analysis.">Remote ischaemic conditioning reduces infarct size in animal in vivo models of ischaemia-reperfusion injury: a systematic review and meta-analysis.</a> Cardiovascular Research. 113 (3), 288-297 (2017).</li><li>Kalogeris, T., Baines, C. P., Krenz, M., Korthuis, 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=Cell+biology+of+ischemia/reperfusion+injury.">Cell biology of ischemia/reperfusion injury.</a> International Review of Cell and Molecular Biology. 298, 229-317 (2012).</li><li>Hogers, B., DeRuiter, M. C., Baasten, A. M., Gittenberger-de Groot , A. C., Poelmann, R. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intracardiac+blood+flow+patterns+related+to+the+yolk+sac+circulation+of+the+chick+embryo.">Intracardiac blood flow patterns related to the yolk sac circulation of the chick embryo.</a> Circ Res. 76 (5), 871-877 (1995).</li><li>Rezzola, S., 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=angiogenesis-inflammation+cross+talk+in+diabetic+retinopathy:+novel+insights+from+the+chick+embryo+chorioallantoic+membrane/human+vitreous+platform.">angiogenesis-inflammation cross talk in diabetic retinopathy: novel insights from the chick embryo chorioallantoic membrane/human vitreous platform.</a> Frontiers in Immunology. 11, 581288 (2020).</li></ol>

The goal of ischemia-reperfusion research is to create therapeutic strategies that prevent cell death and promote recovery29,30. To overcome current constraints in I/R research, we designed a Hook&#160;I/R chick embryo model to produce a reliable and reproducible I/R model. To our knowledge, ours is the first I/R model ever created in a 3-day chick embryo for routine I/R experiments, besides studying stress signals (e.g., oxidative and inflammatory stress). Given...

Ischemia-reperfusion tissue injury has been linked to a number of pathologies, including heart attacks, ischemic stroke, trauma, and peripheral vascular disease1,2,3,4,5. This is primarily due to a lack of a comprehensive understanding of the disease progression and the lack of an effective research model. Ischemic injury occurs when the blood supply to a specific area of the tissue is cut off. As a result, ischemic tissue eventually necrotizes, though the rate varies depending on the tissue. Hence, restoring the blood supply may help to mitigate the damage. However, it has been observed, in some cases, that reperfusion causes more tissue damage than ischemia alone does6,7,8. Therefore, understanding the molecular and cellular mechanisms of ischemia-reperfusion is required to develop an effective therapeutic intervention. Currently, no effective treatment for I/R injuries is known. This disparity has prompted the creation of new experimental models, ranging from in vitro to in vivo models, to address the existing problem9,10,11,12,13.
Chick embryos (Gallus gallus domesticus) are widely used in research because of their ease of access, ethical acceptability, relatively large size (compared to other embryos), low cost, and rapid growth14. We used a chick embryo at 72 h of development to create an in ovo I/R by occluding and releasing the right vitelline artery with the assistance of a spinal needle.&#160;We named it the Hook-I/R ischemia-reperfusion model (Figure 1). The model utilized in this study is capable of accurately simulating all downstream processes, including oxidative and inflammatory pathways, which are frequently associated with I/R damage15,16,17.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>(-80&#176;C) freezer</td>
			<td>Haier, China</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>1.5mL Centrifuge tube</td>
			<td>TARSONS, India</td>
			<td>500010X</td>
			<td></td>
		</tr>
		<tr>
			<td>100mm Petri dish (sterile)</td>
			<td>Tarsons, India</td>
			<td>460050</td>
			<td></td>
		</tr>
		<tr>
			<td>18G Needle (18G&#215;1.5 (1.25&#215;38mm)</td>
			<td>Ramsons, India</td>
			<td>13990</td>
			<td></td>
		</tr>
		<tr>
			<td>1mL Syringe</td>
			<td>DISPO VAN</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>26G Needle (26G&#215;1/2 (10.45x13mm)</td>
			<td>DISPO VAN, india</td>
			<td>30722D</td>
			<td></td>
		</tr>
		<tr>
			<td>37&#176;C egg incubator with adjustable percentage humidity</td>
			<td>Gentek, India</td>
			<td>GL-100</td>
			<td></td>
		</tr>
		<tr>
			<td>37&#176;C laboratory incubator</td>
			<td>SCIENCE TECH, India</td>
			<td>CB 101-14</td>
			<td></td>
		</tr>
		<tr>
			<td>3-Methyladenine (3-MA)</td>
			<td>Sigma Aldrich, USA</td>
			<td>M9281</td>
			<td></td>
		</tr>
		<tr>
			<td>3mL Pasture Pipette</td>
			<td>TARSONS, India</td>
			<td>940050</td>
			<td></td>
		</tr>
		<tr>
			<td>50mL Beaker</td>
			<td>TARSONS, India</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>5mL Syringe</td>
			<td>DISPO VAN, India</td>
			<td>IP53</td>
			<td></td>
		</tr>
		<tr>
			<td>70% ethanol</td>
			<td>Merck Millipore, United States</td>
			<td>64-17-5</td>
			<td></td>
		</tr>
		<tr>
			<td>Adhesive tape/Cello tape</td>
			<td>Sunrise, India</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Ambra1 primers</td>
			<td>Applied Biosystems, Foster city, USA</td>
			<td>Hs00387943_m1</td>
			<td></td>
		</tr>
		<tr>
			<td>Anti-mouse IgG</td>
			<td>Cell Signaling Technology, USA</td>
			<td>7076S</td>
			<td></td>
		</tr>
		<tr>
			<td>Anti-Rabbit IgG</td>
			<td>Jackson Immuno Research Laboratories, USA</td>
			<td>711-035-152</td>
			<td></td>
		</tr>
		<tr>
			<td>Atg7</td>
			<td>R&#38;D Systems, USA</td>
			<td>MAB6608</td>
			<td></td>
		</tr>
		<tr>
			<td>Atg7 primers</td>
			<td>Applied Biosystems, Foster city, USA</td>
			<td>Hs00893766_m1</td>
			<td></td>
		</tr>
		<tr>
			<td>Autoclave Bag</td>
			<td>Tarsons, India</td>
			<td>550022</td>
			<td></td>
		</tr>
		<tr>
			<td>Autoclave Machine</td>
			<td>Local made, Lucknow, India</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Beclin-1</td>
			<td>Proteintech, USA</td>
			<td>66665-1-Ig</td>
			<td></td>
		</tr>
		<tr>
			<td>Beta Actin</td>
			<td>ImmunoTag, USA</td>
			<td>ITT07018</td>
			<td></td>
		</tr>
		<tr>
			<td>Bovine Serum Albumin</td>
			<td>Himedia, Mumbai, India</td>
			<td>TC194</td>
			<td></td>
		</tr>
		<tr>
			<td>Calcium Chloride</td>
			<td>Himedia, Mumbai, India</td>
			<td>GRM534</td>
			<td></td>
		</tr>
		<tr>
			<td>Catalase</td>
			<td>ImmunoTag, USA</td>
			<td>ITT5155</td>
			<td></td>
		</tr>
		<tr>
			<td>Cleaning wipes</td>
			<td>Kimberly-Clark, India</td>
			<td>370080</td>
			<td></td>
		</tr>
		<tr>
			<td>Cleaved Caspase3</td>
			<td>ImmunoTag, USA</td>
			<td>ITT07022</td>
			<td></td>
		</tr>
		<tr>
			<td>di-Sodium hydrogen phosphate heptahydrate</td>
			<td>Himedia, Mumbai, India</td>
			<td>GRM39611</td>
			<td></td>
		</tr>
		<tr>
			<td>Doppler blood flowmeter</td>
			<td>Moors instrument, United Kingdom</td>
			<td>moorVMS-LDF1</td>
			<td></td>
		</tr>
		<tr>
			<td>Egg rack</td>
			<td>-</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Egg rack</td>
			<td>-</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>GAPDH</td>
			<td>ImmunoTag, USA</td>
			<td>M1000110</td>
			<td></td>
		</tr>
		<tr>
			<td>GAPDH primers</td>
			<td>Applied Biosystems, Foster city, USA</td>
			<td>Hs02758991_g1</td>
			<td></td>
		</tr>
		<tr>
			<td>Glycine</td>
			<td>Himedia, Mumbai, India</td>
			<td>MB013</td>
			<td></td>
		</tr>
		<tr>
			<td>Kidney tray</td>
			<td>HOSPITO</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>LC3A/B</td>
			<td>Cell Signaling Technology, USA</td>
			<td>4108S</td>
			<td></td>
		</tr>
		<tr>
			<td>Methanol</td>
			<td>Rankem laboratories, Mumbai, India</td>
			<td>M0252</td>
			<td></td>
		</tr>
		<tr>
			<td>Micromanipulator</td>
			<td>Narishige, Japan</td>
			<td>M-152</td>
			<td></td>
		</tr>
		<tr>
			<td>N-acetyl-L-cysteine (NAC)</td>
			<td>Sigma Aldrich, USA</td>
			<td>A7250</td>
			<td></td>
		</tr>
		<tr>
			<td>Naringenin</td>
			<td>Sigma Aldrich, USA</td>
			<td>67604-48-2</td>
			<td></td>
		</tr>
		<tr>
			<td>NF-k&#946;</td>
			<td>Thermo Fisher Scientific, USA</td>
			<td>51-0500</td>
			<td></td>
		</tr>
		<tr>
			<td>NLRP3</td>
			<td>ImmunoTag, USA</td>
			<td>ITT07438</td>
			<td></td>
		</tr>
		<tr>
			<td>Nose plier</td>
			<td>Local made, Lucknow, India</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Ocular forceps</td>
			<td>Stoelting, Germany</td>
			<td>52106-40</td>
			<td></td>
		</tr>
		<tr>
			<td>Ocular iris</td>
			<td>Tufft Surgical Instruments, Jaipur, India</td>
			<td>Hard Age Vannas Micro Scissors Angled 8CM / 3 1/8&#34;</td>
			<td></td>
		</tr>
		<tr>
			<td>OHP marker pen</td>
			<td>Camlin, India</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>ORP-150</td>
			<td>ImmunoTag, USA</td>
			<td>ITT08329</td>
			<td></td>
		</tr>
		<tr>
			<td>Pointed sharp edge scissor</td>
			<td>Stoelting, Germany</td>
			<td>52132-11</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium Chloride</td>
			<td>Himedia, Mumbai, India</td>
			<td>MB043</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium phosphate monobasic anhydrous</td>
			<td>Himedia, Mumbai, India</td>
			<td>MB050</td>
			<td></td>
		</tr>
		<tr>
			<td>Protease Inhibitor</td>
			<td>Abcam, United States</td>
			<td>Ab65621</td>
			<td></td>
		</tr>
		<tr>
			<td>SOD-1</td>
			<td>ImmunoTag, USA</td>
			<td>ITT4364</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium Chloride</td>
			<td>Fisher Scientific, Mumbai, India</td>
			<td>27605</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium dodecyl sulphate</td>
			<td>Himedia, Mumbai, India</td>
			<td>GRM886</td>
			<td></td>
		</tr>
		<tr>
			<td>Spinal needle 25GA; 3.50 IN (90.51 X 90mm)</td>
			<td>Ramson, India</td>
			<td>GS-2029</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereo Zoom surgical microscope</td>
			<td>Olympus, Japan</td>
			<td>SZ2-STU3</td>
			<td></td>
		</tr>
		<tr>
			<td>Syringe discarder</td>
			<td>BIOHAZARD</td>
			<td>882210</td>
			<td></td>
		</tr>
		<tr>
			<td>Toothed forceps</td>
			<td>Stoelting, Germany</td>
			<td>52102-30</td>
			<td></td>
		</tr>
		<tr>
			<td>Tris Base</td>
			<td>G Biosciences, United States</td>
			<td>RC1217</td>
			<td></td>
		</tr>
		<tr>
			<td>Tris Hydrochloric Acid</td>
			<td>Himedia, Mumbai, India</td>
			<td>MB030</td>
			<td></td>
		</tr>
		<tr>
			<td>Tween 20</td>
			<td>G Biosciences, United States</td>
			<td>RC1227</td>
			<td></td>
		</tr>
		<tr>
			<td>White Leghorn Chicken 0-day eggs</td>
			<td>-</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Z-Val-Ala-Asp(OMe)-FMK</td>
			<td>MP Biomedicals, LLC, USA</td>
			<td>FK009</td>
			<td></td>
		</tr>
	</tbody>
</table>

generation of hook ischemia-reperfusion model using a three-day developing chick embryo

Ischemia and reperfusion (I/R) disorders, such as myocardial infarction, stroke, and peripheral vascular disease, are a few of the leading causes of illness and death. Many in vitro and in vivo models are currently available for studying the I/R mechanism in disease or damaged tissues. However, to date, no in ovo I/R model has been reported, which would allow for a better understanding of I/R mechanisms and faster drug screening. This paper describes I/R modeling using a spinal needle customized hook in a 3-day chick embryo to understand I/R development and treatment mechanisms. Our model can be used to investigate anomalies at the DNA, RNA, and protein levels. This method is simple, quick, and inexpensive. The current model can be used independently or in conjunction with existing in vitro and in vivo I/R models.

The Doppler Blood Flow Imaging technique was used to evaluate the effectiveness of our model. In short, we compared the data from the control group with the data from the RVA group to determine the success of our creation. Figure 4A depicts a typical flux associated with the control animal, while Figure 4B depicts the results obtained from an RVA. The numeric 1-8 represents the various events associated with I/R phases. In brief, numeric 1-3 correspond to the ph...

Watch this Scientific Journal Video about Generation of Hook Ischemia-Reperfusion Model using a Three-Day Developing Chick Embryo at JoVE.com

Generation of Hook Ischemia-Reperfusion Model using a Three-Day Developing Chick Embryo

This paper describes ischemia-reperfusion (I/R) modeling in a 3-day chick embryo using a spinal needle customized hook to better understand I/R development and treatment. This model is simple, quick, and inexpensive.

Ischemia and reperfusion (I/R) disorders, such as myocardial infarction, stroke, and peripheral vascular disease, are a few of the leading causes of ...

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