Bu araştırma genel refah tarafından desteklenen &#38; güvenliği araştırma programı aracılığıyla Ulusal Araştırma Vakfı, Kore (NMG) Milli Eğitim Bakanlığı, Fen ve Teknoloji (NMK-2014M3C8A5030612) tarafından finanse edilen ve Chung Yang, Cha Young tarafından desteklenen Güneş, MD ve Jang Merhaba Joo Yeu Sa Memorial Fonu.

Tüm yöntem tanımlamak burada uluslararası hayvan bakım ve kullanım Komitesi (IACUC), Kore Üniversitesi üniversite ve tıp. tarafından 6 hafta sonra doğum onaylanmış olan, Denemecileri enthansasia ikna etmek ve toplamak cilt ve serum sıçan gibi çeşitli örnekleri.
 1. indüksiyon reklam Neonatal kapsaisin tedavi <ol><li>5 mg/mL kapsaisin çözüm hazırlamak, kapsaisin 50 mg 1 ml Tween 80 ve %100 etanol 15 mL konik tüp içinde 1 mL ekleyin.</li>
	<li>Normal salin 8 mL konik tüp ekleyin. Hiçbir ayrı katmanlar görünür hale gelene kadar tüp şiddetle çalkalanır</li> <li>kapsaisin çözüm (10 µL/g) midline ve neonatal Sprague-Dawley rat boynun arka subkutan doku içine enjekte.
	<ol><li>Teslim (kritik) doğumdan sonra 48 h 12 h arasında kapsaisin. 
		Not: enjeksiyon tarafından birkaç gün gecikirse, dermatit geliştirmek değil veya yalnızca birkaç hafta boyunca devam edecek. Enjeksiyon çok erken, solunum sıkıntısı nedeniyle şiddetli ağrı oluşabilir.</li>
		<li>Yenidoğan sıçanlarından emir kapsaisin enjeksiyon nedeniyle solunum durması önlemek için itmek Yenidoğan Bebeklerin nefes teşvik etmek veya solunum normal. yana capsaicin kaynaklı ağrı olabilir hale gelinceye kadar Yenidoğan Bebeklerin bir oksijen odasında tutmak göğüs nefes vazgeçirmek, genel anestezi ekleme muhtemelen nefes zorluk daha da kötüye. Böylece, Denemecileri anesthesics fareler yaşama artırmak için kullanmamalısınız.</li>
	</ol></li></ol> 2. Kaşıntı ve dermatit reklam, bir fare modeli değerlendirilmesi <ol><li>spontan tırmalamak davranış sayma tarafından prurit ölçümü <ol><li>hazırla plastik chambers (20 cm x 30 cm x 20 cm) için tam kapsama sağlamak için bir ayna ile donatılmış görüntüleme ve nefes farelerin sağlamak için delikleri için.</li>
		<li>Ayarla yukarı ve düzgün ayna sayısı ve fareler açık manzaralarını aynı anda kaydetmek için bir dijital video kamera ayarlayın.</li>
		<li>Yer sıçan plastik odası ve açılmasını engellemek için ağır bir nesne ile odası tavan kapak.</li>
		<li>1 h sıçanlara spontan davranışlarını kayıt, video klipleri oynatmak ve davranışları tırmalamak sayısını saymak.</li>
	</ol></li> <li>Bir skorlama sistemi kullanarak deri lezyonu değerlendirilmesi <ol><li>indüksiyon odasında fare yerleştirin.</li>
		<li>Oksijen akış ölçer 2 L/dk turn akış hızı ve anestezi için % 3 isoflurane Buharlaştırıcı ayarlayın.</li>
		<li>Ölçüde ve dermatit kulaklar ve diğer bölgelerinde bir farenin şiddetini belirlemek.</li>
		<li>0.25 cm 2 ve deri lezyonları aşağıdaki Tablo 1 tarafından şiddeti olarak deri lezyonları ölçüde birim boyutunu define: önem dizin; ölçüde önem çarpımının tarafından Dermatit puanı derecesini (Dermatit puanı görmek < güçlü sınıfı "xfig" = > şekil 1B).</li>
	</ol></li></ol> 3. Gaz halinde olan yakıtlar SK maruz kalma hayvanlar <ol><li>inhalasyon odası hazırlanması <ol><li>bir akrilik cam kutusu hazırlamak (55 cm x 40 cm x 35 cm) karşı taraflarda iki çıkışları olan.</li>
		<li>Dikkat. Dolaşımı SK gaz için giriş yüksek çıkış yerleştirin ve giriş, çıkış boyutunu büyütmek.</li>
		<li>Dikkat. Çıkışları, başka bir tarafında dahili gaz konsantrasyonu ölçümü için kapatılabilmesi için üç küçük pasajlar matkap.</li>
		Her iki Outlet <li>Bağlan tüpler. Bir çıkış tüp kan dolaşımı hood için veya emisyon gaz için bina dışında yere takıp diğer çıkış tüp tıbbi oksijen regülatörü bağlayın.</li>
		<li>Geleneksel fare kafesi akrilik cam kutusunun içine yerleştirir.</li>
		<li>Fareler için 5 hafta (2 h/gün ve haftada 5 gün) doğumdan sonra bir hafta koyarlar.</li>
		<li>Gaz kaçağı önlemek için mümkün olduğunca sıkı bir şekilde akrilik cam kutu mühür. 
		Not: sıkı sızdırmazlık için kauçuk odası kapak içermelidir. Sıkı sızdırmazlık için odası sarmak için sarılmak film uygulamak.</li>
	</ol></li> <li>Bir tıbbi oksijen regülatörü kullanarak gaz halinde olan yakıtlar SK nesil <ol><li>SK 5 g 1 litre su Dissolve.</li>
		<li>%0,5 SK çözüm tıbbi oksijen regülatörü nemlendirici şişeye koy.</li>
		<li>Regülatör, giriş tüp bağladıktan sonra regülatör üzerinde açın ve 5 L/dak için akış hızı ayarlayın</li>
		<li>Çözüm konsantrasyonu ve gaz odasında konsantrasyonu belirlemek için akış hızı ayarlayın.</li>
	</ol>
	</li>
</ol>

<ol>
	<li>Geha, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Allergy+and+hypersensitivity.+Nature+versus+nurture+in+allergy+and+hypersensitivity.">Allergy and hypersensitivity. Nature versus nurture in allergy and hypersensitivity.</a> Curr Opin Immunol. 15 (6), 603-608 (2003).</li><li>Novak, N., Bieber, T., Leung, D. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Immune+mechanisms+leading+to+atopic+dermatitis.">Immune mechanisms leading to atopic dermatitis.</a> The J Allergy Clin Immunol. 112 (Suppl 6), S128-S139 (2003).</li><li>Back, S. K., 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=Chronically+relapsing+pruritic+dermatitis+in+the+rats+treated+as+neonate+with+capsaicin;+a+potential+rat+model+of+human+atopic+dermatitis.">Chronically relapsing pruritic dermatitis in the rats treated as neonate with capsaicin; a potential rat model of human atopic dermatitis.</a> J Dermatol Sci. 67 (2), 111-119 (2012).</li><li>Jin, H., He, R., Oyoshi, M., Geha, R. S. <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+atopic+dermatitis.">Animal models of atopic dermatitis.</a> J Invest Dermatol. 129 (1), 31-40 (2009).</li><li>Han, R. T., 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=Formaldehyde-Induced+Aggravation+of+Pruritus+and+Dermatitis+Is+Associated+with+the+Elevated+Expression+of+Th1+Cytokines+in+a+Rat+Model+of+Atopic+Dermatitis.">Formaldehyde-Induced Aggravation of Pruritus and Dermatitis Is Associated with the Elevated Expression of Th1 Cytokines in a Rat Model of Atopic Dermatitis.</a> PLOS ONE. 11 (12), e0168466 (2016).</li><li>Xu, B., Aoyama, K., Takeuchi, M., Matsushita, T., Takeuchi, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Expression+of+cytokine+mRNAs+in+mice+cutaneously+exposed+to+formaldehyde.">Expression of cytokine mRNAs in mice cutaneously exposed to formaldehyde.</a> Immunol Lett. 84 (1), 49-55 (2002).</li><li>Kim, J. Y., Jeong, M. S., Park, K. Y., Seo, 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=Aggravation+of+atopic+dermatitis-like+symptoms+by+consecutive+low+concentration+of+formaldehyde+exposure+in+NC/Nga+mice.">Aggravation of atopic dermatitis-like symptoms by consecutive low concentration of formaldehyde exposure in NC/Nga mice.</a> Exp Dermatol. 22 (3), 219-221 (2013).</li><li>Laske, N., Niggemann, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Does+the+severity+of+atopic+dermatitis+correlate+with+serum+IgE+levels?.">Does the severity of atopic dermatitis correlate with serum IgE levels?.</a> Pediatr Allergy Immunol. 15 (1), 86-88 (2004).</li><li>Fujii, Y., Takeuchi, H., Sakuma, S., Sengoku, T., Takakura, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+a+2,4-dinitrochlorobenzene-induced+chronic+dermatitis+model+in+rats.">Characterization of a 2,4-dinitrochlorobenzene-induced chronic dermatitis model in rats.</a> Skin Pharmacol Physiol. 22 (5), 240-247 (2009).</li><li>Sugimoto, T., Xiao, C., Ichikawa, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neonatal+primary+neuronal+death+induced+by+capsaicin+and+axotomy+involves+an+apoptotic+mechanism.">Neonatal primary neuronal death induced by capsaicin and axotomy involves an apoptotic mechanism.</a> Brain Res. 807 (1-2), 147-154 (1998).</li><li>Saito, 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=Characterization+of+skin+inflammation+induced+by+repeated+exposure+of+toluene,+xylene,+and+formaldehyde+in+mice.">Characterization of skin inflammation induced by repeated exposure of toluene, xylene, and formaldehyde in mice.</a> Environ Toxicol. 26 (3), 224-232 (2011).</li><li>Jung, W. 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=Formaldehyde+exposure+induces+airway+inflammation+by+increasing+eosinophil+infiltrations+through+the+regulation+of+reactive+oxygen+species+production.">Formaldehyde exposure induces airway inflammation by increasing eosinophil infiltrations through the regulation of reactive oxygen species production.</a> Environ Toixcol Pharmacol. 24 (2), 174-182 (2007).</li></ol>

Yazarlar ifşa gerek yok.

Burada reklam benzeri belirtiler Sıçanlarda indüksiyon için açıklanan protokol reklam patofizyolojisi eğitim için uyarlanmış ve uyuşturucu testi yapmak için bir tarama aracı olarak kullanılan. Gaz halinde olan yakıtlar SK maruz kapsaisin tedavi fareler için araç çeşitli hayvan modelleri çeşitli gazlar maruz etkilerini değerlendirmek için uygulanabilir.
Yenidoğan kapsaisin tedavi reklam modeli DNCB tedavi modeli9 ve epicutaneous Sensitizasyonu mod...

Formaldehit (FA) inhalasyon, Atopik dermatit (AD) gelişimi etkileri sorguya çekmek için bir protokol reklam indüksiyon için kurulan ve geleneksel dolaşım odası SK kronik pozlama için geliştirilmiştir.
Genetik yatkınlık ve çevresel Tetikleyicileri1,2arasındaki karmaşık etkileşimler atfedilen bir kronik enflamatuar deri hastalığı reklamdır. Uygun bir hayvan modeli preklinik durumda reklam çalışması için gereklidir. Bu reklam patogenezinde çevresel faktörlerin etkisi altında elucidating en önemli olur. Ayrıca, daha iyi reklam ilerleme çevre Tetikleyicileri etkilerini araştırmak için bireysel çevresel faktörler ayırmak ve pozlama derecesini ölçmek için önemlidir.
Yenidoğan kapsaisin tedavi tarafından indüklenen reklam roman fare modeli göstermiştir. Bu reklam modeli3 epicutaneous Sensitizasyonu, genetik mühendisliğiyle reklam modelleri ve reklam4spontan fare modelleri tarafından indüklenen reklam modelleri gibi diğer reklam modelleri üzerinde bazı yararları vardır. İlk olarak, bu model yaşında 3 hafta, hangi bebek reklam karşılık gelir ve NC/Nga fare gibi reklam diğer spontan fare modelleri önceki reklam benzeri belirtiler gelişir. Bazı durumlarda, genetik mühendisliğiyle reklam modelleri, aksine reklam modelleri belirtileri çökmek ve kendiliğinden bir 20 haftalık süre içinde Relaps. Ayrıca, reklam benzeri belirtiler modelindeki indüksiyon sürecinin daha Alerjik Kontakt Dermatit indüksiyon işleminin perspektifte benzer epicutaneous Sensitizasyonu modelinin daha kolaydır.
Çocuk ve ergen dönemlerinde SK inhalasyon reklam gelişimi üzerinde etkileri çalışmaya bir akrilik cam odası ve konvansiyonel tıbbi oksijen regülatörü5 kullanarak SK maruz kalma bir hayvan modeli roman Protokolü girmiştik. Regülatör gaz akış oranları ayarlayarak sabit bir konsantrasyon ve çözüm konsantrasyonu hayvanlar ortaya çıkarmak bize izin verdi. Önceki çalışmalarda SK maruz hayvan modelleri için çeşitli yollar bildirdin. Xu et al. hangi SK çözüm için cilt6doğrudan uygulanan pozlama yöntemi açıklanmıştır. Başka bir çalışmada Protokolü SK gaz maruz kalma bir özel uçucu çözücüler jeneratör, modeli 49127kullanarak ele. Önceki çalışmalarda yöntemlerine göre yeni yöntemi daha kolay bulunuyor ve SK ek olarak diğer çözümleri için uygulanabilir.

<table width="776">
	<tbody>
		<tr height="21">
			<td height="21" style="width:231px;height:21px;">Medical Oxygen Regulator</td>
			<td style="width:159px;">Yuyao Jiahua Medical</td>
			<td style="width:129px;"></td>
			<td style="width:257px;">manufacturer will not affect results</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:231px;height:21px;">Tube</td>
			<td style="width:159px;"></td>
			<td style="width:129px;"></td>
			<td>manufacturer will not affect results</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:231px;height:21px;">Acrylic Chamber</td>
			<td style="width:159px;">Taejin Acryl</td>
			<td style="width:129px;"></td>
			<td>manufacturer will not affect results</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:231px;height:21px;">Plastic Chamber</td>
			<td style="width:159px;">Taejin Acryl</td>
			<td style="width:129px;"></td>
			<td>manufacturer will not affect results</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:231px;height:21px;">Paraformaldehyde, powder</td>
			<td style="width:159px;">Sigma</td>
			<td align="right">158127</td>
			<td>manufacturer will not affect results</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:231px;height:21px;">Wrap filme</td>
			<td style="width:159px;">Uniwrap</td>
			<td></td>
			<td>manufacturer will not affect results</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:231px;height:21px;">Capsaicin</td>
			<td style="width:159px;">Sigma</td>
			<td>M2028</td>
			<td>manufacturer will not affect results</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:231px;height:21px;">Tween80</td>
			<td style="width:159px;">Sigma</td>
			<td>P4780</td>
			<td>manufacturer will not affect results</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:231px;height:21px;">Digital Video Camera</td>
			<td style="width:159px;">Sony</td>
			<td>HDR-CX380y</td>
			<td>manufacturer will not affect results</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:231px;height:21px;">Isoflurane</td>
			<td style="width:159px;">JW pharmaceutical</td>
			<td style="width:129px;"></td>
			<td>manufacturer will not affect results</td>
		</tr>
	</tbody>
</table>

effects of exposure of formaldehyde to a rat model of atopic dermatitis induced by neonatal capsaicin treatment

Atopik dermatit kronik pruritik Egzama Relaps ve özellikle gelişmiş ülkelerde dünya çapında hakim. Genetik ve çevresel faktörler arasındaki karmaşık etkileşimler Atopik dermatit patofizyolojisi içinde önemli bir rol oynamak için bilinir. Ancak, hala bu hastalığın patogenezinde detaylı bir resmini eksikliği. Böylece, Atopik dermatit ilerlemesini elucidating için uygun hayvan modelleri geliştirmek için önem taşıyor. Ayrıca, Atopik dermatit hava kirleticiler gibi çevresel faktörlerin etkisini araştıran hastalığın anlayış genişletir. Burada, Yenidoğan kapsaisin tedavi ile Sıçanlarda Atopik dermatit ve formaldehit sürekli konsantrasyon maruz Atopik dermatit çocuk ve ergen gelişimi üzerinde etkileri ortaya çıkarmak için fareler için bir protokol inducing bir yöntem tarif dönemleri. Bu protokoller için çeşitli deneyler başarıyla uygulanan ve diğer maddeler için kullanılabilir.

Kapsaisin tedavi reklam fare modeli 7 hafta-in yaş, AD benzeri deri iltihabı şekil 1Aile sunulur. Reklam sıçan kapsaisin tedavi modelinin sağlam tırmalamak davranışları Video 1gösterilir. Yenidoğan kapsaisin tedavi Dermatit elde edildi ve davranışları Rat 3 hafta-in yaş tırmalamak ağırlaştırılmış ve reklam benzeri belirtiler birkaç hafta (şekil 1B, C) için kalıcı. Der...

Watch this Scientific Journal Video about Effects of Exposure of Formaldehyde to a Rat Model of Atopic Dermatitis Induced by Neonatal Capsaicin Treatment at JoVE.com

Effects of Exposure of Formaldehyde to a Rat Model of Atopic Dermatitis Induced by Neonatal Capsaicin Treatment

Biz burada fareler ve pozlama formaldehit teneffüs Atopik dermatit üzerinde etkilerini araştırmak için buharlaşmış formaldehit Atopik dermatit ikna etmek yenidoğan kapsaisin tedavi yöntemleri açıklanmaktadır.

Formaldehit maruz bir fare modeli Atopik dermatit yenidoğan kapsaisin tedavi tarafından indüklenen etkileri

Atopic dermatitis is chronically relapsing pruritic eczema and prevails around the world especially in developed countries. Complex interactions between ...

effects-exposure-formaldehyde-to-rat-model-atopic-dermatitis-induced

Research

JoVE Journal

Immunology and Infection

9.5K Görüntüleme.  Korea University. Biz burada fareler ve pozlama formaldehit teneffüs Atopik dermatit üzerinde etkilerini araştırmak için buharlaşmış formaldehit Atopik dermatit ikna etmek yenidoğan kapsaisin tedavi yöntemleri açıklanmaktadır.

Video: Effects of Exposure of Formaldehyde to a Rat Model of Atopic Dermatitis Induced by Neonatal Capsaicin Treatment

Obtaining Highly Purified Toxoplasma gondii Oocysts by a Discontinuous Cesium Chloride Gradient

Toxoplasma gondii is an obligate intracellular protozoan pathogen that commonly infects humans. It is a well characterized apicomplexan associated with causing food- and water-borne disease outbreaks. The definitive host is the feline species where sexual replication occurs resulting in the development of the highly infectious and environmentally resistant oocyst. Infection occurs via ingestion of tissue cysts from contaminated meat or oocysts from soil or water. Infection is typically asymptomatic in healthy individuals, but results in a life-long latent infection that can reactivate causing toxoplasmic encephalitis and death if the individual becomes immunocompromised. Meat contaminated with T. gondii cysts have been the primary source of infection in Europe and the United States, but recent changes in animal management and husbandry practices and improved food handling and processing procedures have significantly reduced the prevalence of T. gondii cysts in meat1, 2. Nonetheless, seroprevalence in humans remains relatively high suggesting that exposure from oocyst contaminated soil or water is likely. Indeed, waterborne outbreaks of toxoplasmosis have been reported worldwide supporting the theory exposure to the environmental oocyst form poses a significant health risk3-5. To date, research on understanding the prevalence of T. gondii oocysts in the water and environment are limited due to the lack of tools to detect oocysts in the environment 5, 6. This is primarily due to the lack of efficient purification protocols for obtaining large numbers of highly purified T gondii oocysts from infected cats for research purposes. This study describes the development of a modified CsCl method that easily purifies T. gondii oocysts from feces of infected cats that are suitable for molecular biological and tissue culture manipulation7.

Obtaining Highly Purified Toxoplasma gondii Oocysts by a Discontinuous Cesium Chloride Gradient

This study describes the development of a modified CsCl method that easily purifies T. gondii oocysts from feces of infected cats that are suitable for molecular biological and tissue culture manipulation

Yüksek Saflaştırılmış Alınması Toxoplasma gondii Süreksiz Sezyum Klorür Gradient ookistleri

Toxoplasma gondii is an obligate intracellular protozoan pathogen that commonly infects humans. It is a well characterized apicomplexan associated with ...

İmmünoloji ve Enfeksiyon

Recognition of Epidermal Transglutaminase by IgA and Tissue Transglutaminase 2 Antibodies in a Rare Case of Rhesus Dermatitis

Tissue transglutaminase 2 (tTG2) is an intestinal digestive enzyme which deamidates already partially digested dietary gluten e.g. gliadin peptides. In genetically predisposed individuals, tTG2 triggers autoimmune responses that are characterized by the production of tTG2 antibodies and their direct deposition into small intestinal wall 1,2. The presence of such antibodies constitutes one of the major hallmarks of the celiac disease (CD). Epidermal transglutaminase (eTG) is another member of the transglutaminase family that can also function as an autoantigen in a small minority of CD patients. In these relatively rare cases, eTG triggers an autoimmune reaction (a skin rash) clinically known as dermatitis herpetiformis (DH). Although the exact mechanism of CD and DH pathogenesis is not well understood, it is known that tTG2 and eTG share antigenic epitopes that can be recognized by serum antibodies from both CD and DH patients 3,4.
In this study, the confocal microscopy examination of biopsy samples from skin lesions of two rhesus macaques (Macaca mulatta) with dermatitis (Table 1, Fig. 1 and 2) was used to study the affected tissues. In one animal (EM96) a spectral overlap of IgA and tTG2 antibodies (Fig. 3) was demonstrated. The presence of double-positive tTG2+IgA+ cells was focused in the deep epidermis, around the dermal papillae. This is consistent with lesions described in DH patients 3. When EM96 was placed on a gluten-free diet, the dermatitis, as well as tTG2+IgA+ deposits disappeared and were no longer detectable (Figs. 1-3). Dermatitis reappeared however, based on re-introduction of dietary gluten in EM96 (not shown). In other macaques including animal with unrelated dermatitis, the tTG2+IgA+ deposits were not detected. Gluten-free diet-dependent remission of dermatitis in EM96 together with presence of tTG2+IgA+ cells in its skin suggest an autoimmune, DH-like mechanism for the development of this condition. This is the first report of DH-like dermatitis in any non-human primate.

Recognition of Epidermal Transglutaminase by IgA and Tissue Transglutaminase 2 Antibodies in a Rare Case of Rhesus Dermatitis

Dermatitis herpetiformis (DH) is a chronic inflammatory condition characterized by an autoimmune reaction between IgA and epidermal transglutaminase (eTG). DH develops in a very small portion of gluten-sensitive and/or celiac patients. The results of this study indicate that DH can also develop in a rhesus monkey host with symptoms of idiopatic dermatitis.

Nadir bir Case IgA ve Doku Transglutaminaz 2 Antikorlar Epidermal Transglutaminaz tanınması Rhesus Dermatit

Tissue transglutaminase 2 (tTG2) is an intestinal digestive enzyme which deamidates already partially digested dietary gluten e.g. gliadin peptides. In ...

Murine Model of Allergen Induced Asthma

Asthma is a major cause of morbidity and mortality, affecting some 300 million people throughout the world.1 More than 8% of the US population has asthma, with the prevalence increasing.2 As with other diseases, animal models of allergic airway disease greatly facilitate understanding of the underlying pathophysiology, help identify potential therapeutic targets, and allow preclinical testing of possible new therapies. Models of allergic airway disease have been developed in several animal species, but murine models are particularly attractive due to the low cost, ready availability, and well-characterized immune systems of these animals.3 Availability of a variety of transgenic strains further increases the attractiveness of these models.4 Here we describe two murine models of allergic airway disease, both employing ovalbumin as the antigen. Following initial sensitization by intraperitoneal injection, one model delivers the antigen challenge by nebulization, the other by intratracheal delivery. These two models offer complementary advantages, with each mimicking the major features of human asthma.5
The major features of acute asthma include an exaggerated airway response to stimuli such as methacholine (airway hyperresponsiveness; AHR) and eosinophil-rich airway inflammation. These are also prominent effects of allergen challenge in our murine models,5,6 and we describe techniques for measuring them and thus evaluating the effects of experimental manipulation. Specifically, we describe both invasive7 and non-invasive8 techniques for measuring airway hyperresponsiveness as well as methods for assessing infiltration of inflammatory cells into the airways and the lung. Airway inflammatory cells are collected by bronchoalveolar lavage while lung histopathology is used to assess markers of inflammation throughout the organ. These techniques provide powerful tools for studying asthma in ways that would not be possible in humans.

Experimental mouse models of allergic asthma offer new possibilities for studying disease pathogenesis and developing new therapeutics. These models are well suited to measuring factors governing the allergic immune response, airway inflammation, and pulmonary pathophysiology.

Alerjen Bağlı Astım murin Modeli

Asthma is a major cause of morbidity and mortality, affecting some 300 million people throughout the world.1 More than 8% of the US population has asthma, ...

Primary Microglia Isolation from Mixed Glial Cell Cultures of Neonatal Rat Brain Tissue

Microglia account for approximately 12% of the total cellular population in the mammalian brain. While neurons and astrocytes are considered the major cell types of the nervous system, microglia play a significant role in normal brain physiology by monitoring tissue for debris and pathogens and maintaining homeostasis in the parenchyma via phagocytic activity 1,2. Microglia are activated during a number of injury and disease conditions, including neurodegenerative disease, traumatic brain injury, and nervous system infection 3. Under these activating conditions, microglia increase their phagocytic activity, undergo morpohological and proliferative change, and actively secrete reactive oxygen and nitrogen species, pro-inflammatory chemokines and cytokines, often activating a paracrine or autocrine loop 4-6. As these microglial responses contribute to disease pathogenesis in neurological conditions, research focused on microglia is warranted.
	Due to the cellular heterogeneity of the brain, it is technically difficult to obtain sufficient microglial sample material with high purity during in vivo experiments. Current research on the neuroprotective and neurotoxic functions of microglia require a routine technical method to consistently generate pure and healthy microglia with sufficient yield for study. We present, in text and video, a protocol to isolate pure primary microglia from mixed glia cultures for a variety of downstream applications. Briefly, this technique utilizes dissociated brain tissue from neonatal rat pups to produce mixed glial cell cultures. After the mixed glial cultures reach confluency, primary microglia are mechanically isolated from the culture by a brief duration of shaking. The microglia are then plated at high purity for experimental study.
	The principle and protocol of this methodology have been described in the literature 7,8. Additionally, alternate methodologies to isolate primary microglia are well described 9-12. Homogenized brain tissue may be separated by density gradient centrifugation to yield primary microglia 12. However, the centrifugation is of moderate length (45 min) and may cause cellular damage and activation, as well as, cause enriched microglia and other cellular populations. Another protocol has been utilized to isolate primary microglia in a variety of organisms by prolonged (16 hr) shaking while in culture 9-11. After shaking, the media supernatant is centrifuged to isolate microglia. This longer two-step isolation method may also perturb microglial function and activation. We chiefly utilize the following microglia isolation protocol in our laboratory for a number of reasons: (1) primary microglia simulate in vivo biology more faithfully than immortalized rodent microglia cell lines, (2) nominal mechanical disruption minimizes potential cellular dysfunction or activation, and (3) sufficient yield can be obtained without passage of the mixed glial cell cultures.
	It is important to note that this protocol uses brain tissue from neonatal rat pups to isolate microglia and that using older rats to isolate microglia can significantly impact the yield, activation status, and functional properties of isolated microglia. There is evidence that aging is linked with microglia dysfunction, increased neuroinflammation and neurodegenerative pathologies, so previous studies have used ex vivo adult microglia to better understand the role of microglia in neurodegenerative diseases where aging is important parameter. However, ex vivo microglia cannot be kept in culture for prolonged periods of time. Therefore, while this protocol extends the life of primary microglia in culture, it should be noted that the microglia behave differently from adult microglia and in vitro studies should be carefully considered when translated to an in vivo setting.

Isolating primary microglia from the cellular heterogeneity of the brain is essential to investigate their role in both physiological and pathological conditions. This protocol describes a mechanical isolation and mixed cell culture technique that provides high yield and high purity, viable primary microglial cells for in vitro study and downstream applications.

Yenidoğan Rat Beyin Dokusu Karışık Glial Hücre Kültürleri Primer mikroglia İzolasyonu

Microglia account for approximately 12% of the  total cellular population in the mammalian brain. While neurons and astrocytes are considered  the major ...

A Microplate Assay to Assess Chemical Effects on RBL-2H3 Mast Cell Degranulation: Effects of Triclosan without Use of an Organic Solvent

Mast cells play important roles in allergic disease and immune defense against parasites. Once activated (e.g. by an allergen), they degranulate, a process that results in the exocytosis of allergic mediators. Modulation of mast cell degranulation by drugs and toxicants may have positive or adverse effects on human health. Mast cell function has been dissected in detail with the use of rat basophilic leukemia mast cells (RBL-2H3), a widely accepted model of human mucosal mast cells3-5. Mast cell granule component and the allergic mediator &#946;-hexosaminidase, which is released linearly in tandem with histamine from mast cells6, can easily and reliably be measured through reaction with a fluorogenic substrate, yielding measurable fluorescence intensity in a microplate assay that is amenable to high-throughput studies1. Originally published by Naal et al.1, we have adapted this degranulation assay for the screening of drugs and toxicants and demonstrate its use here.
Triclosan is a broad-spectrum antibacterial agent that is present in many consumer products and has been found to be a therapeutic aid in human allergic skin disease7-11, although the mechanism for this effect is unknown. Here we demonstrate an assay for the effect of triclosan on mast cell degranulation. We recently showed that triclosan strongly affects mast cell function2. In an effort to avoid use of an organic solvent, triclosan is dissolved directly into aqueous buffer with heat and stirring, and resultant concentration is confirmed using UV-Vis spectrophotometry (using &#949;280 = 4,200 L/M/cm)12. This protocol has the potential to be used with a variety of chemicals to determine their effects on mast cell degranulation, and more broadly, their allergic potential.

Mast cell degranulation, the release of allergic mediators, is important in allergy, asthma, and parasite defense. Here we demonstrate techniques1 for assessing effects of drugs and toxicants on degranulation, methodology recently utilized to exhibit the powerful inhibitory effect of antibacterial agent triclosan2.

Organik Solvent kullanımı olmadan Triklosan Etkileri: RBL-2H3 Mast Hücre Degranülasyon üzerinde kimyasal etkileri değerlendirmek için bir Mikroplaka Testi

Mast cells play important roles in allergic disease and immune defense against parasites. Once activated (e.g. by an allergen), they degranulate, a ...

Non-Invasive Model of Neuropathogenic Escherichia coli Infection in the Neonatal Rat

Investigation of the interactions between animal host and bacterial pathogen is only meaningful if the infection model employed replicates the principal features of the natural infection. This protocol describes procedures for the establishment and evaluation of systemic infection due to neuropathogenic Escherichia coli K1 in the neonatal rat. Colonization of the gastrointestinal tract leads to dissemination of the pathogen along the gut-lymph-blood-brain course of infection and the model displays strong age dependency. A strain of E. coli O18:K1 with enhanced virulence for the neonatal rat produces exceptionally high rates of colonization, translocation to the blood compartment and invasion of the meninges following transit through the choroid plexus. As in the human host, penetration of the central nervous system is accompanied by local inflammation and an invariably lethal outcome. The model is of proven utility for studies of the mechanism of pathogenesis, for evaluation of therapeutic interventions and for assessment of bacterial virulence.

Non-Invasive Model of Neuropathogenic Escherichia coli Infection in the Neonatal Rat

Here, a procedure is described for the establishment of systemic infection in the neonatal rat with cultures of Escherichia coli K1. This non-invasive procedure permits colonization of the gastrointestinal tract, translocation of the pathogen to the systemic circulation, and invasion of the central nervous system at the choroid plexus.

Neuropathogenic olmayan İnvaziv Modeli<em&gt; Escherichia coli</em&gt; Yenidoğan Sıçan Enfeksiyon

Investigation of the interactions between animal host and bacterial pathogen is only meaningful if the infection model employed replicates the principal ...

Flow Cytometric Analysis of Particle-bound Bet v 1 Allergen in PM10

Flow cytometry is a method widely used to quantify suspended solids such as cells or bacteria in a size range from 0.5 to several tens of micrometers in diameter. In addition to a characterization of forward and sideward scatter properties, it enables the use of fluorescent labeled markers like antibodies to detect respective structures. Using indirect antibody staining, flow cytometry is employed here to quantify birch pollen allergen (precisely Bet v 1)-loaded particles of 0.5 to 10 &#181;m in diameter in inhalable particulate matter (PM10, particle size &#8804;10 &#181;m in diameter). PM10 particles may act as carriers of adsorbed allergens possibly transporting them to the lower respiratory tract, where they could trigger allergic reactions.
So far the allergen content of PM10 has been studied by means of enzyme linked immunosorbent assays (ELISAs) and scanning electron microscopy. ELISA measures the dissolved and not the particle-bound allergen. Compared to scanning electron microscopy, which can visualize allergen-loaded particles, flow cytometry may additionally quantify them. As allergen content of ambient air can deviate from birch pollen count, allergic symptoms might perhaps correlate better with allergen exposure than with pollen count. In conjunction with clinical data, the presented method offers the opportunity to test in future experiments whether allergic reactions to birch pollen antigens are associated with the Bet v 1 allergen content of PM10 particles &#62;0.5 &#181;m.

Here, we present a protocol to quantify allergen-loaded particles by flow cytometry. Ambient particulate matter particles may act as carriers of adsorbed allergens. We show here that flow cytometry, a method widely used to characterize suspended solids &#62;0.5 &#181;m in diameter, can be used to measure these allergen-loaded particles.

PM10 parçacık bağlı Bet v 1 Allergen Akışı Sitometrik Analizi

Flow cytometry is a method widely used to quantify suspended solids such as cells or bacteria in a size range from 0.5 to several tens of micrometers in ...

A Controlled Mouse Model for Neonatal Polymicrobial Sepsis

Neonatal sepsis remains a global burden. A preclinical model to screen effective prophylactic or therapeutic interventions is needed. Neonatal mouse polymicrobial sepsis can be induced by injecting cecal slurry intraperitoneally into day of life 7 mice and monitoring them for the following week. Presented here are the detailed steps necessary for the implementation of this neonatal sepsis model. This includes making a homogeneous cecal slurry stock, diluting it to a weight- and litter-adjusted dose, an outline of the monitoring schedule, and a definition of observed health categories used to define humane endpoints. The generation of a homogeneous cecal slurry stock from pooled donors allows for the administration into many litters over time, reducing the variation between donors, and preventing the use of potentially toxic glycerol. The monitoring strategy used allows for the anticipation of survival outcome and the identification of mice that would later progress to death, allowing for an earlier identification of the humane endpoint. Two main behavioral features are used to define the health scores, namely, the ability of the neonatal mice to right themselves when placed on their back and their level of mobility. These criteria could potentially be applied to address humane endpoints in other studies of neonatal disease in mice, as long as a pilot study is performed to confirm accuracy. In conclusion, this approach provides a standardized method to model newborn sepsis in mice, while providing resources to assess animal welfare used to define early humane endpoints for challenged animals.

This protocol provides the necessary steps to establish and evaluate neonatal sepsis in 7-day-old mice.

Yenidoğan Polymicrobial Sepsis için kontrollü fare modeli

Neonatal sepsis remains a global burden. A preclinical model to screen effective prophylactic or therapeutic interventions is needed. Neonatal mouse ...

Humanized NOG Mice for Intravaginal HIV Exposure and Treatment of HIV Infection

Humanized mice provide a sophisticated platform to study human immunodeficiency virus (HIV) virology and to test antiviral drugs. This protocol describes the establishment of a human immune system in adult NOG mice. Here, we explain all the practical steps from isolation of umbilical cord blood derived human CD34+ cells and their subsequent intravenous transplantation into the mice, to the manipulation of the model through HIV infection, combination antiretroviral therapy (cART), and blood sampling. Approximately 75,000 hCD34+ cells are injected intravenously into the mice and the level of human chimerism, also known as humanization, in the peripheral blood is estimated longitudinally for months by flow cytometry. A total of 75,000 hCD34+ cells yields 20%&#8211;50% human CD45+ cells in the peripheral blood. The mice are susceptible to intravaginal infection with HIV and blood can be sampled once weekly for analysis, and twice monthly for extended periods. This protocol describes an assay for quantification of plasma viral load using droplet digital PCR (ddPCR). We show how the mice can be effectively treated with a standard-of-care cART regimen in the diet. The delivery of cART in the form of regular mouse chow is a significant refinement of the experimental model. This model can be used for preclinical analysis of both systemic and topical pre-exposure prophylaxis compounds as well as for testing of novel treatments and HIV cure strategies.

We have developed a protocol for the generation and evaluation of a humanized and human immunodeficiency virus-infected NOG mouse model based on stem cell transplant, intravaginal human immunodeficiency virus exposure, and droplet digital PCR RNA quantification.

Intravajinal HIV Maruziyeti ve HIV Enfeksiyonu Tedavisi için İnsanize NOG Fareler

Humanized mice provide a sophisticated platform to study human immunodeficiency virus (HIV) virology and to test antiviral drugs. This protocol describes ...

A Neonatal Imaging Model of Gram-Negative Bacterial Sepsis

Neonates are at an increased risk of bacterial sepsis due to the unique immune profile they display in the first months of life. We have established a protocol for studying the pathogenesis of E. coli O1:K1:H7, a serotype responsible for high mortality rates in neonates. Our method utilizes intravital imaging of neonatal pups at different time points during the progression of infection. This imaging, paralleled by measurement of bacteria in the blood, inflammatory profiling, and tissue histopathology, signifies a rigorous approach to understanding infection dynamics during sepsis. In the current report, we model two infectious inoculums for comparison of bacterial burdens and severity of disease. We find that subscapular infection leads to disseminated infection by 10 h post-infection. By 24 h, infection of luminescent E. coli was abundant in the blood, lungs, and other peripheral tissues. Expression of inflammatory cytokines in the lungs is significant at 24 h, and this is followed by cellular infiltration and evidence of tissue damage that increases with infectious dose. Intravital imaging does have some limitations. This includes a luminescent signal threshold and some complications that can arise with neonates during anesthesia. Despite some limitations, we find that our infection model offers an insight for understanding longitudinal infection dynamics during neonatal murine sepsis, that has not been thoroughly examined to date. We expect this model can also be adapted to study other critical bacterial infections during early life.

Infection of neonatal mice with bioluminescent E. coli O1:K1:H7 results in a septic infection with significant pulmonary inflammation and lung pathology. Here, we describe procedures to model and further study neonatal sepsis using longitudinal intravital imaging in parallel with enumeration of systemic bacterial burdens, inflammatory profiling, and lung histopathology.

Gram-Negatif Bakteriyel Sepsisin Yenidoğan Görüntüleme Modeli

Neonates are at an increased risk of bacterial sepsis due to the unique immune profile they display in the first months of life. We have established a ...