This work was supported by the National Institutes of Health (NIH) AI091759 and the Crohn's and Colitis Foundation of America's William and Shelby Modell Family Foundation Research Award (to MN). Schistosome-infected mice were supplied by the NIAID Schistosome Resource Center (NIAID Contract N01 A130026).

1. Purifying Schistosoma mansoni Eggs
<ol>
 <li>Infect Swiss-Webster mice with schistosome cercariae, which is the infectious stage in the Schistosoma life cycle 8. Alternatively, obtain schistosome-infected mice from the NIAID Schistosomiasis Resource Center (<a href="http://www.schisto-resource.org/" target="_blank">http://www.schisto-resource.org/</a>). Leave mice for 6-7 weeks following infection, by which time-point, eggs are present in the liver and can be recovered as described below...

<ol>
	<li>World Health Organization. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prevention+and+control+of+schistosomiasis+and+soil-transmitted+helminthiasis:+report+of+a+WHO+expert+committee.">Prevention and control of schistosomiasis and soil-transmitted helminthiasis: report of a WHO expert committee.</a> WHO Technical Report Series 912. , (2002).</li><li>Pearce, E. J., MacDonald, A. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+immunobiology+of+schistosomiasis.">The immunobiology of schistosomiasis.</a> Nat. Rev. Immunol. 2, 499-511 (2002).</li><li>Sandler, N. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Global+gene+expression+profiles+during+acute+pathogen-induced+pulmonary+inflammation+reveal+divergent+roles+for+Th1+and+th2+responses+in+tissue+repair.">Global gene expression profiles during acute pathogen-induced pulmonary inflammation reveal divergent roles for Th1 and th2 responses in tissue repair.</a> J. Immunol. 171, 3655-3667 (2003).</li><li>Perrigoue, J. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=IL-31-IL-31R+interactions+negatively+regulate+type+2+inflammation+in+the+lung.">IL-31-IL-31R interactions negatively regulate type 2 inflammation in the lung.</a> J. Exp. Med. 204, 481-487 (2007).</li><li>Sandor, M., Weinstock, J. V., Wynn, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Granulomas+in+schistosome+and+mycobacterial+infections:+a+model+of+local+immune+responses.">Granulomas in schistosome and mycobacterial infections: a model of local immune responses.</a> Trends Immunol. 24, 44-52 (2003).</li><li>Nair, M. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alternatively+activated+macrophage-derived+RELM-{alpha}+is+a+negative+regulator+of+type+2+inflammation+in+the+lung.">Alternatively activated macrophage-derived RELM-{alpha} is a negative regulator of type 2 inflammation in the lung.</a> J. Exp. Med. 206, 937-952 (2009).</li><li>Dalton, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+method+for+the+isolation+of+schistosome+eggs+and+miracidia+free+of+contaminating+host+tissues.">A method for the isolation of schistosome eggs and miracidia free of contaminating host tissues.</a> Parasitology. 115, 29-32 (1997).</li><li>Lewis, F. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Large-scale+laboratory+maintenance+of+Schistosoma+mansoni,+with+observations+on+three+schistosome/snail+host+combinations.">Large-scale laboratory maintenance of Schistosoma mansoni, with observations on three schistosome/snail host combinations.</a> J. Parasitol. 72, 813-829 (1986).</li><li>Tilney, N. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Patterns+of+lymphatic+drainage+in+the+adult+laboratory+rat.">Patterns of lymphatic drainage in the adult laboratory rat.</a> J. Anat. 109 (Pt. 3), 369-383 (1971).</li><li>Lewis, F., Coligan, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Schistosomiasis.">Schistosomiasis.</a> Current protocols in immunology. Chapter 19, Unit 19 (2001).</li><li>Kaplan, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Th2+cells+are+required+for+the+Schistosoma+mansoni+egg-induced+granulomatous+response.">Th2 cells are required for the Schistosoma mansoni egg-induced granulomatous response.</a> J. Immunol. 160, 1850-1856 (1998).</li><li>Wynn, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fibrotic+disease+and+the+T(H)1/T(H)2+paradigm.">Fibrotic disease and the T(H)1/T(H)2 paradigm.</a> Nat. Rev. Immunol. 4, 583-594 (2004).</li><li>Cheever, A. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Variation+of+hepatic+fibrosis+and+granuloma+size+among+mouse+strains+infected+with+Schistosoma+mansoni.">Variation of hepatic fibrosis and granuloma size among mouse strains infected with Schistosoma mansoni.</a> Am. J. Trop. Med. Hyg. 37, 85-97 (1987).</li></ol>

Here, a model employing Schistosoma mansoni eggs to induce type 2 lung inflammation is described. Characteristic features of this model include the potent Th2 immune response, airway inflammation and pulmonary granuloma formation. Since these parameters are dependent on CD4 Th2 cell responses 11, this is a useful model to investigate the effects of protein- specific or lineage-specific deletions on Th2 cell responses. Here the readouts would include quantification of antigen-specific Th2 cytokines, an...

<table border="1">
	<tbody>
		<tr>
			<td>Reagent</td>
			<td>Size/Description</td>
			<td>Company</td>
			<td>Catalog #</td>
		</tr>
		<tr>
			<td>Collagenase/Dispase</td>
			<td>500 mg</td>
			<td>Roche Applied Science</td>
			<td>11 097 113 001</td>
		</tr>
		<tr>
			<td>Diff Quik Hema 3 Stain</td>
			<td>Pack</td>
			<td>Fisher Scientific Co. LLC</td>
			<td>122-911</td>
		</tr>
		<tr>
			<td>DMEM</td>
			<td>Liquid</td>
			<td>Gibco</td>
			<td>11965</td>
		</tr>
		<tr>
			<td>Falcon 50mL</td>
			<td>Pack</td>
			<td>Falcon</td>
			<td>352070</td>
		</tr>
		<tr>
			<td>Intramedic PE Tubing</td>
			<td>0.58 mm</td>
			<td>Becton Dickinson and Co.</td>
			<td>427410</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>250 mL</td>
			<td>Abbot Animal Health</td>
			<td>52600405</td>
		</tr>
		<tr>
			<td>Ketamine</td>
			<td>100 mg/mL</td>
			<td>Fort Dodge, Animal Health</td>
			<td>N/A</td>
		</tr>
		<tr>
			<td>Mannose receptor antibody</td>
			<td>Biotin</td>
			<td>AbD Serotec</td>
			<td>MCA2235B</td>
		</tr>
		<tr>
			<td>Metallic Strainer</td>
			<td>Standard kitchen strainer</td>
			<td>Target</td>
			<td>Laurentiis</td>
		</tr>
		<tr>
			<td>Paraformaldehyde</td>
			<td>16%</td>
			<td>Electron Microsc. Sciences</td>
			<td>15710</td>
		</tr>
		<tr>
			<td>Penicillin/ Streptomycin</td>
			<td>100x</td>
			<td>Gibco</td>
			<td>15070</td>
		</tr>
		<tr>
			<td>Percoll</td>
			<td>1 Liter</td>
			<td>GE Healthcare Biosciences</td>
			<td>17 0891 01</td>
		</tr>
		<tr>
			<td>RELM&alpha; antibody</td>
			<td>50 &mu;g</td>
			<td>Peprotech</td>
			<td>500-P214</td>
		</tr>
		<tr>
			<td>Surgical Thread</td>
			<td>2.0 USP 100 yds</td>
			<td>Surgical Specialties Corp.</td>
			<td>SP118</td>
		</tr>
		<tr>
			<td>Syringe Needle</td>
			<td>23g, 3/4 inch</td>
			<td>BD Vacutainer Lab. Med.</td>
			<td>305143</td>
		</tr>
		<tr>
			<td>USA Standard Testing Sieve</td>
			<td>150 &mu;m, 0.0059"</td>
			<td>W.S. Tyler, Inc.</td>
			<td>ASTME-11, Spec. No. 100</td>
		</tr>
		<tr>
			<td>Xylazine</td>
			<td>20 mg/mL</td>
			<td>Butler</td>
			<td>N/A</td>
		</tr>
	</tbody>
</table>

using eggs from schistosoma mansoni as an in vivo model of helminth-induced lung inflammation

Schistosoma parasites are blood flukes that infect an estimated 200 million people worldwide 1. In chronic infection with Schistosoma, the severe pathology, including liver fibrosis and splenomegaly, is caused by the immune response to the parasite eggs rather than the parasite itself 2. Parasite eggs induce a Th2 response characterized by the production of IL-4, IL-5 and IL-13, the alternative activation of macrophages and the recruitment of eosinophils. Here, we describe injection of Schistosoma mansoni eggs as a model to examine parasite-specific Th2 cytokine responses in the lung and draining lymph nodes, the formation of pulmonary granulomas surrounding the egg, and airway inflammation. 
Following intraperitoneal sensitization and intravenous challenge, S. mansoni eggs are transported to the lung via the pulmonary arteries where they are trapped within the lung parenchyma by granulomas composed of lymphocytes, eosinophils and alternatively activated macrophages 3-6. Associated with granuloma formation, inflammation in the broncho-alveolar spaces, expansion of the draining lymph nodes and CD4 T cell activation can be observed. Here we detail the protocol for isolating Schistosoma mansoni eggs from infected livers (modified from 7), sensitizing and challenging mice, and recovering the organs (broncho-alveolar lavage (BAL), lung and draining lymph nodes) for analysis. We also include representative histologic and immunologic data and suggestions for additional immunologic analysis. 
Overall, this method provides an in vivo model to investigate helminth-induced immunologic responses in the lung, which is broadly applicable to the study of Th2 inflammatory diseases including helminth infection, fibrotic diseases, allergic inflammation and asthma. Advantages of this model for the study of type 2 inflammation in the lung include the reproducibility of a potent Th2 inflammatory response in the lung and draining lymph nodes, the ease of assessment of inflammation by histologic examination of the granulomas surrounding the egg, and the potential for long-term storage of the parasite eggs.

Watch this Scientific Journal Video about Using Eggs from Schistosoma mansoni as an In vivo Model of Helminth-induced Lung Inflammation at JoVE.com

Using Eggs from Schistosoma mansoni as an In vivo Model of Helminth-induced Lung Inflammation

Schistosoma mansoni eggs are potent stimulators of the T helper type 2 (Th2) immune response, characteristic of parasite infection, asthma and allergic inflammation. This protocol utilizes S. mansoni egg injection to generate a CD4 Th2 cytokine-induced inflammatory response in the lung, characterized by lung granuloma formation around the egg, eosinophilia and macrophage alternative activation.

Using Eggs from Schistosoma mansoni as an In vivo Model of Helminth-induced Lung Inflammation

Schistosoma parasites are blood flukes that infect an estimated 200 million people worldwide 1. In chronic infection with Schistosoma, the severe ...

using-eggs-from-schistosoma-mansoni-as-an-vivo-model-helminth-induced

Research

JoVE Journal

Immunology and Infection

13.7K vistas.  University of Pennsylvania. Huevos de Schistosoma mansoni son potentes estimuladores del tipo T helper 2 (Th2) la respuesta inmune, característico de la infección por el parásito, el asma y la inflamación alérgica. Este protocolo utiliza S. mansoni de inyección de huevos para generar un recuento de CD4 Th2 citoquinas inducida por la respuesta inflamatoria en el pulmón, caracterizada por la formación de un granuloma pulmonar alrededor del huevo, la eosinofilia y la activaci?...