Name: generation of a mouse spontaneous autoimmune thyroiditis model
Uploaded: 2023-03-17T14:05:00
Description: Watch this Scientific Journal Video about Generation of a Mouse Spontaneous Autoimmune Thyroiditis Model at JoVE.com

تم توفير الأجسام المضادة وحيدة النسيلة للفأر ل TPO البشري (المستخدمة كضوابط إيجابية) من قبل الدكتور P. Carayon والدكتور J. Ruf (مرسيليا ، فرنسا). يشكر المؤلفون جميع المشاركين في هذه الدراسة وأعضاء فريق البحث لدينا. تم دعم هذا العمل جزئيا بمنح من صندوق دعم ما بعد الدكتوراه لمستشفى غرب الصين ، جامعة سيتشوان ، الصين (2020HXBH057) وبرنامج دعم العلوم والتكنولوجيا في مقاطعة سيتشوان (رقم المشروع 2021YFS0166)

تمت الموافقة على البروتوكول الموضح أدناه من خلال إرشادات الرعاية والاستخدام التي وضعتها اللجنة المؤسسية لرعاية واستخدام الحيوان بجامعة سيتشوان.
1. التحضير
<ol><li>إيواء جميع الفئران في ظروف محددة خالية من مسببات الأمراض تحت دورات الضوء والظلام لمدة 12 ساعة (تبدأ في ا...

<ol>
	<li>Ralli, 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=Hashimoto's+thyroiditis:+An+update+on+pathogenic+mechanisms,+diagnostic+protocols,+therapeutic+strategies,+and+potential+malignant+transformation.">Hashimoto's thyroiditis: An update on pathogenic mechanisms, diagnostic protocols, therapeutic strategies, and potential malignant transformation.</a> Autoimmunity Reviews. 19 (10), 102649 (2020).</li><li>Zhang, Q. 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=Lymphocyte+infiltration+and+thyrocyte+destruction+are+driven+by+stromal+and+immune+cell+components+in+Hashimoto's+thyroiditis.">Lymphocyte infiltration and thyrocyte destruction are driven by stromal and immune cell components in Hashimoto's thyroiditis.</a> Nature Communications. 13 (1), 775 (2022).</li><li>Ruggeri, R. 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=Autoimmune+comorbidities+in+Hashimoto's+thyroiditis:+different+patterns+of+association+in+adulthood+and+childhood/adolescence.">Autoimmune comorbidities in Hashimoto's thyroiditis: different patterns of association in adulthood and childhood/adolescence.</a> European Journal of Endocrinology. 176 (2), 133-141 (2017).</li><li>Resende de Paiva, C., Gr&#248;nh&#248;j, C., Feldt-Rasmussen, U., von Buchwald, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Association+between+Hashimoto's+thyroiditis+and+thyroid+cancer+in+64,628+patients.">Association between Hashimoto's thyroiditis and thyroid cancer in 64,628 patients.</a> Frontiers in Oncology. 7, 53 (2017).</li><li>Ehlers, M., Schott, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hashimoto's+thyroiditis+and+papillary+thyroid+cancer:+are+they+immunologically+linked.">Hashimoto's thyroiditis and papillary thyroid cancer: are they immunologically linked.</a> Trends in Endocrinology and Metabolism. 25 (12), 656-664 (2014).</li><li>Medenica, 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=The+role+of+cell+and+gene+therapies+in+the+treatment+of+infertility+in+patients+with+thyroid+autoimmunity.">The role of cell and gene therapies in the treatment of infertility in patients with thyroid autoimmunity.</a> International Journal of Endocrinology. 2022, 4842316 (2022).</li><li>Rose, N. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+genetics+of+autoimmune+thyroiditis:+the+first+decade.">The genetics of autoimmune thyroiditis: the first decade.</a> Journal of Autoimmunity. 37 (2), 88-94 (2011).</li><li>Kolypetri, P., King, J., Larijani, M., Carayanniotis, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genes+and+environment+as+predisposing+factors+in+autoimmunity:+acceleration+of+spontaneous+thyroiditis+by+dietary+iodide+in+NOD.H2(h4)+mice.">Genes and environment as predisposing factors in autoimmunity: acceleration of spontaneous thyroiditis by dietary iodide in NOD.H2(h4) mice.</a> International Reviews of Immunology. 34 (6), 542-556 (2015).</li><li>Terplan, K. L., Witebsky, E., Rose, N. R., Paine, J. R., Egan, R. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+thyroiditis+in+rabbits,+guinea+pigs+and+dogs,+following+immunization+with+thyroid+extracts+of+their+own+and+of+heterologous+species.">Experimental thyroiditis in rabbits, guinea pigs and dogs, following immunization with thyroid extracts of their own and of heterologous species.</a> The American Journal of Pathology. 36 (2), 213-239 (1960).</li><li>Alexopoulos, H., Dalakas, M. C. <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+autoimmune+encephalitides.">The immunobiology of autoimmune encephalitides.</a> Journal of Autoimmunity. 104, 102339 (2019).</li><li>Noviello, C. M., Kreye, J., Teng, J., Pr&#252;ss, H., Hibbs, 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=Structural+mechanisms+of+GABA+receptor+autoimmune+encephalitis.">Structural mechanisms of GABA receptor autoimmune encephalitis.</a> Cell. 185 (14), 2469-2477 (2022).</li><li>Pudifin, D. J., Duursma, J., Brain, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+autoimmune+thyroiditis+in+the+vervet+monkey.">Experimental autoimmune thyroiditis in the vervet monkey.</a> Clinical and Experimental Immunology. 29 (2), 256-260 (1977).</li><li>Esquivel, P. S., Rose, N. R., Kong, Y. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+of+autoimmunity+in+good+and+poor+responder+mice+with+mouse+thyroglobulin+and+lipopolysaccharide.">Induction of autoimmunity in good and poor responder mice with mouse thyroglobulin and lipopolysaccharide.</a> The Journal of Experimental Medicine. 145 (5), 1250-1263 (1977).</li><li>Kong, Y. C., 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=HLA-DRB1+polymorphism+determines+susceptibility+to+autoimmune+thyroiditis+in+transgenic+mice:+definitive+association+with+HLA-DRB1*0301+(DR3)+gene.">HLA-DRB1 polymorphism determines susceptibility to autoimmune thyroiditis in transgenic mice: definitive association with HLA-DRB1*0301 (DR3) gene.</a> The Journal of Experimental Medicine. 184 (3), 1167-1172 (1996).</li><li>Kotani, T., Umeki, K., Hirai, K., Ohtakia, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+murine+thyroiditis+induced+by+porcine+thyroid+peroxidase+and+its+transfer+by+the+antigen-specific+T+cell+line.">Experimental murine thyroiditis induced by porcine thyroid peroxidase and its transfer by the antigen-specific T cell line.</a> Clinical and Experimental Immunology. 80 (1), 11-18 (1990).</li><li>Ng, H. P., Banga, J. P., Kung, A. W. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+a+murine+model+of+autoimmune+thyroiditis+induced+with+homologous+mouse+thyroid+peroxidase.">Development of a murine model of autoimmune thyroiditis induced with homologous mouse thyroid peroxidase.</a> Endocrinology. 145 (2), 809-816 (2004).</li><li>Ng, H. P., Kung, A. W. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+of+autoimmune+thyroiditis+and+hypothyroidism+by+immunization+of+immunoactive+T+cell+epitope+of+thyroid+peroxidase.">Induction of autoimmune thyroiditis and hypothyroidism by immunization of immunoactive T cell epitope of thyroid peroxidase.</a> Endocrinology. 147 (6), 3085-3092 (2006).</li><li>Ellis, J. S., Braley-Mullen, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+by+which+B+cells+and+regulatory+T+Cells+influence+development+of+murine+organ-specific+autoimmune+diseases.">Mechanisms by which B cells and regulatory T Cells influence development of murine organ-specific autoimmune diseases.</a> Journal of Clinical Medicine. 6 (2), 13 (2017).</li><li>Fang, Y., Yu, S., Braley-Mullen, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contrasting+roles+of+IFN-gamma+in+murine+models+of+autoimmune+thyroid+diseases.">Contrasting roles of IFN-gamma in murine models of autoimmune thyroid diseases.</a> Thyroid. 17 (10), 989-994 (2007).</li><li>Fang, Y., Zhao, L., Yan, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemokines+as+novel+therapeutic+targets+in+autoimmune+thyroiditis.">Chemokines as novel therapeutic targets in autoimmune thyroiditis.</a> Recent Patents on DNA &amp; Gene Sequences. 4 (1), 52-57 (2010).</li><li>Chen, C. 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=Antibodies+to+thyroid+peroxidase+arise+spontaneously+with+age+in+NOD.H-2h4+mice+and+appear+after+thyroglobulin+antibodies.">Antibodies to thyroid peroxidase arise spontaneously with age in NOD.H-2h4 mice and appear after thyroglobulin antibodies.</a> Endocrinology. 151 (9), 4583-4593 (2010).</li><li>Ruf, J., 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=Relationship+between+immunological+structure+and+biochemical+properties+of+human+thyroid+peroxidase.">Relationship between immunological structure and biochemical properties of human thyroid peroxidase.</a> Endocrinology. 125 (3), 1211-1218 (1989).</li><li>McLachlan, S. M., Aliesky, H. A., Chen, C. R., Chong, G., Rapoport, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Breaking+tolerance+in+transgenic+mice+expressing+the+human+TSH+receptor+A-subunit:+thyroiditis,+epitope+spreading+and+adjuvant+as+a+'double+edged+sword'.">Breaking tolerance in transgenic mice expressing the human TSH receptor A-subunit: thyroiditis, epitope spreading and adjuvant as a 'double edged sword'.</a> PLoS One. 7 (9), e43517 (2012).</li><li>McLachlan, S. M., Aliesky, H. A., Chen, C. 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=Breaking+tolerance+in+transgenic+mice+expressing+the+human+TSH+receptor+A-subunit:+thyroiditis,+epitope+spreading+and+adjuvant+as+a+'double+edged+sword’[J].">Breaking tolerance in transgenic mice expressing the human TSH receptor A-subunit: thyroiditis, epitope spreading and adjuvant as a 'double edged sword’[J].</a> PLoS One. 7 (9), e43517 (2012).</li><li>Hutchings, P. 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=Both+CD4(+)+T+cells+and+CD8(+)+T+cells+are+required+for+iodine+accelerated+thyroiditis+in+NOD+mice.">Both CD4(+) T cells and CD8(+) T cells are required for iodine accelerated thyroiditis in NOD mice.</a> Cellular Immunology. 192 (2), 113-121 (1999).</li><li>Xue, 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=Dynamic+changes+of+CD4+CD25+++regulatory+T+cells+in+NOD.H-2h4+mice+with+iodine-induced+autoimmune+thyroiditis.">Dynamic changes of CD4+CD25 + regulatory T cells in NOD.H-2h4 mice with iodine-induced autoimmune thyroiditis.</a> Biological Trace Element Research. 143 (1), 292-301 (2011).</li><li>Hou, 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=Effect+of+halofuginone+on+the+pathogenesis+of+autoimmune+thyroid+disease+in+different+mice+models.">Effect of halofuginone on the pathogenesis of autoimmune thyroid disease in different mice models.</a> Endocrine, Metabolic &amp; Immune Disorders Drug Targets. 17 (2), 141-148 (2017).</li><li>McLachlan, S. 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=Dissociation+between+iodide-induced+thyroiditis+and+antibody-mediated+hyperthyroidism+in+NOD.H-2h4+mice.">Dissociation between iodide-induced thyroiditis and antibody-mediated hyperthyroidism in NOD.H-2h4 mice.</a> Endocrinology. 146 (1), 294-300 (2005).</li><li>Danailova, 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=Nutritional+management+of+thyroiditis+of+hashimoto.">Nutritional management of thyroiditis of hashimoto.</a> International Journal of Molecular Sciences. 23 (9), 5144 (2022).</li><li>Carayanniotis, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+parameters+linking+thyroglobulin+iodination+with+autoimmune+thyroiditis.">Molecular parameters linking thyroglobulin iodination with autoimmune thyroiditis.</a> Hormones. 10 (1), 27-35 (2011).</li><li>Verginis, P., Li, H. S., Carayanniotis, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tolerogenic+semimature+dendritic+cells+suppress+experimental+autoimmune+thyroiditis+by+activation+of+thyroglobulin-specific+CD4+CD25++T+cells.">Tolerogenic semimature dendritic cells suppress experimental autoimmune thyroiditis by activation of thyroglobulin-specific CD4+CD25+ T cells.</a> Journal of Immunology. 174 (11), 7433-7439 (2005).</li><li>Flynn, J. C., 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=Superiority+of+thyroid+peroxidase+DNA+over+protein+immunization+in+replicating+human+thyroid+autoimmunity+in+HLA-DRB1*0301+(DR3)+transgenic+mice.">Superiority of thyroid peroxidase DNA over protein immunization in replicating human thyroid autoimmunity in HLA-DRB1*0301 (DR3) transgenic mice.</a> Clinical and Experimental Immunology. 137 (3), 503-512 (2004).</li><li>Akeno, N., 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=IFN-&#945;+mediates+the+development+of+autoimmunity+both+by+direct+tissue+toxicity+and+through+immune+cell+recruitment+mechanisms.">IFN-&#945; mediates the development of autoimmunity both by direct tissue toxicity and through immune cell recruitment mechanisms.</a> Journal of Immunology. 186 (8), 4693-4706 (2011).</li></ol>

يحدث HT بسبب اضطراب في جهاز المناعة الذاتية الناجم عن الخلايا الليمفاوية التي تتسلل إلى الغدة الدرقية ، مما يزيد من ضعف وظيفة الغدة الدرقية ، مع إنتاج أجسام مضادة خاصة بالغدة الدرقية. مستويات مصل TSH و TgAb و TPOAb في مرضى HT مرتفعة بشكل ملحوظ27. في الوقت الحاضر ، يتم استخدام نوعين رئيس?...

An erratum was issued for:&#160;<a href="https://www.jove.com/t/64609">Generation of a Mouse Spontaneous Autoimmune Thyroiditis Model</a>. The Protocol section was updated.
Step 3.1.1 of the Protocol was updated from:
After the induction, anesthetize the mice with a volume of 0.01 mL/g anesthetic by intraperitoneal injection. Prepare the anesthetic by mixing midazolam (40 &#181;g/100 &#181;L for sedation), medetomidine (7.5 &#181;g/100 &#181;L for sedation), and butorphanol tartrate (50 &#181;g/100 &#181;L for analgesia) in phosphate-buffered saline (PBS).
to
After the induction, anesthetize the mice with a volume of 0.01 mL/g anesthetic by intraperitoneal injection. Prepare the anesthetic by mixing midazolam (40 &#181;g/100 &#181;L for sedation), medetomidine (7.5 &#181;g/100 &#181;L for sedation), and butorphanol tartrate (50 &#181;g/100 &#181;L for analgesia) in phosphate-buffered saline (PBS). 
NOTE: The specific concentrations of each component in the anesthesia mixture are: midazolam 13.33&#181;g/100&#181;L, medetomidine 2.5&#181;g/100&#181;L, and butorphanol 16.7&#181;g/100&#181;L. For specific dosages used in mice, the doses are: midazolam 4&#181;g/g, medetomidine 0.75&#181;g/g, and butorphanol 1.67&#181;g/g.&#160;Anesthesia depth was confirmed when the mouse&#39;s limb muscles relaxed, the whiskers had no touch response, and there was loss of&#160; pedal reflex.
Step 3.1.2 of the Protocol was updated from:
After the mice are anesthetized, cut off their whiskers with ophthalmic scissors to prevent blood from flowing down the whiskers and causing hemolysis. Fix the mouse with one hand and press the skin of the eye to make the eyeball protrude. Quickly remove the eyeball and draw 1 mL of blood into the microcentrifuge tube via a capillary tube.
to
After the mice are anesthetized, prepare the peripheral blood samples, by fixing the mouse with one hand and pressing the eye skin to protrude the eyeball. Then, insert the capillary tube into the inner corner of the eye and penetrate at a 30-45 degree angle to the plane of the nostril. Apply pressure while gently rotating the capillary tube. Blood will flow into the tube via capillary action.
Step 3.2.1 of the Protocol was updated from:
Dissect the chest wall to expose the heart, cut open the right atrium, and infuse saline into the left ventricle by an intravenous infusion needle attached to a 20 mL syringe until the tissue turns white.
to
Humanely euthanize the animal according to the institutional policies. Then, dissect the chest wall to expose the heart, cut open the right atrium, and infuse saline into the left ventricle by an intravenous infusion needle attached to a 20 mL syringe until the tissue turns white.

An erratum was issued for:&#160;<a href="https://www.jove.com/t/64609">Generation of a Mouse Spontaneous Autoimmune Thyroiditis Model</a>. The Protocol section was updated.

Erratum: Generation of a Mouse Spontaneous Autoimmune Thyroiditis Model

تم الإبلاغ عن التهاب الغدة الدرقية هاشيموتو (HT) ، المعروف أيضا باسم التهاب الغدة الدرقية الليمفاوي المزمن أو التهاب الغدة الدرقية المناعي الذاتي ، لأول مرة في عام 19121. يتميز HT بتسلل الخلايا اللمفاوية وتلف الأنسجة المسامية الدرقية. تتجلى الاختبارات المعملية بشكل أساسي في زيادة الأجسام المضادة الخاصة بالغدة الدرقية ، بما في ذلك الأجسام المضادة لهرمون الغدة الدرقية (TgAb) والأجسام المضادة للبيروكسيديز المضاد للغدة الدرقية (TPOAb) 2. يتراوح معدل الإصابة ب HT بين 0.4٪ -1.5٪ ، وهو ما يمثل 20٪ -25٪ من جميع أمراض الغدة الدرقية ، وقد زادت هذه القيمة في السنوات الأخيرة3. بالإضافة إلى ذلك ، أفاد عدد كبير من الدراسات أن HT يرتبط بتكوين الأورام وتكرار سرطان الغدة الدرقية الحليمي (PTC)4,5 ؛ الآليات المحتملة لا تزال مثيرة للجدل. التهاب الغدة الدرقية المناعي الذاتي هو أيضا عامل مهم في العقم عند النساء6. لذلك ، يجب أن يكون التسبب في HT واضحا ، حيث يعد النموذج الحيواني المستقر والبسيط ضروريا.
لدراسة مسببات HT ، تم استخدام نوعين رئيسيين من نماذج الفئران ، بما في ذلك التهاب الغدة الدرقية المناعي الذاتي التجريبي (EAT) والتهاب الغدة الدرقية المناعي الذاتي التلقائي (SAT) في الدراسات الحالية 7,8. تم تحصين الفئران الحساسة بمستضدات الغدة الدرقية المحددة (بما في ذلك الغدة الدرقية الخام ، وثيروجلوبولين المنقى [TG] ، وبيروكسيديز الغدة الدرقية [TPO] ، والمجال الخارجي ل TPO المؤتلف ، وببتيدات TPO المختارة) لإنشاء نموذج الفئران EAT. بالإضافة إلى ذلك ، يتم أيضا استخدام المواد المساعدة ، بما في ذلك عديد السكاريد الشحمي (LPS) ، ومساعد فرويند الكامل (CFA) ، وغيرها من المواد المساعدة غير العادية ، أثناء التحصين لتحطيم التسامح المناعي9،10،11،12،13،14،15،16،17.
نموذج SAT هو نموذج مهم لدراسة التطور التلقائي لالتهاب الغدة الدرقية المناعي الذاتي ، والذي يعتمد على NOD. الفئران H-2H4. الإيماءة. الماوس H-2h4 هو سلالة جديدة تم الحصول عليها من صليب NOD و B10. الفئران A (4R) ، تليها عدة تهجينات خلفية إلى NOD ، مع جين الحساسية لالتهاب الغدة الدرقية المناعي الذاتي IAk18,19. موافقه. لا تصاب الفئران H-2h4 بمرض السكري ، ولكن لديها نسبة عالية من التهاب الغدة الدرقية المناعي الذاتي ومتلازمة سجوجرن (SS) 19. وقد وجدت الدراسات أن جزيء الالتصاق داخل الخلايا -1 (ICAM-1) يتم التعبير عنه بشكل كبير في أنسجة الغدة الدرقية من NOD. الفئران H-2h4 في 3-4 أسابيع من العمر. علاوة على ذلك ، مع زيادة تناول اليود ، يتم تعزيز مناعة جزيء ثيروجلوبولين ، مما يزيد من تنظيم التعبير عن ICAM-1 ، الذي يلعب دورا مهما في عملية تسلل الوحيدات21. يحاكي هذا النموذج عملية المناعة الذاتية مع التحقق من العلاقة بين جرعة اليود وشدة المرض. الطريقة المعمول بها مستقرة ، مع احتمال كبير للنجاح. تم تطبيق نموذج SAT للحث على التهاب الغدة الدرقية المناعي الذاتي لسنوات عديدة ولا يزال وسيلة فعالة لدراسة التسبب في التهاب الغدة الدرقية المناعي الذاتي. ومع ذلك ، فإن طريقة البناء الحالية لنموذج EAT أكثر تعقيدا وتكلفة. تستخدم المختبرات المختلفة طرق تحصين ومواقع حقن مختلفة. علاوة على ذلك ، فإن الفئران ذات الخلفيات الجينية المختلفة لديها معدلات مختلفة من الحث ، والتي تحتاج إلى مزيد من الدراسة للكشف عن الآلية القوية.
ومع ذلك ، يرتبط تطور التهاب الغدة الدرقية في نموذج SAT بيوديد الصوديوم ، وإزدواج الشكل الجنسي ، وظروف التربية. للكشف عن الإجراء المناسب لالتهاب الغدة الدرقية المناعي الذاتي في نموذج SAT ، وصفت هذه المقالة طريقة تحريض التهاب الغدة الدرقية المناعي الذاتي في ظروف مختلفة. بالإضافة إلى ذلك ، فإنه يسمح لدراسة التسبب والتقدم المناعي لالتهاب الغدة الدرقية المناعي الذاتي في مراحل مختلفة من هذا المرض.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Butorphanol tartrate</td>
			<td>Supelco</td>
			<td>L-044&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Dexmedetomidine hydrochloride&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>145108-58-3</td>
			<td></td>
		</tr>
		<tr>
			<td>Enzyme-linked immunosorbent assay (ELISA) well</td>
			<td>Sigma-Aldrich</td>
			<td>M9410-1CS</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethanol</td>
			<td>macklin</td>
			<td>64-17-5&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Freund&#8217;s Adjuvant, Complete&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>F5881&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Freund&#8217;s Adjuvant, Incomplete&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>F5506</td>
			<td></td>
		</tr>
		<tr>
			<td>Goat anti-Mouse IgG&#160;</td>
			<td>invitrogen</td>
			<td>SA5-10275&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Midazolam solution&#160;</td>
			<td>Supelco</td>
			<td>M-908&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Mouse/rat thyroxine (T4) ELISA</td>
			<td>Calbiotech</td>
			<td>DKO045</td>
			<td></td>
		</tr>
		<tr>
			<td>Paraformaldehyde</td>
			<td>macklin</td>
			<td>30525-89-4&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Propidium iodide</td>
			<td>Sigma-Aldrich</td>
			<td>P4864</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium Iodine</td>
			<td>Sigma-Aldrich&#160;</td>
			<td>7681-82-5</td>
			<td></td>
		</tr>
		<tr>
			<td>Thyroglobulin</td>
			<td>Sigma-Aldrich&#160;</td>
			<td>T1126</td>
			<td></td>
		</tr>
		<tr>
			<td>Thyroglobulin&#160; ELISA Kit</td>
			<td>Thermo Scientific</td>
			<td>EHTGX5</td>
			<td></td>
		</tr>
		<tr>
			<td>TSH ELISA</td>
			<td>Calbiotech</td>
			<td>DKO200</td>
			<td></td>
		</tr>
		<tr>
			<td>Xylene</td>
			<td>macklin</td>
			<td>1330-20-7</td>
			<td></td>
		</tr>
	</tbody>
</table>

generation of a mouse spontaneous autoimmune thyroiditis model

في السنوات الأخيرة ، أصبح التهاب الغدة الدرقية هاشيموتو (HT) أكثر أمراض الغدة الدرقية المناعية الذاتية شيوعا. يتميز بتسلل الخلايا اللمفاوية والكشف عن الأجسام المضادة الذاتية المحددة في الدم. على الرغم من أن الآلية المحتملة لا تزال غير واضحة ، إلا أن خطر الإصابة بالتهاب الغدة الدرقية هاشيموتو مرتبط بعوامل وراثية وبيئية. في الوقت الحاضر ، هناك عدة أنواع من نماذج التهاب الغدة الدرقية المناعي الذاتي ، بما في ذلك التهاب الغدة الدرقية المناعي الذاتي التجريبي (EAT) والتهاب الغدة الدرقية المناعي الذاتي التلقائي (SAT).
EAT في الفئران هو نموذج شائع ل HT ، والذي يتم تحصينه مع عديد السكاريد الدهني (LPS) جنبا إلى جنب مع ثيروجلوبولين (Tg) أو تستكمل مع مساعد فرويند الكامل (CFA). تم تأسيس نموذج الماوس EAT على نطاق واسع في العديد من أنواع الفئران. ومع ذلك ، من المرجح أن يرتبط تطور المرض باستجابة الأجسام المضادة Tg ، والتي قد تختلف في التجارب المختلفة.
يستخدم SAT أيضا على نطاق واسع في دراسة HT في NOD. ماوس H-2H4. الإيماءة. الماوس H2h4 هو سلالة جديدة تم الحصول عليها من صليب فأر السكري غير البدين (NOD) مع B10. A (4R) ، والذي يتم تحريضه بشكل كبير ل HT مع أو بدون تغذية اليود. أثناء الحث ، NOD. يحتوي فأر H-2h4 على مستوى عال من TgAb مصحوبا بتسلل الخلايا الليمفاوية في الأنسجة المسامية الدرقية. ومع ذلك ، بالنسبة لهذا النوع من نماذج الماوس ، هناك عدد قليل من الدراسات لتقييم شامل للعملية المرضية أثناء تحريض اليود.
تم إنشاء نموذج فأر SAT لأبحاث HT في هذه الدراسة ، ويتم تقييم عملية التغيير المرضي بعد فترة طويلة من تحريض اليود. من خلال هذا النموذج ، يمكن للباحثين فهم التطور المرضي ل HT بشكل أفضل وفحص طرق العلاج الجديدة ل HT.

كانت التغيرات النسيجية مختلفة بشكل لافت للنظر في الإناث والذكور ، ومدة تناول اليود ، وحل NaI. كما هو موضح في الشكل 1 ، ~ 10٪ من NOD. طورت الفئران H-2h4 SAT حتى بدون تحريض اليود في عمر 24 أسبوعا ، وطورت جميع الفئران في النهاية التهاب الغدة الدرقية. عند إعطاء الماء العادي ، لم يكن هناك فرق...

Watch this Scientific Journal Video about Generation of a Mouse Spontaneous Autoimmune Thyroiditis Model at JoVE.com

Generation of a Mouse Spontaneous Autoimmune Thyroiditis Model

تم إنشاء عدة أنواع من النماذج الحيوانية لالتهاب الغدة الدرقية هاشيموتو ، وكذلك التهاب الغدة الدرقية المناعي الذاتي التلقائي في فأر NOD. الفئران H-2h4 هي نموذج بسيط وموثوق لتحريض HT. توضح هذه المقالة هذا النهج وتقيم العملية المرضية لفهم أفضل لنموذج الفئران SAT.

جيل من الفأر نموذج التهاب الغدة الدرقية المناعي الذاتي التلقائي

In recent years, Hashimoto's thyroiditis (HT) has become the most common autoimmune thyroid disease. It is characterized by lymphocyte infiltration and ...

التهاب الغدة الدرقية المناعي الذاتي ، نتيجة المرض الذي يؤثر بشكل خطير على صحة الإنسان. على الرغم من عدم وجود علاج فعال حاليا ، إلا أن النماذج الحيوانية توفر منصة لدراسة هذا المرض. هذا النموذج سهل البناء بخطوات بسيطة ومعدل نجاح مرتفع.إنه يحفز عملية المناعة الذاتية بالتحقق من العلاقة بين موتنا وشدة المرض. للبدء ، قم بإصلاح الماوس على طاولة التشريح باستخدام المسامير. لتحضير عينة أنسجة الغدة الدرقية ، قم بتشريح جدار الصدر وكشف القلب.افتح الأذين الأيمن وحقن المحلول الملحي عن طريق الوريد في البطين الأيسر ، باستخدام حقنة 20 ملليلتر حتى يتحول النسيج إلى اللون الأبيض. تعقيم الرقبة مع 75 ٪ من الإيثانول. باستخدام مقص الأنسجة ، قم بقص جلد الفأر على طول الرقبة ، خط الوسط من أعلى القص إلى الفك السفلي ، لفضح أنسجة الرقبة بالكامل.حدد موقع الغدد تحت الفك السفلي وعضلة القصبة الهوائية الأمامية وافصلها باستخدام مقص العيون والملقط. بمجرد تعرض القصبة الهوائية وغضروف الغدة الدرقية ، قم بإزالة الأنسجة الموجودة أسفل الغضروف باستخدام ملقط منحني. بعد التقاط الغضروف والقصبة الهوائية ، قم بقطع العدسة البعيدة للغضروف والعدسة القريبة من القصبة الهوائية.ثم قم بإزالة الغدة الدرقية والقصبة الهوائية. أظهر تلطيخ HE تغيرات مرضية في الغدة الدرقية ، تحت تناول اليود. طورت جميع الفئران التهاب الغدة الدرقية ، حتى بدون تحريض اليود.ومع ذلك ، لوحظ تطور متسارع لالتهاب الغدة الدرقية مع زيادة تركيز يوديد الصوديوم. تميل إناث الفئران إلى الإصابة بالتهاب الغدة الدرقية أكثر حدة من الذكور تحت نفس الحالة ، لكن الاختلافات في شدة تسلل الخلايا اللمفاوية كانت ضئيلة. لم يتم زيادة مستويات الأجسام المضادة لثيروجلوبولين بشكل ملحوظ في الماء العادي في عمر 16 أسبوعا.في حين أن مكملات يوديد الصوديوم أظهرت ارتفاعا مماثلا في مستويات الأجسام المضادة لدى ذكور وإناث الفئران. تم العثور على الأجسام المضادة لبيروكسيديز الغدة الدرقية في 24 أسبوعا في الماء العادي ؛ وأظهرت الفئران الإناث مستويات أعلى من الذكور في 72 أسبوعا. ومع ذلك ، أظهر إصدار يوديد الصوديوم 0.05٪ أعلى الأجسام المضادة في 64 أسبوعا.بالإضافة إلى ذلك ، كانت مستويات TSH أعلى في ذكور الفئران من إناث NODH2 H4 ؛ مع أو بدون مكملات اليوديد. علاوة على ذلك ، أظهرت مستويات T4 انخفاضا في الميل إلى وقت الحث في ماء اليود. وتجدر الإشارة إلى أن 0.05٪ صوديوم في أنسب تركيز لتقليل نموذج الملح.أيضا ، يجب توخي المزيد من الحذر أثناء عزل العينات. لأن خلية الماوس صغيرة نسبيا ويمكن إتلافها بسهولة. إن إنشاء النموذج الحيواني مفيد لدراسة مسار الإبادة الجماعية ومعالجة حالة خلايا المناعة الذاتية.ويوفر أساسا لتطوير طريقة علاج جديدة.

generation-of-a-mouse-spontaneous-autoimmune-thyroiditis-model

Research

JoVE Journal

Immunology and Infection

Preparation of Mouse Pituitary Immunogen for the Induction of Experimental Autoimmune Hypophysitis

Autoimmune hypophysitis is a chronic inflammation of the pituitary gland caused or accompanied by autoimmunity1. It has traditionally been considered a rare disease but reporting has increased markedly in recent years. Hypophysitis, in fact, develops not uncommonly as a "side effect" in cancer patients treated with antibodies that block inhibitory receptors expressed on T lymphocytes, such as CTLA-42 and PD-1 receptors. Autoimmune hypophysitis can be induced experimentally by injecting mice with pituitary proteins mixed with an adjuvant3. In this video article we demonstrate how to extract proteins from mouse pituitary glands and how to prepare them in a form suitable for inducing autoimmune hypophysitis in SJL mice.

Autoimmune hypophysitis can be reproduced in mice by injecting an extract of mouse pituitary proteins.

إعداد مستمنع ماوس النخامية للتحريض التهاب النخامية الذاتية التجريبي

Autoimmune hypophysitis is a chronic inflammation of the pituitary gland caused or accompanied by autoimmunity1.  It has traditionally been considered a ...

Induction of Experimental Autoimmune Hypophysitis in SJL Mice

Autoimmune hypophysitis can be reproduced experimentally by the injection of pituitary proteins mixed with an adjuvant into susceptible mice1. Mouse models allow us to study how diseases unfold, often providing a good replica of the same processes occurring in humans. For some autoimmune diseases, like type 1A diabetes, there are models (the NOD mouse) that spontaneously develop a disease similar to the human counterpart. For many other autoimmune diseases, however, the model needs to be induced experimentally. A common approach in this regard is to inject the mouse with a dominant antigen derived from the organ being studied. For example, investigators interested in autoimmune thyroiditis inject mice with thyroglobulin2, and those interested in myasthenia gravis inject them with the acetylcholine receptor3. If the autoantigen for a particular autoimmune disease is not known, investigators inject a crude protein extract from the organ targeted by the autoimmune reaction. For autoimmune hypophysitis, the pathogenic autoantigen(s) remain to be identified4, and thus a crude pituitary protein preparation is used. In this video article we demonstrate how to induce experimental autoimmune hypophysitis in SJL mice.

This video shows how to induce autoimmune hypophysitis in SJL mice and how to assess its severity by histopathology.

تحريض التهاب النخامية الذاتية التجريبية في الفئران SJL

Autoimmune hypophysitis can be reproduced experimentally by the injection of pituitary proteins mixed with an adjuvant into susceptible mice1. Mouse ...

Parasite Induced Genetically Driven Autoimmune Chagas Heart Disease in the Chicken Model

The Trypanosoma cruzi acute infections acquired in infancy and childhood seem asymptomatic, but approximately one third of the chronically infected cases show Chagas disease up to three decades or later. Autoimmunity and parasite persistence are competing theories to explain the pathogenesis of Chagas disease 1, 2. To separate roles played by parasite persistence and autoimmunity in Chagas disease we inoculate the T. cruzi in the air chamber of fertilized eggs. The mature chicken immune system is a tight biological barrier against T. cruzi and the infection is eradicated upon development of its immune system by the end of the first week of growth 3. The chicks are parasite-free at hatching, but they retain integrated parasite mitochondrial kinetoplast DNA (kDNA) minicircle within their genome that are transferred to their progeny. Documentation of the kDNA minicircle integration in the chicken genome was obtained by a targeted prime TAIL-PCR, Southern hybridizations, cloning, and sequencing 3, 4. The kDNA minicircle integrations rupture open reading frames for transcription and immune system factors, phosphatase (GTPase), adenylate cyclase and phosphorylases (PKC, NF-Kappa B activator, PI-3K) associated with cell physiology, growth, and differentiation 3, 5-7, and other gene functions. Severe myocarditis due to rejection of target heart fibers by effectors cytotoxic lymphocytes is seen in the kDNA mutated chickens, showing an inflammatory cardiomyopathy similar to that seen in human Chagas disease. Notably, heart failure and skeletal muscle weakness are present in adult chickens with kDNA rupture of the dystrophin gene in chromosome 1 8. Similar genotipic alterations are associated with tissue destruction carried out by effectors CD45+, CD8&gamma;&delta;+, CD8&alpha; lymphocytes. Thus this protozoan infection can induce genetically driven autoimmune disease.

The inoculation of Trypanosoma cruzi in fertile eggs prior to incubation renders the parasite kDNA minicircle integration in embryo cells genome. Crossbreeding reveals the vertical transfer of the mutations to progeny. The kDNA integrates into coding regions at several chromosomes and the chickens die with an inflammatory autoimmune heart disease.

المستحث الطفيلي قيادة وراثيا القلب المناعة الذاتية ومرض شاغاس في نموذج الدجاج

The Trypanosoma cruzi acute infections acquired in infancy and childhood seem asymptomatic, but approximately one third of the chronically infected cases ...

Isolation of Infiltrating Leukocytes from Mouse Skin Using Enzymatic Digest and Gradient Separation

Dissociating murine skin into a single cell suspension is essential for downstream cellular analysis such as the characterization of infiltrating immune cells in rodent models of skin inflammation. Here, we describe a protocol for the digestion of mouse skin in a nutrient-rich solution with collagenase D, followed by separation of hematopoietic cells using a discontinuous density gradient. Cells thus obtained can be used for in vitro studies, in vivo transfer, and other downstream cellular and molecular analyses including flow cytometry. This protocol is an effective and economical alternative to expensive mechanical dissociators, specialized separation columns, and harsher trypsin- and dispase-based digestion methods, which may compromise cellular viability or density of surface proteins relevant for phenotypic characterization or cellular function. As shown here in our representative data, this protocol produced highly viable cells, contained specific immune cell subsets, and had no effect on integrity of common surface marker proteins used in flow cytometric analysis.

This protocol describes enzymatic digestion of mouse skin in nutrient-rich medium followed by gradient separation to isolate leukocytes. Cells thus derived can be used for diverse downstream applications. This is an effective, economical, and improved alternative to tissue dissociation machines and harsher trypsin and dispase-based tissue digestion protocols.

عزل التسلل الكريات البيض من ماوس الجلد عن طريق الأنزيمية دايجست والفصل التدرج

Dissociating murine skin into a single cell suspension is essential for downstream cellular analysis such as the characterization of infiltrating immune ...

Generation and Identification of GM-CSF Derived Alveolar-like Macrophages and Dendritic Cells From Mouse Bone Marrow

Macrophages and dendritic cells (DCs) are innate immune cells found in tissues and lymphoid organs that play a key role in the defense against pathogens. However, they are difficult to isolate in sufficient numbers to study them in detail, therefore, in vitro models have been developed. In vitro cultures of bone marrow-derived macrophages and dendritic cells are well-established and valuable methods for immunological studies. Here, a method for culturing and identifying both DCs and macrophages from a single culture of primary mouse bone marrow cells using the cytokine granulocyte macrophage colony-stimulating factor (GM-CSF) is described. This protocol is based on the established procedure first developed by Lutz et al. in 1999 for bone marrow-derived DCs. The culture is heterogeneous, and MHCII and fluoresceinated hyaluronan (FL-HA) are used to distinguish macrophages from immature and mature DCs. These GM-CSF derived macrophages provide a convenient source of in vitro derived macrophages that closely resemble alveolar macrophages in both phenotype and function.

Bone marrow cells cultured with granulocyte macrophage colony stimulating factor (GM-CSF) generate a heterogeneous culture containing macrophages and dendritic cells (DCs). This method highlights using MHCII and hyaluronan (HA) binding to differentiate macrophages from the DCs in the GM-CSF culture. Macrophages in this culture have many similarities to alveolar macrophages.

جيل وتحديد GM-CSF المشتقة مثل السنخية الضامة والخلايا الجذعية من الفأر نخاع العظم

Macrophages and dendritic cells (DCs) are innate immune cells found in tissues and lymphoid organs that play a key role in the defense against pathogens. ...

Simple and Efficient Production and Purification of Mouse Myelin Oligodendrocyte Glycoprotein for Experimental Autoimmune Encephalomyelitis Studies

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS), thought to occur as a result of autoimmune responses targeting myelin. Experimental autoimmune encephalomyelitis (EAE) is the most common animal model of CNS autoimmune disease, and is typically induced via immunization with short peptides representing immunodominant CD4+ T cell epitopes of myelin proteins. However, B cells recognize unprocessed protein directly, and immunization with short peptide does not activate B cells that recognize the native protein. As recent clinical trials of B cell-depleting therapies in MS have suggested a role for B cells in driving disease in humans, there is an urgent need for animal models that incorporate B cell-recognition of autoantigen. To this end, we have generated a new fusion protein containing the extracellular domain of the mouse version of myelin oligodendrocyte glycoprotein (MOG) as well as N-terminal fusions of a His-tag for purification purposes and the thioredoxin protein to improve solubility (MOGtag). A tobacco etch virus (TEV) protease cleavage site was incorporated to allow the removal of all tag sequences, leaving only the pure MOG1-125 extracellular domain. Here, we describe a simple protocol using only standard laboratory equipment to produce large quantities of pure MOGtag or MOG1-125. This protocol consistently generates over 200 mg of MOGtag protein. Immunization with either MOGtag or MOG1-125 generates an autoimmune response that includes pathogenic B cells that recognize the native mouse MOG.

We describe a simple protocol using only basic lab equipment to generate and purify large quantities of a fusion protein that contains mouse Myelin Oligodendrocyte Glycoprotein. This protein can be used to induce experimental autoimmune encephalomyelitis driven by both T and B cells.

إنتاج بسيطة وفعالة وتنقية ماوس المايلين دبقية قليلة التغصن البروتين السكري لالتجريبية المناعة الذاتية الدراسات التهاب الدماغ

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS), thought to occur as a result of autoimmune responses ...

Cefoperazone-treated Mouse Model of Clinically-relevant Clostridium difficile Strain R20291

Clostridium difficile is an anaerobic, gram-positive, spore-forming enteric pathogen that is associated with increasing morbidity and mortality and consequently poses an urgent threat to public health. Recurrence of a C. difficile infection (CDI) after successful treatment with antibiotics is high, occurring in 20-30% of patients, thus necessitating the discovery of novel therapeutics against this pathogen. Current animal models of CDI result in high mortality rates and thus do not approximate the chronic, insidious disease manifestations seen in humans with CDI. To evaluate therapeutics against C. difficile, a mouse model approximating human disease utilizing a clinically-relevant strain is needed. This protocol outlines the cefoperazone mouse model of CDI using a clinically-relevant and genetically-tractable strain, R20291. Techniques for clinical disease monitoring, C. difficile bacterial enumeration, toxin cytotoxicity, and histopathological changes throughout CDI in a mouse model are detailed in the protocol. Compared to other mouse models of CDI, this model is not uniformly lethal at the dose administered, allowing for the observation of a prolonged clinical course of infection concordant with the human disease. Therefore, this cefoperazone mouse model of CDI proves a valuable experimental platform to assess the effects of novel therapeutics on the amelioration of clinical disease and on the restoration of colonization resistance against C. difficile.

Cefoperazone-treated Mouse Model of Clinically-relevant Clostridium difficile Strain R20291

This protocol outlines the cefoperazone mouse model of Clostridium difficile infection (CDI) using a clinically-relevant and genetically-tractable strain, R20291. Emphasis on clinical disease monitoring, C. difficile bacterial enumeration, toxin cytotoxicity, and histopathological changes throughout CDI in a mouse model are detailed in the protocol.

المعالجة سيفوبيرازون ماوس نموذج سريريا ذات صلة<em&gt; المطثية العسيرة</em&gt; سلالة R20291

Clostridium difficile is an anaerobic, gram-positive, spore-forming enteric pathogen that is associated with increasing morbidity and mortality and ...

A Chronic Autoimmune Dry Eye Rat Model with Increase in Effector Memory T Cells in Eyeball Tissue

Dry eye disease is a very common condition that causes morbidity and healthcare burden and decreases the quality of life. There is a need for a suitable dry eye animal model to test novel therapeutics to treat autoimmune dry eye conditions. This protocol describes a chronic autoimmune dry eye rat model. Lewis rats were immunized with an emulsion containing lacrimal gland extract, ovalbumin, and complete Freund&#39;s adjuvant. A second immunization with the same antigens in incomplete Freund&#39;s adjuvant was administered two weeks later. These immunizations were administered subcutaneously at the base of the tail. To boost the immune response at the ocular surface and lacrimal glands, lacrimal gland extract and ovalbumin were injected into the forniceal subconjunctiva and lacrimal glands 6 weeks after the first immunization. The rats developed dry eye features, including reduced tear production, decreased tear stability, and increased corneal damage. Immune profiling by flow cytometry showed a preponderance of CD3+ effector memory T cells in the eyeball.

This report describes a method to induce chronic experimental autoimmune dry eye in Lewis rats through immunization with an emulsion of rat lacrimal gland extract, ovalbumin, and complete Freund's adjuvant, followed by the injection of lacrimal gland extract and ovalbumin into the forniceal subconjunctiva and lacrimal glands six weeks later.

A المناعة الذاتية الجفاف العين الفئران نموذج مع زيادة في المفعول الذاكرة خلايا T في الأنسجة مقلة العين

Dry eye disease is a very common condition that causes morbidity and healthcare burden and decreases the quality of life. There is a need for a suitable ...

Influenza A Virus Studies in a Mouse Model of Infection

Influenza viruses cause over 500,000 deaths worldwide1 and are associated with an annual cost of 12 - 14 billion USD in the United States alone considering direct medical and hospitalization expenses and work absenteeism2. Animal models are crucial in Influenza A virus (IAV) studies to evaluate viral pathogenesis, host-pathogen interactions, immune responses, and the efficacy of current and/or novel vaccine approaches as well as antivirals. Mice are an advantageous small animal model because their immune system is evolutionarily similar to that found in humans, they are available from commercial vendors as genetically identical subjects, there are multiple strains that can be exploited to evaluate the genetic basis of infections, and they are relatively inexpensive and easy to manipulate. To recapitulate IAV infection in humans via the airways, mice are first anesthetized prior to intranasal inoculation with infectious IAVs under proper biosafety containment. After infection, the pathogenesis of IAVs is determined by monitoring daily the morbidity (body weight loss) and mortality (survival) rate. In addition, viral pathogenesis can also be evaluated by assessing virus replication in the upper (nasal mucosa) or lower (lungs) respiratory tract of infected mice. Humoral responses upon IAV infection can be rapidly evaluated by non-invasive bleeding and secondary antibody detection assays aimed at detecting the presence of total or neutralizing antibodies. Here, we describe the common methods used to infect mice intranasally (i.n) with IAV and evaluate pathogenesis, humoral immune responses and protection efficacy.

Influenza A viruses (IAVs) are important human respiratory pathogens. To understand the pathogenicity of IAVs and to perform preclinical testing of novel vaccine approaches, animal models mimicking human physiology are required. Here, we describe techniques to evaluate IAV pathogenesis, humoral responses and vaccine efficacy using a mouse model of infection.

فيروس الإنفلونزا أ الفيروس دراسات في نموذج ماوس للإصابة

Influenza viruses cause over 500,000 deaths worldwide1 and are associated with an annual cost of 12 - 14 billion USD in the United States alone ...

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.

نموذج التحكم ماوس لأنه الانتان الوليدي

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