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

Summary

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

Abstract

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.

Protocol

Experimental Protocol

The first step in this experimental protocol is to collect a large number of mouse pituitary glands. The second step is to homogenize the glands to prepare a protein extract. The third step is to emulsify these proteins with an oily mixture.

Step 1: Collection and storage of mouse pituitary glands

The mouse pituitary gland sits at the base of the skull in a depression of the sphenoid bone called the sella turcica, surrounded laterally by the trigeminal nerves and anteriorly by the optic chiasm. The gland is composed of a larger anterior lobe and smaller posterio....

Discussion

Despite the fact that the first patient with autoimmune hypophysitis was reported in 1962⁴, the pathogenic pituitary autoantigens remain to be identified⁵. Animal models can be very useful to identify these autoantigens, aiding the discovery of biomarkers that are translatable to patient care. This article has demonstrated a straightforward procedure for preparing pituitary proteins in a form that is adequate for inducing autoimmune hypophysitis in experimental animals.

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Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
15-ml Falcon tubes
homogenization buffer
phosphate buffered saline
protease inhibitor cocktail
Polytron homogenizer
BCA assay	Pierce, Thermo Scientific	23225
Complete Freund’s Adjuvant (CFA)	Sigma-Aldrich	F-5881
Mycobacterium tuberculosis	BD Biosciences	231141
2.5 mL Hamilton gastight syringe
22-gauge micro-emulsifying needle