**2. Physiology of thymus gland**

*Thymus*

Thymus gland in dog is a compressed bilobed structure located in the cranial mediastinum that is laying cranial to the heart and behind the sternum. Its size is largest in young which is followed by atrophy with the age progression until only a trace remains [3]. When it is fully developed, its caudal part is melded on the cranial surface of pericardium. Divisions of the inferior thyroid, internal thoracic arteries, and superior thyroid artery supply blood to the thymus. These arteries travel along the connective tissue septa, which is extended from the covering capsule into the thymic parenchyma [2]. Histologically, however, it is an old technique, but it is still used excessively in the medical field for the understanding of the organ's microarchitecture [4]. The septa divide the parenchyma into small incomplete microscopic lobules, Where they entered the thymus gland. Veins, inferior thyroid, internal thoracic and left brachiocephalic vein takes blood away from thymus gland. Nerve supply of the thymus gland arises from the sympathetic nerves of the cervical chain and the vagus nerve. Extended divisions of the phrenic nerves stretch up to the covering capsule of the thymus but are not arrive to the gland parenchyma [3]. The function of such enervations to the thymus gland is not well comprehended. Lymphatic vessels drain into the lymph nodes viz. parasternal, tracheobronchial, and brachiocephalic. Histologically, the thymus gland appears as a lobulated lymphoid organ, enclosed with a capsule, made up of a fibrous connective tissue (FCT). Capsule surrounding the organ have blood vessels which supply blood to the thymus gland parenchyma. The CT-composed trabeculae descended downward from the capsule, splits the thymus parenchyma into many incomplete lobules by extending into the interior of the organ [5]. These lobules consist of the following two parts: the cortex is a dark staining outer region just beneath the FCT capsule. It contains densely packed lymphocyte that is not involved in the formation of lymphatic nodules. This portion support the early thymocyte development also positively selects the major self-histocompatibility complex. This portion is very thick at the earlier age. The junctional point between the two compartments is called as corticomedullary junction. This is the specific area where the thymic precursor cells enter in the adult age and few of them differentiate into NK cells and the dendritic cells later few reached to the subcapsular sinuses. This corticomedullary area is also very clear and become blurting and even more fuzzy with the progression of age [6]. The medulla is a light staining inner/central region. Medulla contains later thymocyte differentiation to subpopulation like CD-4 and CD-8, also have fewer lymphocytes than cortex but have more epithelial reticular cells. It also has many thymic (Hassall's) corpuscles which differentiate it from other lymphoid tissues/glands [3]. The Hassall's corpuscles are variable sized ovoid structures composed of granule cells, epithelioid cells, and concentric layer of reticular cells containing keratohyalin and eosinophilic fibers. Under microscope the Hassall's corpuscles and the bubble-shaped adipose tissue appears in the area and their number increases with the age progression [7]. Medulla also shows the continuity between the lobules, because the lobules are incomplete. Thymocytes mature, downregulate, and reach the medullary regions. Cellular components of the thymus glands comprise of emerging thymusderived T cells (later population reached to 95%), the stromal cellular system including the microvasculature, the mesenchymal cells, the dendritic cells, and the very important thymic epithelial cells (TEC) [8]. Few macrophages are present in almost all parts of the gland but in medulla it plays important role in the apoptosis. The TEC are categorized into three key classes including cortical, medullar and subcapsular/ perivascular based on localization in the thymic parenchyma. The dendritic cells are mostly found in the corticomedullary junction and in the medulla. All the aforementioned cells participated in the thymocyte function started from the receiving of the progenitor cells till its final training and maturation. During the period of advance gestation, the thymus in the fetus has unclear cortex and medullary regions,

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Major role of the thymus gland is the training of variety of T cells that respond to the antigens. Its function is mainly regulating by the response of the cytokines and for this the equilibrium among anti-inflammatory and proinflammatory cytokines of the body is crucial. It has been observed that thymic atrophy is associated with age linked with diminished interleukin-7 expression [6]. Thymic epithelial cells are originated from a mutual bipotent ancestor and are also the main constituents in the growth of T cells in the thymic microstructure. It comprised of the two regions including the cortex TECs which is positioned in the cortical regions and the medulla TECs which is in the internal medulla. They experience a sequential progress which is organized by various signals, which later leads to support in physiological maturation and development of the thymocyte. The TECs playing a role in the selection of T- cells in the thymus parenchyma [5]. Both cortical and medullar TECs play distinctive responsibilities in the positive and negative selections of the thymocyte.

## **3. Differentiation, proliferation, and development of lymphocytes**

The undifferentiated lymphocytes are migrated from the reservoir, that is, bone marrow to the thymus gland by means of blood stream. The thymic cortex contains the reticular cells, also known as thymic nurse cells. These cells surround the lymphocytes and enhance the differentiation, proliferation, and maturation of the cells [11]. The lymphocytes get matured and get transformed into immunocompetent cytotoxic T cells, helper T cells, and the T cell. At this stage, the receptor is being attached at surface of the lymphocytes for the recognition of antigens. This process starts just before the birth and continues till some month after the birth. Almost 1% of the mature lymphocytes are getting out of thymus toward the margin on daily basis. The differentiation and further activations of T cells to CD-4 and CD-8, and then established T cells travels from thymus to the marginal blood vessels and secondary immune organs [9, 11]. Thus, the size and mass of thymus reflect the maturation of the immune system.
