Ultrastructural Study of the Outer Cortex, Deep Cortex and Medulla of Mesenteric Lymph Node in Adult Goats

Although goats are disease-resistant livestock, they are subject to some diseases such as pneumonia, foot-rot and internal parasites. Goats can become infected with various viral and bacterial diseases, such as foot-and-mouth disease, caprine arthritis encephalitis, caseous lymphadenitis, pink eye, mastitis and pseudorabies. They also can transmit several zoonotic diseases to people such as tuberculosis, brucellosis, Q-fever and rabies (Smith, 1994).
       
Nagamani et al., (2009) stated that the survival of an animal depends on the successful defense of the body against microbial invasion. As the lymph nodes play a significant role in the disease process and the lesions of lymph nodes are the reflections of lesions of tissues they drain. The high incidence of the mesenteric lymph node lesions observed in many investigations indicated that the spread of infection frequently occurred through the oral route in comparison to the aerosol route in goats. Therefore it is very much essential to understand the normal ultrastructure of mesenteric lymph nodes.

The elongated mesenteric lymph nodes from 6 numbers of 1-3 year old slaughtered goats were collected during 2018 at Veterinary College and Research Institute, Namakkal. The tissue pieces of 5 mm cube was initially fixed in 2.5% glutaraldehyde in 0.1M phosphate buffer at pH 7.4 for 2 days. Then it was washed well with 0.1M phosphate buffer at pH 7.4 followed by post fixation in 1% osmium tetroxide. The specimens were dehydrated in a series of ascending concentrations of acetone. Then the specimens were dried according to the critical point method of Kumaravel et al., (2018). The specimens dehydrated with alcohol and immersed in amyl acetate were put in liquid carbon dioxide in a high pressure chamber. Then the chamber was warmed to 40°C and the high pressure carbon dioxide was slowly let out. Thus critical point dried and freeze fractured specimen was fixed on a metal stub with conductive paste holding the fractured surface upward. In order to obtain sufficient electrical conductivity and yield of secondary electrons, the fractured surface of the specimen was doubly vacuum evaporation coated with carbon and ion sputtered with gold-palladium. The tissues were observed under a VEGA3 TESCAN scanning electron microscope at SASTRA University, Thanjavur with an accelerating voltage of 3 kV at a magnification of 8500x.

The lymph node was made up of capsule, outer cortex, cortical lymph nodules, paracortex, deep cortex, medullary cords, lymph sinuses and blood vessels. For the descriptive purpose, Mori and Lennert (1969) coined the word “Pulp” for the parenchyma, which comprised lymph nodules, medullary cords, lymph sinuses and blood vessels. The parenchyma of the lymph node does not include a capsule and trabeculae.
 
Capsule and Trabeculae
 
The capsule revealed densely piled layers of collagen fibers. The inner surface of the capsule was attached to the spidery reticular cells by their feet traversing the marginal sinus. Trabeculae of variable thickness extended from the capsule into the sinuses. The cut end of the trabeculae appeared like a stump of wood. Trabeculae had numerous branches. The trabecular ramification and the reticular cobweb of the sinus switched over to each other smoothly to form a continuous structure. The sinus lining sheet directly covered the trabeculae. Light microscope and electron microscopic studies of lymph nodes have demonstrated that the inner surface of the capsule and the surfaces of the trabeculae were coated by a basement membrane-like substance and a layer of flattened cells (Sorenson, 1960). The reticular thread in the lymph node parenchyma was thus formed by a bundle of collagen fibrils with occasional elastic fibers covered by a sheath of reticular cell cytoplasm (Clark, 1962).
 
Lymphatic sinuses
 
The lymphatic sinuses formed tunnel-like spaces running beneath the capsule of the node and was also surrounding the cortical nodules and medullary cords. The sinuses were lined by a thin cell sheet and contained a very loose network of spidery reticular cells. In some portions, the sinus appeared like a hollow vessel because of the presence of few reticular cells in that portion.
       
The medullary sinuses contained free macrophages. The lymph sinus was separated from the parenchyma by a sheet of flattened sinusoidal endothelial cells called sinus lining cells (SLC) (Fig 1) between which fenestrations are commonly present. The SLCs were continuous and their cytoplasmic processes (Moe 1964) were connected to the processes of sinus reticular cells (SRC) (Fig 1). The fibrous reticular cells (FRC) (Fig 3) whose fibre-like projections (marked with an arrowhead in Fig 3) were in close spatial relationship with sinus lining cells. The surface of the lining cells was equally rough as that of the reticular cells at higher magnification.
 

 

 
Parenchyma
 
The sponge-like parenchyma was comprised of lymph nodules and medullary cords. The framework of the parenchyma was formed by stellate reticular cells. The bodies of stellate reticular cells were smaller in the parenchyma than those in the sinuses. The meshes were loosely filled with lymphocytes, plasma cells and macrophages. The secondary nodule was a dense, granular area that showed numerous, small round cells and appeared in the cortical portions of the parenchyma. Some smaller cells and large macrophages were found entangled in the meshes formed by the reticular cells. The interdigitating cells form part of the cortical parenchyma (Fig 1 and 2).
 

 
Macrophage
 
The macrophages measured 10 -15 µ in diameter and appeared rough in the surface as they were densely covered by clubbed cytoplasmic processes. Some of them had tentacle-like projections with either clubbed or tapered endings. The macrophages were found to be tightly embraced by the reticular cell processes which appeared like broomstick ramifications. The macrophages in the lymph node, though they usually are round or ovoid, may extend pseudopodial processes during amoeboid movement (Sivagnanam et al., 2017).
       
The medullary sinuses contain free macrophages (Maximow and Bloom, 1957). The macrophages as seen under the SEM appear differently from the reticular cells in their shape and their relation to their neighbours and there are no transitional forms between either. The rounded shape of the macrophages in our specimens may have made it easier to discriminate between the two types of cells. Round macrophages appeared numerously and constantly in the sinus than in the pulp. The electron microscopic study of the rat lymph node indicated that the sinus macrophages had an irregular surface with many small finger-like micropseudopods (Nopajaroonsri et al., 1971). This description was partly in accordance with our observations in goats.
 
Reticular Cells
 
The reticular cells were polygonal or stellate in the form (Fig 2), with their thread-like processes. In most places the cell bodies were flat plates. The neighboring cell plates were arranged in a uniform direction. The perikaryon of the reticular cell was thick. The cell surface appeared smooth when observed under lower magnification but was rough at higher magnification showing irregular elevations and tiny pits.
       
Present observations indicated that the reticular cells, even though phagocytosed small particles such as the carbon particles of Indian ink were specialized for the formation of the skeleton of the lymph node. The reticular cells of the lymph sinus were equivalent to the lymphatic valves. Reticular cells formed the skeleton of the lymph node. The reticular cells in the cortex were smaller than those in the medulla. The post-capillary venules were located in the outer cortex and were not clearly surrounded by perivascular spaces. Interdigitating cells were found both in outer and deep cortices of the mesenteric lymph node (Fig 1 and 2).

The ultrastructure of the mesenteric lymph node revealed that the parenchyma consisted of the outer cortex, cortical lymph nodules, paracortex, deep cortex, medullary cords, lymph sinuses and blood vessels. The capsule and trabeculae formed the supporting framework for the lymph node. The sinuses were lined by a thin cell sheet and contained a very loose network of spidery reticular cells with fenestrations.  The T lymphocytes and B lymphocytes were entangled in the network of reticular and macrophage type of cells. The post-capillary venules were surrounded by perivascular spaces. The interdigitating cells were found only in the cortical parenchyma. The sinus lining cells and fibrous reticular cells were in close spatial relationship.