Slide 29 (small intestine, H&E)View Virtual Slide
Look at the connective tissue in thesubmucosawhich is the lighter staining areaView Imagebetween the intestinal epithelium and the smooth muscle layer. In this area note the irregular, wavycollagenfibersarranged singly or in small groups. The collagen of fibrous supporting tissues, the dermis of the skin, tendon, ligaments and bone istype I collagen, which providestensilestrength. The background will be clear, reflecting a "ground substance"-rich connective tissue. Look for elongated nuclei, usually solitary, from which a modest amount of tapered cytoplasm extends from either one or both poles of the nucleus. These arefibroblastsView Image(as opposed to clusters of similar appearing elongate nuclei that are usually smooth muscle cells or Schwann cells in a nerve that you will learn to recognize soon). Note that the nucleoplasm of a fibroblast has a generally fine stippled (dot-like) chromatin pattern with occasional coarse chromatin clumps (heterochomatin) and one or two nucleoli. Look for more examples of fibroblasts and note that you rarely see much cytoplasm and that the nuclei of these cells can be quite condensed and hyperchromatic depending upon the plane of section or the metabolic state. You should be able to recognize a range of nuclear morphologies and be able to identify the cells as fibroblasts.Most of the rounded cells you may see in the submucosa are likely white blood cells (monocytes, macrophages, and occasional neutrophils) that have migrated out into the tissue.
Now look at the regionimmediately underneath the intestinal epithelium,which is called thelaminapropriaView Image. This region is also aloose, irregular connective tissuebut can be so extensively infiltrated by white blood cells andplasmacellsthat the supporting fibers and ground substance are obscured.Neutrophilsandmacrophagesare also present and both are discussed below.
A. Plasma Cells
Slide 29 (small intestine, H&E)View Virtual Slide
Slide 40 (trachea, H&E)View Virtual Slide
Look for plasma cells within thelamina propriaofslide29View Image. They are round to oval shaped cells with a distinct cell boundary and a nucleus set to one side. Note the coarse chromatin clumps organzed as radial spokes in the round nuclei which is therefore often described as having a "clockface" or "wagon-wheel" appearance. You should note that the cytoplasm is quite basophilic (i.e. "base loving" so it binds hematoxylin and stains dark blue/purple), and, in well fixed tissue, the cytoplasm in many of the plasma cells is frequently granular (the rough endoplasmic reticulum, really) rather than smooth or even in appearance.
Find theGolgicomplex, a pale or slightly eosinophilic (=eosin "loving", an area rich in membranes containing basic amino acids, syn. = acidophilic) region adjacent to the cell nucleus. The Golgi complex in these particular plasma cells is usually in the form of a fine crescent adjacent to the nucleus and it takes some practice to recognize.
Recall that the primary function of plasma cells is antibody secretion, so they are a prominent constituent of loose connective tissue wherever antigens may enter the body, such as thegastrointestinal, urogenital, and respiratory tracts. Plasma cells may also be foundwithin the connective tissueof many of theglandsthat secrete into these regions. An excellent example of this isslide 40from the trachea (part of the respiratory tract). Look at the areas outlined in theorientation diagram of the tracheaand locate the loose, cellular connective tissue within the glands (the "glands" are coiled tubes of columnar epithelial cells; some the epithelial cells are tall and eosinophilic, whereas others are shorter and more basophilic). In addition to some fibroblasts and a few delicate collagen fibers, you should see quite a few plasma cellsView Imageamongst the epithelial tubes
Slide 40is also a very good specimen to examine the pseudostratified, ciliated columnar epithelium of the trachea. Note also that thebasementmembraneunderlying this particular epithelium is especially prominent.Type IVcollagen, which does not form fibrils, but rather a fine meshwork, is present in all basement membranes. The basal lamina is anchored to the underlying connective tissue by fine fibrils oftype VIIcollagen(you obviously can't tell this looking at it in the light microscope, but you should recall this from lecture).
B. Neutrophils
Slide 29 (small intestine, H&E)View Virtual Slide
Look in the lamina propria amongst the plasma cells and you will find neutrophilsthat have emigrated from the bloodstream into the tissue space as part of the immune response. Neutrophils can be identified by their granular cytoplasm and their multilobular, condensed nuclei. Because of their nuclear morphology, they are frequently also called “polymorphonuclear leukocytes” (aka "PMNs" or “polys”). Neutrophils generally enter tissues in large numbers only in response to a disease stimulus. However, as seen in this slide, it is quite normal to find them in tissues such as the gut where foreign substances frequently invoke an inflammatory response. You will study neutrophils in much greater detail in other sequences and in your histopathology course, but it is useful for now to at least be able to recognize them in various tissues and organs.
C. Macrophages
Slide 26 (lymph node, H&E)View Virtual Slide
With low power, locate themedulla(the interior) of the lymph node. Look for a region characterized by interlacing cords of cells.Macrophagesare the biggest, rounded cells that are floating free in the spaces between the cords of cells. Many of the free cells in these medullary sinuses cannot be identified; however, the large rounded cells, with eccentrically placed, vesicular nuclei are the ones you should try to find.
Many of these macrophages contain phagocytosed red blood cells or the brownish breakdown pigment, hemosiderin (which is the result of lysosomal action on the ingested red blood cells.). Be sure you can identify a macrophage and not just a bunch of cells superimposed upon one another. Macrophages can be seen also in thesubcapsularsinus(the lighter staining area just under the capsule at the periphery of the lymph node).
The "mononuclear phagocyte system" (also called the "reticuloendothelial system" for historic reasons) consists of free and fixed macrophages throughout the body. These cells are important in removing all kinds of debris from the body as well as playing a major role in the immune response.
D. Fat Cells
Slide 152 (pharynx, H&E)View Virtual Slide
Slide 30 (mesentery, H&E)View Virtual Slide
Slide H2 (fetal thorax, H&E)View Virtual Slide(virtual slide courtesy of Western University)
Slide 152is a section from the pharynx. Locate thelarge clearcirclesView Imagein the connective tissue that sits beneath the epithelium. These arecells(oradipocytes). In white or unilocular adipose tissue, lipids are stored as a single, non-membrane bound droplet in these cells. A fatty tissue called brown or multilocular fat, produced during fetal development, has adipocytes that containmultiplefat droplets. Brown fat is important for thermoregulation in newborns and hibernating mammals. In humans, brown fat is widely distributed throughout the body in the first decade of life, but it then disappears except for regions around the kidney, suprarenal glands, aorta, neck and mediastinum. None of our slides of adult tissue shows any brown fat, however this rather unique tissue can be seen inslideH2View Image, which is from a developing fetus.
Look foradiposetissueinSlide 30which is taken from abdominal mesentery (the connective tissue that suspends the viscera within the abdominal cavity). Some of the individual fat cells are often broken during tissue preparation, but the overall impression of what the tissue looks like is the important point.
E. Mast Cells
Slide 160(stomach, PAS & Azure II)View Virtual Slide
Mast cells can only be definitively recognized with special stains such asAzure IIandtoluoidine bluethat identify the heparin storage granules (Azure metachromatically stains the heparin purple). Mast cells are most abundant in the connective tissue associated with the lining of the digestive and respiratory systems, and your collection just so happens to contain a tissue section from the stomach that has been stained with PAS and Azure II. As you look at this section, you will see a very obvious layer of mucous epithelial cells (PAS also reacts with the carbohydrate-rich mucin). However, to see the mast cells, you will need to look deeper in thesubmucosawhere you should find small, ovoid cells amongst the collagen fibers with spherical, eccentric nuclei andintensely basophilic(dark purple to black)granulesView Image. The granules are often so dark that they obscure the nucleus.