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LABORATORY 3 Histology Epithelial Tissue, Connective Tissue and Epithelial Membranes Objectives 1. Identify the following epithelial tissues using a prepared slide or a diagram: simple squamous, simple cuboidal, simple columnar, pseudostratified columnar, stratified squamous, stratified cuboidal, stratified columnar and transitional. Indicate the function and location(s) of the tissue 2. Identify the following connective tissues using a prepared slide or a diagram: Connective tissues: embryonic (mesenchyme), areolar, adipose, reticular, dense regular, dense irregular, hyaline cartilage, elastic cartilage and fibrocartilage. Indicate the function and location(s) of the tissue 3. Given a prepared slide of a mucous membrane, identify it as belonging to the trachea or esophagus. Be able to identify the different epithelium that may be associated with a mucous membrane. Introduction A tissue is composed of groups of cells that are similar in structure, and function together to carry out one or more common or related activities. Body tissues are classified into four basic categories: epithelial tissue, connective tissue, muscle tissue and nervous tissue. The tissues of the body are vastly different with respect to the types of cells present, the arrangement of cells and the amount and type of extracellular material. Epithelial membranes are simple organs that are composed of a superficial layer of epithelial tissue and an underlying layer of vascular connective tissue. The primary epithelial membranes of the body are mucous membranes, serous membranes and the cutaneous membrane (skin). The microscopic study of tissue is called histology. This laboratory exercise will provide an introduction to the microscopic study of epithelial tissues, connective tissues and two epithelial membranes (mucous membranes and serous membranes). This will ensure a better understanding of organ structure and function when the body systems are studied in the future. Muscle tissue and nervous tissue will be the focus of future lab exercises. 37 Materials 1. Each student should have a compound microscope. 2. Each pair of students should have: Lens Paper Immersion Oil Lens Cleaner Activity 1: Resources: Colored Pencils Box of Prepared Slides Examination of Epithelial Tissues Textbook: Photographic Atlas: pages 118-124 pages 19-22 Examine prepared slides of the following epithelial tissues under the microscope and prepare labeled sketches of each one: Simple Squamous Simple Cuboidal Simple Columnar Stratified Squamous Stratified Cuboidal Stratified Columnar Transitional Pseudostratified Tips: 1. Before you begin to search for a particular tissue on a prepared slide, review the photomicrographs in the textbook and in the photographic atlas. 2. Frequently, the prepared slide is a section of an organ that includes multiple types of tissue, including the one that you are interested in studying. It is a good idea to scan the specimen under low power to locate the tissue that you want to examine. For epithelial tissues, you should look at the edge of the specimen, or the lining of tubes, because epithelial tissue often lines spaces. Once the tissue has been identified, move to progressively higher power until you see the tissue in sufficient detail. 3. For most specimens, the cell cytoplasm will appear pale pink or gray and the nucleus will appear dark blue or violet. The plasma membrane may be distinct or indistinct. 4. Note the shape of the nucleus. Squamous cells will have a disc shaped nucleus, cuboidal cells will have a rounded nucleus and for columnar cells, the nucleus is oval shaped. 38 5. For simple epithelium, observe the cell shape and try to locate a basement membrane and other details, such as any goblet cells, microvilli or cilia that might be present. Microvilli and cilia can be seen best using the 100X (oil immersion) objective. For stratified epithelium, observe the shape of the apical cells, the shape of the basal cells, and the number of cell layers. Try to locate the basement membrane as well. You might want to draw the apical cells and the basal cells using different colors. 6. Keep in mind that the cell shape that you visualize is related to the type of section that was made. For example, viewed from the side, simple squamous epithelium looks like a line of thin cells. If, on the other hand, it is viewed as a sheet from above, the tissue looks like a sheet polygonal cells, much like a tile floor. 7. If you are having trouble distinguishing between two tissue types (say, transitional vs. stratified squamous), consider comparing them on two different microscopes placed along side of each other. Look at each one, going back and forth between the two microscopes, until you feel comfortable differentiating them. Try to determine the unique characteristics that make these tissues different. 8. Use this key to help you determine an epithelial tissue classification: Epithelial Tissue Simple Stratified single layer of cells Pseudostratified more than one layer of cells Squamous Cuboidal Columnar flat cells cube shaped cells tall thin cells with nucleus in basal portion single layer of columnar cells but looks multilayered Ciliated Nonciliated Squamous Cuboidal Columnar Transitional flat apical cells; cubical or columnar basal cells apical cells and basal cells are cuboidal (generally 2 layers) columnar shaped apical cells and cuboidal basal cells rounded cube shaped or squamous apical cells and cuboidal or columnar basal cells Keratinized Nonkeratinized 39 Activity 2: Examination of Connective Tissues Resources: Textbook: Photographic Atlas: pages 127-136 pages 22-24 Examine prepared slides of the following connective tissues under the microscope and prepare labeled sketches of each one: embryonic (mesenchyme) reticular hyaline cartilage areolar dense regular elastic cartilage adipose dense irregular fibrocartilage Tips: Most of the above tips still apply, but here a few more: 1. In most cases there is a great deal of nonliving extracellular matrix between the cells of the tissue. The matrix consists of ground substance (interstitial fluid, cell adhesion proteins and proteoglycans) and fibers (collagen, elastic and/or reticular). 2. Stained elastic fibers often appear as dark, thin jagged strands. Stained collagen fibers often appear as thicker hazy pink or light purple, sometimes crisscrossing lines. Reticular fibers are not usually visible unless they are stained with special dyes. If they are visible they may appear as delicate green or black fibers that branch to form a three dimensional network. 3. The chondrocytes of cartilage are located in spaces called lacunae. In stained preparations the chondrocytes are usually pink to violet and appear to have shriveled within their respective lacunae. The collagen fibers of hyaline cartilage are not distinct in the smooth, pinkish or lavender matrix. Elastic fibers are obvious in the matrix of elastic cartilage and collagen fibers are obvious in the matrix of fibrocartilage. 4. Adipose tissue lacks some of the typical connective tissue described above. The adipocytes (cells) store fat in vesicles that are so large the cytoplasm and organelles are pushed into a thin ring just under the plasma membrane. As the cells fill up with fat and enlarge, they crowd the fibers and other cells. Therefore, adipose tissue has little extracellular material compared to other types of connective tissue. 40 5. Use this key to help you to determine a connective tissue classification (note: there are connective tissues such as blood and bone that are not included in this flow chart; they will be examined in other lab sessions): Connective Tissue Mesenchyme Bone embryonic tissue; star shaped cells and abundant fluid matrix Connective Tissue Proper varied cell types; cells not in Cartilage cells called chondrocytes are located in lacunae; fibers may or may not be apparent lacunae; matrix variable in in composition and abundance Loose Dense widely spaced fibers Areolar widely spaced fibers; variety of cell types Hyaline abundant fibers collagen fibers not visible; pink/ purple matrix Reticular closely spaced adipocytes that contain a storage area for fat and a peripheral nucleus elastic fibers visible Fibrocartilage delicate, thin branching fibers; reticular cells Adipose Elastic Regular roughly parallel bundles of collagen and some elastic fibers collagen fibers visible Irregular collagen fibers and some elastic fibers are arranged in an irregular pattern 41 Blood Activity 3: Resources: Examination of Epithelial Membranes Textbook: Photographic Atlas: pages 140-142 trachea: page 134 (figure 14.1) esophagus: page 140 (figures 15.14 and 15.15) Examine prepared slides of the following epithelial membranes under the microscope and prepare labeled sketches of each: mucous membranes: trachea, esophagus Tips: 1. 2. Epithelial membranes are simple organs that contain some type of epithelium resting on a layer of loose connective tissue. The epithelium may be stratified squamous, simple columnar, or pseudostratified columnar with cilia. 42 Checklist: A. Epithelial Tissues: _____ simple squamous epithelium _____ simple cuboidal epithelium _____ simple columnar epithelium _____ pseudostratified columnar epithelium _____ stratified squamous epithelium _____ stratified cuboidal epithelium _____ stratified columnar epithelium _____ transitional epithelium B. Connective Tissues _____ mesenchyme _____ areolar connective tissue _____ adipose tissue _____ reticular tissue _____ dense regular connective tissue _____ dense irregular connective tissue _____ hyaline cartilage _____ elastic cartilage _____ fibrocartilage C. Epithelial Membranes _____ mucous membrane: trachea _____ mucous membrane: esophagus 43 44 Lab 3 worksheet Name: _______________ Score: _______________ 1. Prepare labeled sketches of the following epithelial tissues as seen under the microscope: Simple Squamous Epithelium Simple Cuboidal Epithelium Total magnification: _________ Label a nucleus, cytoplasm and basement membrane (if visible) Total magnification: ________ Label a nucleus, cytoplasm and basement membrane (if visible) Simple Columnar Epithelium Pseudostratified Columnar Epithelium Total Magnification: _________ Label a nucleus, cytoplasm, basement membrane (if visible) and goblet cell (if visible) Total magnification: _________ Label a nucleus, cytoplasm, basement membrane (if visible), goblet cell (if visible) and cilia (if visible) 45 Stratified Squamous Epithelium Stratified Cuboidal Epithelium Total magnification: __________ Label the apical cells, the basal cells and basement membrane (if visible) Total magnification: _________ Label the apical cells, the basal cells and basement membrane (if visible) Stratified Columnar Epithelium Transitional Epithelium Total magnification: __________ Label the apical cells, the basal cells and basement membrane (if visible) Total magnification: __________ Label the apical cells, the basal cells and basement membrane (if visible) 46 2. Prepare labeled sketches of the following epithelial membranes as seen under the microscope: Mucous Membrane: Trachea Mucous Membrane: Esophagus Total magnification: __________ Determine the type of epithelium present and label it; label the lamina propria Total magnification: __________ Determine the type of epithelium present and label it; label the lamina propria 47 3. Prepare labeled sketches of the following connective tissues as seen under the microscope: Mesenchyme Areolar connective tissue Total magnification: __________ Label the cells, matrix Total magnification: _________ Label the collagen fibers, elastic fibers matrix Adipose connective tissue Dense regular connective tissue Total magnification: __________ Label the adipocytes, adipocyte nucleus, and vacuole Total magnification: __________ Label the fibroblast nuclei, collagen fibers 48 Dense irregular connective tissue Reticular connective tissue Total magnification: __________ Label the fibroblast nuclei, collagen fibers Hyaline cartilage Total magnification: _________ Label the reticular fibers Fibrocartilage Total magnification: __________ Label the chondrocytes, lacuna and matrix 49 Total magnification: __________ Label the chondrocytes, lacuna and matrix Elastic cartilage Total magnification: __________ Label the chondrocytes, lacuna, and matrix 50 Post lab worksheet lab 3 1. List the four tissue categories: ___________________________ __________________________ ___________________________ __________________________ 2. Epithelial Tissue – Basic Structure Provide the appropriate term(s): Projections from cells used to propel substances over cell surfaces. __________________________ Term for a single layer of epithelial cells: __________________________ The three common shapes of epithelial cells: __________________________ __________________________ __________________________ The two lamina of the basement membrane that are found between epithelial tissue and underlying connective tissue. __________________________ __________________________ Fingerlike extensions of the plasma membrane that increase surface area for cells involved in absorption or secretion __________________________ An epithelium with many layers. __________________________ Term for a single layer of epithelial cells of varying heights: __________________________ Multilayered epithelium with in which the shape of the apical cells changes due to distension of the organ. __________________________ 51 3. Epithelium – Functions and Locations Match the epithelial tissue with its description. Tissues may be used more than once: a. b. c. d stratified squamous stratified columnar stratified cuboidal transitional e. f. g. h. simple squamous simple columnar simple cuboidal pseudostratified These two tissue types are specialized for secretion and absorption. ________________ ________________ This type of tissue is rare. It can be found in small amounts in the pharynx (throat), lining the male urethra and lining the ducts of some glands. ________________ This tissue permits stretching and is found lining the ureters, urethra and urinary bladder. ________________ This type of tissue protects deeper tissues and lines the mouth, esophagus and vagina (nonkeratinzed) and forms the epidermis of the skin (keratinized). ________________ This rare type of tissue often exists in two layers. It can be found in the ducts of large glands. ________________ This mucus secreting tissue lines ducts of the male reproductive system (non-ciliated) and the trachea and other portions of the upper respiratory tract (ciliated). ________________ This tissue is specialized for filtration (kidney glomeruli) and diffusion (lungs); it lines the heart, blood vessels and lymphatic vessels and secretes fluid when found in a serous membrane. ________________ This tissue makes up portions of kidney tubules, the ducts and secretory portions of small glands and is found on the surface of the ovary. ________________ The nonciliated variety of this tissue lines portions of the digestive tract (from the stomach to the anal canal), the gall bladder and ducts of some glands; the ciliated variety lines small bronchi, uterine tubes and portions of the uterus. ________________ 52 4. Connective Tissue (Basic Structure), Provide the correct term(s): The three types of fibers that can exist in connective tissue. _____________________ _____________________ _____________________ The fiber that has great tensile strength. _____________________ The term for embryonic connective tissue. _____________________ Amorphous substance consisting of interstitial fluid, cell adhesion molecules and proteoglycans that fills spaces between cells. _____________________ Connective tissue that consists primarily of parallel collagen fibers ______________________ Connective tissue cells that are phagocytes. ______________________ Connective tissue that contains all three fiber types and has a gel like matrix. ______________________ Term for cells that secrete cartilage. ______________________ 5. Connective Tissue – Functions and Locations. Match the connective tissue with its description. a. b. c. d. reticular tissue adipose tissue areolar tissue dense irregular tissue e. f. g. h. dense regular tissue elastic cartilage hyaline cartilage fibrocartilage Cartilage that has great tensile strength and is found in intervertebral discs, the symphysis pubis and discs of the knee. ______________________ Strong tissue that is found in tendons, ligaments and aponeuroses. ______________________ Strong cartilage that forms most of an embryo’s skeleton, covers the ends of long bones and is associated with the ribs, nose, trachea and larynx. ______________________ Connective tissue that stores energy, protects against heat loss, and supports organs. ______________________ 53 Connective tissue that forms the internal supporting framework for lymph nodes, the spleen and bone marrow. ______________________ Connective tissue that provides strength even when forces are exerted in many directions; it forms the capsules of joints, the dermis of the skin and the sumacs of the digestive tract. ______________________ This flexible cartilage is used to maintain the shape of structures such as the external ear and the epiglottis of the larynx (voice box). ______________________ This connective tissue is abundant deep to the skin, in the abdomen, in breasts and around the kidneys and eyes. ______________________ This connective tissue is found deep to epithelial tissue; it wraps around organs and capillaries, holds fluid and contains phagocytes which destroy bacteria. ______________________ 6. Epithelial Membranes a. What type of epithelial tissue is found in mucous membranes? __________________________________________________________________ b. Where are mucous membranes found in the body? ____________________________________________________________ c. What type of epithelium is found in serous membranes? _____________________________________________________________ d. Where are serous membranes found in the body? _______________________________________________________________ e. What type of connective tissue is found deep to the epithelium in all epithelial membranes? _______________________________________________________________ f. What is a lamina propria? _______________________________________________________________ 54