Histology notes.docx

Histology notes 1 – Cell & Tissue Biology Intro Definitions Angstrom – 1x 10-10 Fixation – prevent autolysis (by enzymes or bact.) & preserve physical structure Light (Bright field) microscope – type of LM, dead specimen Phase contrast microscope – type of LM, live specimen; based on differ...

Histology notes 1 – Cell & Tissue Biology Intro Definitions Angstrom – 1x 10-10 Fixation – prevent autolysis (by enzymes or bact.) & preserve physical structure Light (Bright field) microscope – type of LM, dead specimen Phase contrast microscope – type of LM, live specimen; based on different refractive indices, no stain needed Differential Interference Contrast (DIC) - type of LM, produces 3D image of living cells by polarized light Fluorescent microscope - type of LM, uses fluorescent substances that are irradiated by light to visualize Confocal laser scanning microscope – type of LM, lasers → 3-D image; living specimen Resolving power / resolution – smallest distance b/n 2 particles that are seen as separate objects; LM = 0.1 um; TEM = 0.1 nm; (magnification is independent of resolving power) Transmission electron microscope – cell interior Scanning electron microscope – cell surface; pseudo 3-D Microtome – machine that cuts a specimen into 1-10 um pieces Acidophilia – structures have net positive charge; attracted to acid dyes (eosin) → stain pink; RBC, collagen, mitochondria, lysosomes Basophilia – structures have net negative charge; attracted to basic dyes (hematoxylin) → stain blue; nuclei, DNA/RNA, RER, ECM Hematoxylin & Eosin (H&E) - most common stain, it is an acid-base stain- H- blue (nuc, RER, ECM) and E-pink (RBC, collagen) Periodic acid Schiff (PAS) - carbohydrates (basement membrane, glycoproteins) Silver stain (Ag stain) – reticular fibers (collagen III) Iron hematoxylin (Fe hematoxylin) - elastic fibers What have you learned? What are the 4 fundamental tissues? Epithelial - cover Connective - connect Muscle - contract Nerve - conduct Name some ways to study an embryo, a tissue, a cell. Embryo – incubator; modification to egg Tissue – LM, TEM Cell – differential centrifugation What are the essential steps for making histological sections? Fixation – prevent autolysis & preserve structure Dehydration & embedding – add paraffin or plastic to keep cell from collapsing Sectioning – cut into 1 – 10 um thick by microtome Staining Elastic Fibers - similar to collagen because both are rich in Gly and Pro, but elastic fibers (elastin) contain two unusual amino acids, desmosine and isodesmosine (give elastin its rubber-like properties). Elastin stains uniquely well with Iron-Hematoxylin Reticular Fibers - extremely thin and responsible for forming extensive networks in some organs, composed of collagen type III. Are very argyrophilic (meaning they love silver) Stain with Silver Salts - appear as very fine, black fibers forming a delicate network do NOT stain with H&E 2 – Overview of Cell Ultrastructure Definitions Replication – duplicate DNA; occurs during S phase Transcription – DNA → RNA Translation – RNA → protein Eukaryotic – containing a nucleus Karyotype – organisms complete DNA sequence, staining of metaphase chromosomes (b/c they are found arranged and in their most tightly compact form during metaphase) Plasma membrane (a.k.a. plasmalemma) – has higher protein content than lipid content (60:30) by weight; lipid bilayer with proteins and carbohydrates are associated and in the case of proteins, inserted. In TEM - “railroad track” - alternating bands of electron-dense regions (rich in proteins) and electron-lucent regions (containing fewer proteins and more lipid molecules). Mitochondria – powerhouse; generates ATP, CO2, + H2O Smooth ER – steroid + phospholipid production Rough ER – start of the secretory pathway for proteins, protein secretion Golgi complex – secretory granules (hormones + enzymes) + pigment granules (melanin); protein sorting; carbohydrate hydrolysis, terminal glycosylation, phosphorylation + sulfation, and initial proteolysis Christae – foldings of inner compartment of mitochondria Lysosomes – high content of acid hydrolases that function in the hydrolytic degradation of endocytosed materials play an important role in tooth eruption and movement Peroxisome – catabolize FA via beta-oxidation; contains D-amino acid oxidases and L-amino acid oxidases, + catalase (degrades peroxide to O2 + H2O); Secretory granule – store products until signaled to release via metabolic, hormonal, or neuronal message Mast Cells - secretory granules contain histamine and heparin granules Goblet Cells - secretory granules contain mucus - found in small intestine and respiratory tract Adipocytes - contain lipid droplets unilocular (white) adipocytes - thin ring of cytoplasm surrounding a single, large vacuole filled with lipid droplets, nucleus is displaced to the periphery of the cell. “Signet Ring” cell shape Cytoskeleton – give internal support + scaffolding to cell; facilitate movement + positioning of organelles Microtubules – hollow core; made of tubulin; backbone for centrioles, basal bodies, flagella, + cilia Microfilaments – made up of actin; involved w/ exocytosis, endocytosis, + movement of organelles Intermediate filaments-made up of desmin, vimentin, cytokeratin, or glial fibrillar protein Microvilli – core of 20-30 microfilaments (actin); inserted into terminal web; 1 um long Functions: ↑ Surface area for absorption + secretion Terminal web – network of microfilaments; connects w/ microvilli + inserts into zonula adherens Cilia – mobile; 9 doublets + 2 singlets of microtubules; attached @ basal body; 5-10 um long Centriole – create mitotic spindle made up of microtubules Actin – protein of microfilaments Tubulin – protein of microtubules What have you learned? Life’s Central Dogma =DNA to RNA to protein how to survive and replicate: DNA replication → transcription to RNA → translation to protein Laws of Thermodynamics 1) Energy is always conserved within any process; the energy gained by the system is exactly equal to the energy lost by the surroundings 2) There is always an increase in Entropy in any naturally occurring process Cells must capture energy and utilize it to ensure order within the cell to perform normal processes of growth, reproduction, movement and response to stimuli. An imbalance in the capture and utilization of energy leads to sickness and disease. Discuss cell compartmentalization and how it aids cell function efficiency, protection, and regulation defined by presence of cellular membranes (phospholipid membranes) Name 4 subcellular organelles and one main function of each Nucleus – cell’s largest discrete organelle, site of DNA storage and replication cancer cells often exhibit bizarrely shaped nuclei with deep invaginations and chromatin concentrated around the periphery of the nucleus (marginated chromatin) Mitochondria - power house of cell Rough ER - protein secretion, start of secretory pathway Golgi apparatus - protein sorting; terminal glycosylation, secretory vesicle formation (hormones + enzymes) + pigment granules (melanin), carbohydrate hydrolysis, phosphorylation + sulfation Compare and contrast location, molecular composition and size of microtubules, microfilaments, and intermediate filaments Cytoskeleton Location Molecular composition Size Microtubules Cilia, around outside of cell, mitotic spindle, flagella Tubulin (dynamic and stable) D: 25nm Microfilaments Microvilli Actin (dynamic) D: 5-8nm Intermediate filaments Inside cell spanning cytoplasm, embedded in nuclear membrane many – e.g. keratin (stable) D: 12-15nm Cytoskeleton (“Macrotrabecular Network”) - internal support and scaffolding of the cell, as well as movement and positioning of organelles Microtubules - 25 nm in diameter, hollow core, composed of tublin subunits arranged in a helical profile of 13 subunits “shag carpet” appearance MT are in a constant state of flux polymerization-depolymerization, the MT structure is stabilized by other proteins which are associated with MAP (microtubule associated proteins). MAPs function to1) protect MT from disassembling 2) inhibit tubulin dissociation 3) link adjacent MT and other structures 4) to move items along MT MT form the backbone for other cellular structures: centrioles, basal bodies, flagella and cilia Cilia - 9 doublets +2 singlets = axoneme Basal Body - at the base of each cilium or flagellum, 9 triplets + 0 (nothing in center), MTOC Centriole - MTOC, resides near the nucleus and organizes MT Microvilli - core of microfilaments, 5 nm diameter and are characterized as actin filaments, insert into the terminal web play a role in exocytosis, endocytosis, and the movement of subcellular organelles within the cytoplasm “crew cut” appearance, but in cross section may have glycocalyx creating a “fuzzy” appearance, yielding a “brush boarder” Intermediate Filament create “ghost” nucleus appearance Major Subcelluar Compartments nucleus cytoplasm membrane limited organelles the microtrabecular network (cytoskeleton) cytosol - NOT as organized - contains H2O, enzymes, and other molecules Prokaryotic Cell Structure lack membrane-limited nucleus lack mitochondria and other membrane bound organelles (except ribosomes) have cell wall and different cell membrane BOTH prokaryotes and eukaryotes contain: ribosomes, DNA, RNA and protein