ISK 101 - Cytology Short Notes PDF

ISK 101 – Cytology Short Notes in Cytology ISK 101 PROF. Eman Abas Farag Histology department 0 ISK 101 – Cytology INTRODUCTION HISTOLOGY Is the science which study microscopic...

ISK 101 – Cytology Short Notes in Cytology ISK 101 PROF. Eman Abas Farag Histology department 0 ISK 101 – Cytology INTRODUCTION HISTOLOGY Is the science which study microscopical structures of the NORMAL components (cells, tissues, organs) of humans, animals and plants and correlate this structure with their functions. ❖ Cells are very small, so microscopes as light microscope (LM) and electron microscopes (EM) are needed for magnification. ❖ For light microscope (LM), tissues are routinely stained with Hematoxylin & Eosin = H&E 1 ISK 101 – Cytology Hematoxylin=H BASIC & BLUE Stain acidic structures = basophilic as NUCLEI Eosin=E ACIDIC & RED Stain basic structures = Acidophilic CYTOPLASM in some cells SPECIAL STAINS (= stains other than H&E) are sometimes needed for demonstrations of certain structures e.g. Silver (AG), Indian ink, toluidine blue, Sudan III, iron haematoxylin & orcein…….. 2 ISK 101 – Cytology Chapter 1 CYTOLOGY By the end of this chapter, the student should be able to: ▪ Identify and analyse images and drawings of animal cell organelles under a light and electron microscope. ▪ Recognise membranous and non-membranous organelles. ▪ Recognise types of inclusions. ▪ Correlate organelles' structural elements to their respective functions. ▪ Explain relation of certain organelles to some diseases. ▪ Enumerate organelles needed to produce lipids and proteins. ▪ Identify components of the nucleus & types of chromatin. ▪ Recognise heterochromatin and euchromatin. Cell is the structural and functional unit of living tissues. It is formed of nucleus and cytoplasm. Nucleus: Definition: The largest component, present in all cells except RBCs and platelets. Number: One (mononucleated), Two nuclei (binucleated), More than two nuclei (multinucleated). Position: Central, basal, eccentric &peripheral. Shape: Rounded, Oval, Flat,…. LM: Basophilic????? due to its content of DNA & RNA. 3 ISK 101 – Cytology Appears by LM Either: 1-Pale stained with prominent nucleolus= vesicular nucleus (nerve cell, liver)????? = active or 2-Dark stained= condensed nucleus =????? (small lymphocyte) = inactive By EM = Structure of the nucleus: 1. Nuclear membrane or envelop 2. Chromatin. 3. Nucleolus. 4. Nuclear sap= solution fills the spaces between nucleolus& chromatin. 4 ISK 101 – Cytology 1-Nuclear Membrane = Nuclear Envelop LM: basophilic.??? As its attached to Chromatin on its inner side Ribosomes on its outer side EM: Double membranes [outer &inner] Outer nuclear membrane - Rough & granular due to ribosornes - Continuous with rER cisternae. Inner nuclear membrane - Fibrillar due to chromatin - It is associated with nuclear lamina formed mainly of lamins to stabilize nuclear envelope. The inner & outer membranes are separated by perinuclear space. - The inner & outer membranes fuse at intervals forming nuclear pores Which allow Transport of proteins & and ribosomal subunits between nucleus & cytoplasm 5 ISK 101 – Cytology 2-Chromatin: = the genetic material, formed of strands of DNA + proteins. LM: Basophilic???......[pale or deep???] EM: Two forms according to cell activity - Euchromatin: It represents uncoiled=extended chromatin Contains the active genes In active cells as protein forming cells. It appears as electron lucent - Heterochromatin: It represents coiled condensed chromatin Contains the inactive genes. In inactive cells. It appears as electron dense Sites of heterochromatin: 1- Peripheral chromatin: attached to the inner nuclear membrane. 2- Chromatin islands: masses in the nuclear sap. 3- Nucleolus-associated chromatin: around the nucleolus. Functions of Chromatin: 1- Carries genetic information. 2- Control protein formation. 3- Formation of RNA (mRNA, rRNA& tRNA). Q: In a table compare between two types of chromatin 6 ISK 101 – Cytology 3-Nucleolus: LM: round Basophilic mass??? (DNA+RNA). EM: spongy, not limited by membrane, has dark and light areas. Functions Of Nucleolus: Formation of rRNA of ribosomes Medical application: Rapidly growing malignant cells have large nucleoli 7 ISK 101 – Cytology Cytoplasm It is formed of: Cytosol = Solution contains Carbohydrates, +lipid+ptn+enz Organelles: Essential, living, permanent e.g. mitochondria Inclusions. Not…………………. e.g. stored food like fat or pigments like haemoglobin and myoglobin. I- Organelles They are classified according to presence of membranes into: Membranous organelles Non membranous organelles Covered by membrane Not covered by membrane Plasma membrane Ribosomes Mitochondria Cytoskeleton Endoplasmic reticulum Golgi apparatus Lysosomes Peroxisomes 8 ISK 101 – Cytology (A) MEMBRANOUS ORGANELLES: 1- CELL MEMBRANE Definition: A very thin membrane around cells as envelop LM: difficult to be seen by H&E (??) Can be stained by PAS & AG EM: Tri lamellar = unit membrane = Two dark (electron-dense) lines separated by a light one (electron-lucent). It also has a fuzzy layer found on the outer surface only, which represents the cell coat or (glycocalyx). Not required-just to understand EM appearance 9 ISK 101 – Cytology Functions of cell membrane: Transport 1- Simple transport =diffusion with concentration gradient: passive & facilitated by carrier 2- Active = against concentration gradient - need energy 3- Bulk transport = for large molecules: ❖ Exocytosis = Exit of large substance to outside of the cell ❖ Endocytosis = Entry (phagocytosis & pinocytosis & receptor mediated): 10 ISK 101 – Cytology 1- Phagocytosis (Cell eating): Pseudopodia extended from the cell to surround solid particle e.g. bacteria then fuse forming phagosome. As WBCs engulf bacteria. 2. Pinocytosis (cell drinking): Cell membrane engulf and surround extracellular fluid forming pinocytic vesicles. As in thyroid hormone synthesis. 3-Receptor mediated endocytosis: Drugs or hormones bind to specific receptors on outer surface of cell membrane This site is coated from the cytoplasmic side by clathrin protein. Cell membrane. invaginate and form coated vesicles. Therefore, specific drug act only on specific cells. As in uptake of growth hormones. Cell coat: -Cell coat is formed of molecules of glycoproteins & glycolipids. - Present on the outer surface only of cell membrane. Functions of Cell coat: 1. Receptor for drugs, hormones and bacteria 2. Cell recognition 3. Cell immunity 4. Cell protection 5. Cell adhesion 11 ISK 101 – Cytology 2- MITOCHONDRIA Definition: Membranous organelles, responsible for cell respiration and energy production. Number: More in active cell as liver and muscle cells LM: granules or rods With iron Haematoxylin: dark blue with Janus green: green. EM: Oval or rounded membranous organelle. Surrounded with two membranes separated by an inter- membranous space: The outer membrane: smooth, permeable has porins protein The inner membrane: selective, form shelf-like folds called cristae.????For more energy production? Cristae Increase surface area for elementary particles. Elementary particles: = globular structures connected to the cristae with stalk Contain enzymes for ATP production & oxidative phosphorylation 12 ISK 101 – Cytology Mitochondrial Matrix contains: - Oxidative enzymes of Krebs cycle for ATP production. - Ca2+ and Mg2+ act as catalysts for mitochondrial enzymes. - DNA and RNA (divide) (self-replication). Functions: Cell respiration: Obtaining energy from food, most of this energy is stored as ATP and some is liberated as heat to maintain body temperature= power house of the cell Medical application: Defects in mitochondrial enzymes leads to muscular weakness?? Failure in ATP production which is needed for all vital activities as muscle contraction. 13 ISK 101 – Cytology 3-ENDOPLASMIC RETICULUM Definition: Membranous organelle formed of communicating tubules and sacs (cisternae) forming reticulum inside cytoplasm. Types: two types; 1-Rough 2- Smooth Rough ER = rER Site Protein forming cells as: Plasma cell, fibroblast LM Basophilic???Ribosomes [basophilia may be diffuse or localized] EM Parallel tubules & cisternae Regular arrangement. Ribosomes [ electron-dense granules] on outer surface Ribosomal large subunit is attached to rER by specific receptors (Ribophorins). Pores are present under the receptor for passage of proteins to cisternae. 14 ISK 101 – Cytology Functions 1.Protein synthesis by attached ribosomes. 2.Isolation of formed proteins. 3. Initial glycosylation of proteins. 4.Packing of formed proteins as transfer vesicles →Golgi apparatus. 5.Protection of cytoplasm from lysosomes enzymes 15 ISK 101 – Cytology Smooth ER = sER Site Steroid, lipid& carbohydrate forming or storing cells: endocrine glands, liver cells & muscle cells LM Not visible ????. when abundant → acidophilia EM Anastomosing tubules &cisternae Irregular arrangement. No ribosomes. Functions 1. Lipid synthesis 2.Steroid hormones synthesis as cortisone. 3. Glycogen synthesis and store in liver 4. Detoxification of drugs in liver 5. Muscle contraction and relaxation [by Calcium ions pump &store] Q: Compare between rER& sER in a table form 16 ISK 101 – Cytology 4-GOLGI APPARATUS Definition: Membranous organelle, concerned with secretion [secretory apparatus] Site: Well developed in secretory cells like plasma cells. LM: 1) H & E: Not appear But In protein forming cells (as plasma cells) with deeply basophilic cytoplasm, it can be seen as a pale unstained area near the nucleus called “Negative Golgi image”. 2) Silver stain (Ag): brown network of granules & fibrils as in: - Nerve cells [perinuclear] - Secretory cells [supranuclear] EM: (formed of Golgi saccules – transfer vesicles – secretory vesicles) o 3-10 flat curved membranous interconnected saccules o arranged one above the other forming a stack. o Stack has 2 faces: -Immature (convex-entry- Cis) face receives transfer vesicles from rER -Mature (concave-exit- Trans) face: form Secretory vesicles or Lysosomes. Functions: 1. Protein &lipid modifications [by addition of CHO] 2. Protein concentration& packaging 3. Secretory vesicles formation 4. Cell membrane renewal 5. Lysosomes formation 17 ISK 101 – Cytology 5- Lysosomes Definition: Membranous organelles, containing hydrolytic enzymes, considered as digestive apparatus = degrade nutrients, foreign or dead materials. Origin: requires 2 organelles rER: form enzymes →carried to Golgi apparatus as transfer vesicles → then released as primary lysosomes. Site: More in phagocytic cells as macrophages LM: Histochemical stains…. detect enzymes in lysosomes in tissue (like acid phosphatase). EM =2Types: 1. Primary -Newly released from GA. -Not enter in digestive process -Homogenous vesicle 2. Secondary -Formed by fusion of primary lysosome with digestive vacuoles -Appears heterogeneous. Types of 2ry lysosome: 1-Heterolysosomes: 1ry lysosome fuses with phagosomes. 2-Multivesicular bodies: 1ry lysosome fuses with pinocytic vesicle. 3- Autolysosomes: 1ry lysosome fuses with old organelles 4- Residual bodies: contains undigested materials: Either Released by exocytosis or Remain in the cell as lipofuscin pigment as in heart & nerve cells???? 18 ISK 101 – Cytology Functions of lysosomes (DIGESTIVE) 1. Digest nutrients= Digestive 2. Digest bacteria &viruses= Defensive 3. Digest old organelles= maintain cell health 4. Digest dead cell following death 5. Fertilization?? by helping the head of sperm to penetrate ovum. N.B. Characters of protein forming cells: By LM: 1- Pale nucleus with prominent nucleolus 2- Deep basophilic cytoplasm By EM: 1- Euchromatic nucleus 2- Many rough endoplasmic reticulum= rER 3-Many mitochondria 4-Well developed GA Q. Compare in a table between different membranous organelles regarding: definition, site, number, LM, special stains, EM& functions 19 ISK 101 – Cytology (B) NON MEMBRANOUS ORGANELLES: 1-RIBOSOMES Definition: Non-membranous electron dense granules-formed of rRNA and proteins- concerned with protein synthesis Origin: rRNA synthesized in nucleolus + Protein (formed in cytoplasm) and pass-through nuclear pores both unite in the nucleolus into small & large subunits then pass into the cytoplasm through nuclear pores & join during protein formation Number: Abundant in protein forming cells as plasma cells,………. LM: When abundant they cause Cytoplasmic basophilia ?? which may be: Diffuse: As in embryonic cells Localized: As in pancreatic cells Focal (spotty): As in nerve cells (Nissl's granules). EM: Small electron-dense granules. Formed of 2 subunits small & large ones. The 2 subunits unite by mRNA. The large subunit contains a groove in its center which contains the polypeptide chain. Ribosomes are found in 2 forms: 1-Free: Scattered singly in the cytoplasm or Polysomes (polyribosomes) : linked together with mRNA and appear as spiral or rosettes chains 2- Attached: Bind to the outer surface of rER by the large subunits at receptors (ribophorins) Function: Protein synthesis. Free ribosomes: proteins used within the cell for its own use. Attached ribosomes proteins secreted by the cells as hormones& enzymes 20 ISK 101 – Cytology 2- CYTOSKELETON Formed of microtubules, micro filaments & intermediate filaments linked with some proteins to each other and to cell membrane forming a micro trabecular lattice. A. Microtubules [MTs] Structure: Hollow cylindrical tubules Has Constant diameter (24nm) (wall formed of 13 protofilaments),but unfixed length. formed of alpha & beta tubulin protein molecules. Their length varies according to polymerization of tubulin molecules. This is directed by microtubular organizing center (MTOC) which contains gamma tubulin. LM: seen by Immunofluorescent technique Medical application: Functions: Cancer chemotherapy 1. Determine cell shape. 2. Help intracellular transport of organelles, vesicles stop tumor cells proliferation 3. Formation of mitotic spindle during cell by preventing MTs & mitotic division. spindle formation 4. Formation of centrioles, cilia and flagella. 21  ISK 101 – Cytology 1- Centrioles Definition: Cylindrical structures formed of MTs. LM: by iron hematoxylin: Appear as two dark bodies near the nucleus. EM: In non- dividing cells: 2 cylinders at right angles to each other surrounded by a matrix of tubulin (= centrosome). Each cylinder is formed is formed of 27 microtubules arranged as 9 bundles, each bundle is formed of 3 MT (triplets)[9 x 3 =27]. Each triplets is named as A,B&C from inner to outer Functions of Centrioles: 1- Formation of mitotic spindle during cell division [after duplication] 2- Share in the formation of cilia and flagella. 2- Cilia Definition: Motile hair-like processes with core formed of microtubules & covered by cell membrane. Origin: Centrioles duplicate thousands of times to form basal bodies of cilia basal bodies migrate under apical cell membrane. Shaft grows up from each basal body& rootlet grows down into cytoplasm. LM: hair-like striations. EM: a cilium is formed of 3 parts: a basal body, shaft & rootlets. 22 ISK 101 – Cytology 1- Basal body: Structure: single centriole = 27 microtubules in 9 triplets[A,B&C]. It is embedded in the cytoplasm. 2- Shaft (axoneme): Finger-like projection from the surface of the cell covered by cell membrane. Structure: 9 peripheral doublets MTS [A&B] + 2 singlet MTS [newly formed] in the center = 20 MTs. 3-Rootlets: Structure: growth of the microtubule (C) of each triplet into the cytoplasm (9 MT). Function: Fix the basal body and shaft to the cytoplasm. Functions of Cilia: Move secretions or particles in one direction e.g.in the respiratory system & female genital system. Cilia Modify to act as receptors e.g. rods & cones of the retina. 3- Flagella Definition: Motile projections from the cell to move the cell itself. Structure: same as the axoneme of the cilium (9 peripheral doublets+ 2 central singlets)= 20 MTs. But extremely longer than cilia. Function: forms the tail of the sperm to helps its movements. Medical application: Cancer chemotherapy is used to arrest cell proliferation in tumors by preventing mitotic spindle formation. Inability of the cilia to move results in chronic respiratory infections. Immotile flagella cause male infertility. 23 ISK 101 – Cytology B-Filaments (1) Microfilaments (Actin filaments) 5:7nm Structure: dynamic strands of 2 chains of globular G actin, coiled around each other to form filamentous F actin. LM: Immunofluorescent technique Sites &Functions: 1. Form a network under the cell membrane involved in cell shape changes as endocytosis, exocytosis 2. Intracellular movement of organelles & vesicles 3. Interact with myosin for cleavage [separation] of dividing cells. 4. Interact with myosin for muscle contraction. 5. Formation of core of microvilli and stereocilia. Structures Formed by Microfilaments 1-Microvilli 2-Stereocilia: Non motile solid cilia Finger like projections (shorter than cilia) from cell Not true cilia but long microvilli. membrane of some cells. LM Apical straited brush border. Hair like processes from free surface of some cells. EM A core of actin filaments Their core has actin filaments (to maintain its shape), (no microtubules). covered by cell membrane Function Increase surface area for Help absorption in male genital system e.g. more absorption e.g. small epididymis intestine. 24 ISK 101 – Cytology (2) Intermediate filaments 8:10nm Intermediate in size between microfilaments and microtubules Structure: Formed of rod shaped subunits LM: seen by Immunofluorescent technique Sites &Types 1. keratin: in epithelial cells, hairs, nails 2. Vimentin: in CT & muscle. 3. Desmin: in muscle 4. Neurofilaments: in neurons. 5. Glial fibrillar acidic protein [GFAP]: in glial cells 6. Lamins: in nuclear envelope Function: all are (supportive).filaments (myosin) 15 nm Medical application: Identification of intermediate filaments protein is important in diagnosis & treatment of tumors. The cell origin of the tumor can be recognized. Q. In table form, compare between cytoskeleton components regarding: structure, site, LM, special stains, functions & medical importance 25 ISK 101 – Cytology II- Cytoplasmic Inclusions Definition: Non-living, temporary, nonessential [except myoglobin & haemoglobin] cytoplasmic contents. They include stored food and pigments A- Stored Food: 1- Carbohydrates Site: Stored as glycogen granules in liver and muscle cells. L.M: H & E → dissolve appear as vacuoles. Best's carmine stain → RED granules. PAS → MAGENTA RED granules. 2 2- Fats Site: In fat cells and in liver cells. L.M: H & E: vacuolated as it dissolves during preparation [in fat cells, it has signet ring appearance] Sudan Ill stain: ORANGE globules. B- Pigment: Definition: Coloured particles, having their own colour without staining, produced by the cell or taken from outside. Types: Endogenous: produced by the cell: Exogenous: taken from outside: 1) Hemoglobin (Hb): in RBCs, carries 1) Carbon and Dust particles: in gases & Myoglobin in muscles lung. 2) Melanin: in skin & eye, give color 2) Carotene pigments: in carrots. &protects from UV rays. 3) Tattoo marks: dyes are injected 3) Lipofuscin: under the skin & taken by phagocytic in nerve &cardiac cells. cells. = waste products increase with age. Q: Correlate lipofuscin pigment to types of lysosomes 26