Lecture 2

semester

General medicine_2nd

Hyaloplasm and c ytoskeleton Cell organelles - their basic structural characteristics and function Cell inclusions and pigments Cell cycle, cell division, and cel l differentiation

HYALOPLASM AND CYTOSKELETON hyaloplasm (cytosol, cytoplasmic ground substance) - a portion of the cytoplasm surrounding
organelles and inclusions that forms millieau for their functioning; seems to be structureless

consists of: H20

Macromolecules Low

molecular substances (aminoacids, mono- and oligo-saccharides) Ions (K+, Na+, Mg2+, Ca2+), phosphate + chloride anions etc.

Cytoskeleton
a part of cytoplasmic matrix that is responsible for its dynamic properties formed by very fine network extending between nuclear envelope and cell mebrane and that is closely associated to the cell organelles

shape of cells, movement of organelles, movement of entire cells

Components of the cytoskeleton:

intermediate filaments

microtubules

microfilaments
(actin filaments)

Microtubules: diameter 25 nm
are hollow tubes composed of 13 strands of protofilaments that are formed from proteinaceous tubulin subunits (alpha and beta-tubulin) microtubules are bound to other cytoskeletal elements and cytoplasmic organelles

Function of microtubules: they are responsible for organization of the cytoplasm and intracellular transport of organelles and vesicles they help to determine cell shape and polarity they participate in a variety of motile activities (the movement chromosomes during mitosis, the beating of cilia)
disruption or depolymerisation of microtubules or inhibition of their synthesis stop mitotic division, phagocytosis, processes of releasing of secretory granules, result also in the loss of cell symmetry etc.

Microtubules provide the structural basis for cilia and flagella 9 sets of microtubules arranged in doublets that surround two central microtubules = axoneme
centrioles and kinetosomes - 9 sets of microtubules arranged in triplets with microtubules are associated special proteins called motor proteins (take participation in transporting processes in cells with utilization of ATP)

Chief distribution of microtubules: interphase kinetosomes cell in mitosis cilia +

Microfilaments
have diameter only 5-7 nm
(actin filaments)

are composed of actin, a protein involved in muscle contraction each microfilament is formed with hundreds of globular subunits - G-actin - organized into a double-stranded helix with a 36 nm repeat -F- actin microfilaments are very dynamic structures that are continually dissociated and reassembled

distribution of microfilaments: may be attached to the plasma membrane - are involved in defining the surface morphology of the cell - C may penetrate the cytoplasm and be intimately associated with several cell organelles, vesicles or granules - B cytoplamic streaming may support microvilli as terminal web and maintain their shape - A may be organized in constriction ring - D

in muscle cells rhabomyocytes and cardiomyocytes, microfilaments are associated with myosin filaments and form stable structures called as myofibrils

Intermediate filaments have average diameter 10-12 nm are of proteinaceous character and of tissue specific are non-contractile and provide cells with mechanical strength - resistance in the traction and pressure can be visualized with the use of immunocytochemical methods and TEM recently, the microscopic visualization of filaments (their proteins) is used in human pathology for diagnosis of tumours filaments are classified into 5 groups:

INTERMEDIATE FILAMENTS
type thickness protein cell type detection

-------------------------------------------------------------------------------------------------keratin filaments 8-11nm cytokeratin epithelial cells TEM

tonofilaments (about 20 kinds) immun ------------------------------------------------------------------------------------------------------------------------------------

vimentin filaments

TEM, immun

8-11nm

vimetin

mesenchymal cells

(fibroblasts, chondroblasts, endothelial cells, vascular smooth muscle, macrophages) ------------------------------------------------------------------------------------------------------------------------------------

desmin filaments

striated + smooth TEM, immun muscle cells (except vascular smooth muscle) ---------------------------------------------------------------------------------------------------------------glial filaments 8-11nm glial fibrillary astrocytes TEM, immun acidic protein (GFAP) ----------------------------------------------------------------------------------------------------------------

8-11nm

desmin

neurofilaments

8-11 nm

neurofilament triplet (NT) protein

neurons

TEM, impreg

CELL ORGANELLES
are the "little organs" of the cell that posses a distinctive structure and well established function all organelles act in concert and to allow the cell to exist they standardly occur in most cells, in several highly specialized cells organelles may be missing List of organelles: - mitochondria - endoplasmic reticulum - ribosomes - Golgi apparatus - lysosomes - peroxisosomes - centrioles

Mitochondria

are largest and generally most abundant of all cytoplasmic organelles discovered by Altmann in 1890 year their volume can reach to 25 % of the entire one of the cytoplasm except red blood cells and terminal keratinocytes mitochondria occur practically in all cells of the human body

mitochondria are spherical corpuscles or filamentous corpuscles varying from 0.5 to 2 m in diameter and from 5 to 10 m in length function: transform the energy of metabolites obtained by their oxidation into energy bound to ATP molecules the transformation process is known as oxidative (aerobic) fosforylation
the mitochondrion consists of a wall and matrix the wall: an outer mitochondrial membrane smooth, 6 -7 nm an inner mitochondrial membrane thinner, it projects folds called as cristae (in steroid-producing cells, the inner mitochondrial membrane
forms invaginations of tubular or vesicular shape - termed as mitochondria with tubules or mitochondria with tubular cristae)

intermembrane space

the matrix contains enzymes of the tricarboxylic

acid cycle,

ADP, ATP, DNA, RNA, ribosomes

Main differences between mitochondrial membranes: outer mitochondrial membrane - contains integral protein porin that functions as channels they serve for transportation of ions and metabolites from cytosol into the interior of the mitochondrion inner mitochondrial membrane - two kinds of proteins are associated with it electron transport system (cytochromes, coenzyme Q, cytochrome oxidase), globular units with ATPase activity

mitochondria are autonomic organelles contain own DNA and ribosomes that differ from cytoplasmic ones and produce several own proteins new mitochondria originate from preexisting mitochondria by division

Golgi apparatus
organelle is known similar as mitochondria since the end of the 19-century G. a. is ubiquitous cell compartment found usually in close proximity to elements of ER and lysosomes in most cells 3 components - the all are limited by a single and smooth membrane 3 8 flattened sacs termed as cisternae with dilated rims, cisternae are usually curved and parallel vesicles (200 nm) mostly numerous vacuoles - ( 1 or more m)

on thin sections, G. a. may occupy even several separate areas that are termed as Golgi fields

the cis-face is closest to the ER and represents the site at which the material enters the organelle (forming

face)

the trans-face is adjacent to secretion vesicles and storage granules and represents the exit site for most constituents

(maturing face)

Function of G.a.: is engaged in segregation of newly formed or synthesized cellular

constituents and their directing to their final destination in cells

are very small particles, observable only by the electron microscope 15 to 20 nm in diameter chemically, they are defined as complexes of ribonucleic acid (RNA) and proteins in cells they occur ubiquitous and form the catalytic sites for translation of messenger ribonucleic acid into a peptide chain ribosomes are composed of a large and a small subunit, each of which contain specific RNA and protein molecule in cells ribosomes occur in two forms: free in the cytoplasmic matrix (cytoplasmic ground substance or cytosol) attached to outer surfaces of membranes of ER (bound ribosomes) free ribosomes are scattered throughout the hyaloplasm either singly as monosomes or are grouped in spiral or rosette patterns termed as polysomes polysomes originate by joining of individual monosomes along the molecule (strand) of mRNA

Ribosomes

bound ribosomes

Function: free ribosomes /polysomes/ are sites of protein synthesis for the use of the cell

Endoplasmic reticulum
was discovered with introduction of electron microscope into cytology the organelle is identical with the cytoplasmic compartment that stains with basic dyes and is known as the ergastoplasm in the light microscope 2 forms: the rough ER, with ribosomes bound to the outer membrane surface (RER), and smooth ER that is ribosome-free (SER)

Rough endoplasmic reticulum consists of anastomosing flattened sacs - cisternae with polysomes on external surface of limiting membrane

RER is abundant especially in glandular cells and corresponds to basophilic regions of the cytoplasm

glucose-6-phosphatase is used as marker for this compartment

Function of the RER: is a major site of synthesis of proteins that are produced for the use of other cells

is made up convoluted tubules or small vesicles that intercommunicate each other contains no attached ribosomes is usually found more peripherally in the cell

Smooth endoplasmic reticulum

Golgiho apart

it corresponds with cytoplasmic areas that are neutrophilic or sligthly acidophilic the relationship between both forms of ER is very flexible

Function: the site of lipid synthesis, the site of synthesis and break-down of glycogen, is

angaged in formation of membran in the cell, converts toxic molecules to nontoxic derivatives that can be excreted in muscle cells functions as pool of Ca ions

steroidogenic cells abundant SER mitochondria with tubules

are involved in digesting processes in cells firstly have been identified biochemically and then histochemically are spherical bodies of diameter varying between 0,05- 2 m is limited by a single membrane, the content of organelle is termed matrix
it shows very variable electron density and composition lysosomes contain large number of degradative enzymes - more than 60, most of them are active at an acid pH

Lysosomes

3 subgroups of lysosomes: primary secondary residual bodies

Primary lysosomes

are small vesicles with a dense matrix 0.05 to 0.2. m in diameter are located near the trans-face of the G.a contain newly synthesized hydrolytic enzymes that have not been utilised in digestion processes in the cell detection of them needs the use of histochemical methods

Secondary lysosomes are larger than primary ones; the diameter of them ranges between 1 to 2 mm originate as a result of fusion of primary lysosomes with the structures containing the substance to be digested
subdivided on the basis of morphology into: phagolysosomes originate by fusion of primary lysosomes with phagocytic Vacuoles autophagic vacuoles by fusion of primary lysosomes with cellular components such as e.g. mitochondria or RER structural picture of s. l. - variable

Residual bodies (telolysosomes) = are final forms of secondary lysosomes that are enzymic inactive contain material not digested by lysosomal enzymes

Function: degradation not only of intracellular substances and damaged structures but

also exogenous provenience

Peroxisomes (microbodies)
are spherical, membrane-limited organelles, about 0,5 - 1.0 m in diameter
are usually found close to the ER contain more than 40 enzymes that are responsible for a variety of catabolic and anabolic reactions very typical enzyme of this organelle is the catalase destroys hydrogen peroxide urate oxidase, oxidase of D- amino acids in several cells (in liver and kidney), they contain a dense crystalline core termed a nucleoid

Function: take participation in synthetic as well as catabolic processes in cells (synthesis of bile acids, synthesis of phospholipids etc)

Centriole
paired organelle ovoid body (cca 0,2 m wide x 0,2-0,5 m long) in all cells that divide mitotically (except blood and nerve cells) located nearly the nucleus and is surrounded by an area of cytoplasm different from the rest by lesser staining = the centrosome ultrastructural appearance 9 sets of microtubule triplets composed of 3 microtubules of 15 to 20 nm in diameter supporting the periphery pericentriolar structures
(appendages)

triplets are turned counterclockwise to each other at a constant angle

Function of centrioles: mitosis induce the organisation of mitotic spindle ciliogenesis centrioles give rise to kinetosomes, from which cilia originate (kinetosomes show the same structure as centriole)

CELL INCLUSIONS
cytoplasmic deposits

are lifeless structures in ther cytoplasm with temporary occurrence; in most cases they are of a result of the cell activity by accumulation of metabolites or substances with stored function by phagocytosis of material from the cell surrounding or even external environment

stored foods crystals pigments secretion granules Stored foods: proteins, lipids and carbohydrates proteins are found in animal cells rarely they show crystalic organization is supposed to be identical with crystals occurrence: in intestitial Leydig cells, Sertoli sells (both in the testis) lipids - 2 forms: as lipid droplets composed mostly of neutral fats masked or bound lipids that are contained mainy in membranes

variable diameter 100 nm - 10 m

Lipid droplets

they serve as a local store of energy and also as a source of short carbon chains
in ordinary histological sections these are likely appear as round clear areas in the cytoplasm, because the lipid is extracted by solvents used in the preparation of the specimen, after osmium tetroxide fixation,

lipid droplets transit into insoluble and extraction resistant form and appear as black spherical structures in the light microscope

the same appearance lipid droplets show also in electron micrographs the degree of blackening or electron density depends upon the degree of unsaturation of the lipid and the nature of the fixative used

Carbohydrates
are mostly stored in the form of glycogen
is not apparent in routine histological sections but may be selectively stained by the Periodic Acid-Schiffs reaction (PAS) (brilliant magenta).

in electron micrographs, the glycogen shows granular appearance (in dependence of fixative used) it occurs in 2 forms: as dense, roughly izodiametric, 15 to 30 nm particles, referred to as the beta particles or as rosette-like aggregates of larger size called alpha particles

glycogen in TEM

- granules/ cca 20 nm

-granules/ cca 450 - 600 nm

Pigments
are inclusions that possess color and they do not have to be stained by histological dyes 2 groups: exogenous - formed outside the cell and later taken into it, or as endogenous - formed within the cell cytoplasm Exogenous pigments include carotin, lipochromes, dusts (carbon), and minerals such as lead and silver (an artificial introducing of some dyes into the deeper layers of the skin is called tattooing) Endogenous pigments involve: - hemoglobin and its breakdown products (hemosiderin, hematoidin) - melanin and lipofuscin Hemosiderin is a golden-brown, iron containing pigment that is accumulated in the cytoplasm of the phagocytes (for example in the normal spleen, liver, and bone marrow) Hemosiderin shows staining reactions for iron. In electron micrographs the masses of pigment include large numbers of 9 nm dense particles of the iron-containing protein, called the ferritin Hematoidin is iron-free pigment that is produced after longer time, mostly located extracellulary, near to blood vessels, especially in extravation sites Melanin is a brown pigment synthesized by special cells called melanocytes in cells it occur in the form of granules melanosomes on the ultrastructural level, melanosomes appear as a homogenous dense granule separated from cytoplasmic matrix by a membrane unit

Lipofuscin

a tan or light brown pigment that fluoresces a golden-brown in ultraviolet light an amount of lipofuscin is progressively increased with advancing age ("wear and tear" pigment) neurons (cells of the autonomic ganglion) recently lipofuscin is thought to be end product of lysosomal activity and is regarded as the indigestible residues of phagocytosed material or degenerated organelles

THE CELL CYCLE

cell cycle or generation time (individual history of the cell) = time from one mitosis to the
beginning of the next, it occurs in all tissues with cell turnover; characterised by many events in both the cell nucleus and the cytoplasm

cell cycle: interphase mitosis

and

Interphase is divided

G1 - postmitotic S- DNA synthesis G2- premitotic mitosis

(pro-, meta-, ana, telophase)

G1 8 - 25 h, intense synthesis of RNA and proteins, number of organelles increases, the cell volume of daugter cell is restored to normal size S - 8 h, synthesis of DNA and its duplication, after S phase completion chromosomes composed of 2 identical chromatids, centriol is replicated G2- (post DNA duplication)- 3 h, cell continues in growth and cumulates energy, tubulin is synthesized, preparation to mitosis,

2 checking (restriction) points included:

to the end of G1 in the middle G2

when the cell passes the first restriction point, it continues through the S, G2 and mitosis the first restriction point stops the cycle under conditions unfavourable to the cell

in G1 the cell can leave the cycle and enter a quiescent phase - G0 , from this phase it can return to the cycle
neurons and muscle cells stay in G0 for their entire lifetime cells are highly metabolic active but have no proliferative potential or capacity

Mitosis cca 1 h (40-120 min)

Metaphase

Late anaphase
constriction ring from microfilaments is organized in the equatorial plane of the parent cell further narrowing of the ring leads to separation of both daughter cells