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HYALOPLASM AND CYTOSKELETONHYALOPLASM AND CYTOSKELETON

hyaloplasm (cytosol, cytoplasmic ground substance)- a portion of the cytoplasm surroundingorganelles 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 finenetwork extending between nuclear envelope and cell mebrane and that is closely associated tothe cell organelles

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

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Components of the cytoskeleton:

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

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Function of microtubules:

they are responsible for organization of the cytoplasm andintracellular transport of organelles and vesicles

they help to determine cell shape and polaritythey 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 synthesisstop mitotic division, phagocytosis, processes of releasing of secretory granules,result also in the loss of cell symmetry etc.

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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 (takeparticipation in transporting processes in cells with utilization of ATP)

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Microfilaments(actin filaments)

have diameter only 5-7 nm

are composed of actin, a protein involved in

muscle contraction

each microfilament is formed with hundredsof globular subunits - G-actin - organized intoa double-stranded helix with a 36 nm repeat-F- actin

microfilaments are very dynamic structuresthat are continually dissociated andreassembled

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distribution of microfilaments:

may be attached to the plasma membrane - are involved in defining the surfacemorphology of the cell - C

may penetrate the cytoplasm and be intimately associated with several cellorganelles, vesicles or granules - Bcytoplamic streaming

may support microvilli as terminal web and maintain their shape -Amay be organized in constriction ring - D

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

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have average diameter 10-12 nmare of proteinaceous character and of

are non-contractile and provide cells with mechanical strength - resistance inthe traction and pressure

can be visualized with the use of and

recently, the microscopic visualization of filaments (their proteins) is used inhuman pathology for diagnosis of tumours

filaments are classified into 5 groups:

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type thickness protein cell type detection

TEMtonofilaments (about 20 kinds) immun------------------------------------------------------------------------------------------------------------------------------------

TEM, immun

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

TEM, immun

(except vascular smooth muscle)

TEM, immun

TEM, impreg(NT)

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are the "little organs" of the cell that posses a distinctive structure and wellestablished functionall organelles act in concert and to allow the cell to exist

they standardly occur in most cells, in several highly specialized cells organelles may bemissingList of organelles:

are largest and generally most abundant of all cytoplasmic organellesdiscovered by Altmann in 1890 yeartheir volume can reach to 25 % of the entire one of the cytoplasm

except red blood cells and terminal keratinocytes mitochondria occur practically in all cellsof the human body

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mitochondria are spherical corpuscles orfilamentous corpuscles varying from 0.5 to 2m in diameter and from 5 to 10 m inlength

function: transform the energy of metabolitesobtained by their oxidation into energy boundto ATP molecules

the transformation process is known asoxidative (aerobic) fosforylation

the mitochondrion consists of a wall and matrixthe wall:

an outer mitochondrial membranesmooth, 6 -7 nm

an inner mitochondrial membranethinner, it projects folds called as cristae(in steroid-producing cells, the inner mitochondrial membraneforms invaginations of tubular or vesicular shape - termed as

mitochondria with tubules or mitochondria with tubular cristae)intermembrane space

the matrix contains enzymesof the tricarboxylic

acid cycle, ADP, ATP, DNA, RNA, ribosomes

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Main differences between mitochondrial membranes:

outer mitochondrial membrane - contains integral protein porin that functions aschannels

they serve for transportation of ions and metabolites from cytosol into the interior ofthe 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

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mitochondria are autonomic organelles

contain own DNA and ribosomes thatdiffer from cytoplasmic ones andproduce several own proteins

new mitochondria originate frompreexisting mitochondria by division

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Golgi apparatusorganelle is known similar as mitochondria since the end of the 19-centuryG. a. is ubiquitous cell compartment found usually in close proximity to elements of ER andlysosomes 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 curvedand parallel vesicles (200 nm) mostly numerous vacuoles - ( 1 or more m)

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

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

constituents and their directing to their final destination in cells

the cis-face isclosest to the ERand represents thesite at which thematerial enters the

organelle (formingface)

the trans-face isadjacent tosecretion vesiclesand storagegranules andrepresents the exit

site for mostconstituents

(maturing face)

Rib

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Ribosomesare very small particles, observable only by the electron microscope15 to 20 nm in diameterchemically, they are defined as complexes of ribonucleic acid (RNA) and proteinsin cells they occur ubiquitous and form the catalytic sites for translation of messengerribonucleic acid into a peptide chain

ribosomes are composed of a large and a small subunit, each of which contain specific RNA andprotein 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 aregrouped in spiral or rosette patterns termed as polysomespolysomes originate by joining of individual monosomes along the molecule (strand) of mRNA

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

bound ribosomes

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Endoplasmic reticulumwas discovered with introduction of electron microscope into cytology

the organelle is identical with the cytoplasmic compartment that stains with basic dyes and isknown as the ergastoplasm in the light microscope

2 forms: the rough ER, with ribosomes bound to the outer membrane surface (RER), and smoothER that is ribosome-free (SER)

Rough endoplasmic reticulumconsists of anastomosing flattened sacs - cisternae with polysomes on external surface oflimiting membrane

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

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 useof other cells

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Smooth endoplasmic reticulumis made up convoluted tubules or small vesicles that intercommunicate each othercontains no attached ribosomesis usually found more peripherally in the cell

it corresponds with cytoplasmic areas that are neutrophilicor sligthly acidophilic

the relationship between both forms of ER is very flexible

Golgiho apart

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Function: the site of lipid synthesis, the site of synthesis and break-down of glycogen, isangaged in formation of membran in the cell, converts toxic molecules to nontoxic derivativesthat can be excreted

in muscle cells functions as pool of Ca ions

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steroidogenic cells

abundant SER

mitochondria with tubules

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Lysosomesare involved in digesting processes in cellsfirstly 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 matrixit 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

3 subgroups of lysosomes:primarysecondaryresidual bodies

Primary lysosomes

are small vesicles with a dense matrix0.05 to 0.2. m in diameterare located near the trans-face of the G.acontain newly synthesized hydrolytic enzymes that have not been utilised in digestionprocessesin the cell

detection of them needs the use of histochemical methods

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Secondary lysosomes

are larger than primary ones; the diameter of them ranges between 1 to 2 mmoriginate as a result of fusion of primary lysosomes with the structures containingthe substance to be digested

subdivided on the basis of morphologyinto:

phagolysosomes originate by fusionof primary lysosomes with phagocytic

Vacuolesautophagic vacuoles by fusion ofprimary lysosomes with cellularcomponents such as e.g. mitochondriaor RER

structural picture of s. l. - variable

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Residual bodiesResidual bodies

((telolysosomestelolysosomes))= are= are final formfinal formss ofofsecondarysecondarylysosomeslysosomes thatthat areareenzymic inactiveenzymic inactiveccontainontain material notmaterial not

digested bydigested bylysosomal enzymeslysosomal enzymes

Function: degradation not only of intracellular substances and damaged structures butalso exogenous provenience

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Peroxisomes (microbodies)

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Peroxisomes (microbodies)

are spherical, membrane-limited organelles, about 0,5 - 1.0 m in diameterin several cells (in liver and kidney), they contain a dense crystalline core termed a nucleoid

are usually found close to the ERcontain more than 40 enzymesthat are responsible for a varietyof catabolic and anabolicreactions

very typical enzyme of this organelle

is the catalasedestroys hydrogen peroxide

urate oxidase, oxidase of D- aminoacids

Function:take participation in synthetic as well as catabolic processes in cells

(synthesis of bile acids, synthesis of phospholipids etc)

C t i l

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Centriole

paired organelle

ovoid body (cca 0,2 m wide x0,2-0,5 m long)

in all cells that divide mitotically(except blood and nerve cells)

located nearly the nucleus and issurrounded by an area ofcytoplasm different fromthe rest by lesser staining =the centrosome

ultrastructural appearance

9 sets of microtubule tripletscomposed of 3 microtubules of15 to 20 nm in diametersupporting the periphery

pericentriolar structures(appendages)

triplets are turned counterclockwise to each

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triplets are turned counterclockwise to eachother at a constant angle

Function of centrioles:

mitosis induce the organisation ofmitotic spindle

ciliogenesis centrioles give rise to

kinetosomes, from which cilia originate(kinetosomes show the same structure ascentriole)

CELL INCLUSIONSCELL INCLUSIONS

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CELL INCLUSIONSCELL INCLUSIONScytoplasmic deposits

are lifeless structures in ther cytoplasm with temporary occurrence; in most cases they are of aresult of the cell activity

by accumulation of metabolites or substances with stored functionby phagocytosis of material from the cell surrounding or even external environment

stored foodscrystalspigmentssecretion granules

Stored foods:proteins, lipids and carbohydrates

proteins are found in animal cells rarely they show crystalic organizationis supposed to be identical with crystalsoccurrence: in intestitial Leydig cells, Sertoli sells (both in the testis)

lipids - 2 forms: aslipid droplets composed mostly of neutral fatsmasked or bound lipids that are contained mainy in membranes

Li id d l

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Lipid dropletsvariable diameter 100 nm - 10 m

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 thelipid is extracted by solvents used in the preparation of the specimen, after osmium tetroxide fixation,lipid droplets transit intoinsoluble and extractionresistant formand appearas black spherical structures

in the light microscope

the same appearance lipiddroplets show also in electronmicrographs

the degree of blackening orelectron density dependsupon the degree of unsaturationof the lipid and the natureof the fixative used

Carbohydrates

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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 fixativeused)

it occurs in 2 forms: as dense, roughly izodiametric, 15 to 30 nm particles, referred to as thebeta particles or as rosette-like aggregates of larger size called alpha particles

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-granules/ cca 20 nm

-granules/

cca 450 - 600 nm

glycogen in TEM

Pigments

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Pigmentsare 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 leadand silver (an artificial introducing of some dyes into the deeper layers of the skin is calledtattooing)

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 cytoplasmof the phagocytes (for example in the normal spleen, liver, and bone marrow) Hemosiderinshows staining reactions for iron. In electron micrographs the masses of pigment include largenumbers 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 locatedextracellulary, 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

Li f i

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Lipofuscina tan or light brown pigment that fluoresces a golden-brown in ultraviolet lightan 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

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

cell cycle: interphase and

mitosis

Interphase is divided

G1 - postmitotic

S- DNA synthesisG2- 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 sizeS - 8 h, synthesis of DNA and its duplication, after S phase completion chromosomes composedof 2 identical chromatids, centriol is replicatedG

2

- (post DNA duplication)- 3 h, cell continues in growth and cumulates energy, tubulin is

synthesized, preparation to mitosis,

2 checking (restriction) points

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2 checking (restriction) pointsincluded:

to the end of G1in the middle G2

when the cell passes the firstrestriction point, it continues throughthe S, G2 and mitosis

the first restriction point stops thecycle under conditions unfavourable tothe 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 buthave no proliferative potential orcapacity

Mitosis

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Mitosis

cca 1 h (40-120 min)

Metaphase

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Metaphase

Late anaphase

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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

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