LectureLecture 11 GeneralGeneral med_2nd semestermed_2nd semester

Introduction. The object and significance of

histology. Short history of the histology. Brno histological and embryological school. Methods used to study of cells and tissues

Cytology: The cell - definition and general characteristics. Cell membrane, cell surfaces and intercellular junctions. Cell nucleus - its structure and function. Chromosomes

http://www.med.muni.cz/histol/histolc.html

lectures - (Prof. Dr. Dr. S. Čech, DrSc.)

+ practicals

http://www.med.muni.cz/histol/histolc.html

lectures (Prof. Dr. Dr. S. Čech, DrSc.)

+ practicals

Textbooks recommended tostudy:

ISBN: 978-1-4160-3706-4

The Developing Human, 8th Edition - Clinically Oriented Embryology With STUDENT CONSULT Online AccessBy Keith L. Moore, BA, MSc, PhD, FIAC, FRSM and T. V. N. Persaud, MD, PhD, DSc, FRC Path(Lond)

536 pages 1805 illsTrim size 8 1/2 X 10 7/8 in$69.95, Softcover

2008

ISBN: 978-1-4160-3705-7

Before We Are Born, 7th Edition - Essentials

of Embryology and Birth Defects With STUDENT CONSULT Online Access

By Keith L. Moore, BA, MSc, PhD, FIAC, FRSM and T. V. N. Persaud, MD, PhD, DSc, FRC Path(Lond)

368 pages 1308 ills

($54.95, Softcover)

2008

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The object and significance of The object and significance of histologyhistology andand embryologyembryology

Short history of the Short history of the branchbranch

Brno histological and embryological Brno histological and embryological schoolschool

Methods used to study of cells and Methods used to study of cells and tissuestissues

histology and embryology are fundamental branches on medical schools origin of the term of histology

greek histos meaning "tissue„ + logos meaning "the study off" (knowledge or science); the term was primarily used in strict sense of the word as a denotation for study of microscopic parameters of animal and plant cells and tissues

recently, histology = as a branch of science that treats microscopic and submicroscopic structure (organization) of animal or plant bodies

on medical schools - it deals with microscopic and submicroscopic structure of the human body

animal and plant organisms consist of organs, organs of one or more tissues and tissues arecomposed of cells that are considered for elementary units of the living substance

in accordance with 3 mentioned organizational levels – cell, tissue, and organ – histologydivides into 3 sections:

cytology - deals with microscopic and ultrastructural organization of cells

histology proper (or histology in strict sense of this word) - describes light microscopic structure and function of tissues

microscopic anatomy - a section of branch studying microscopic and fine structure of individual

organs

major interdisciplinary branches:

histochemistry, histophysiology, pathological histology, and electron microscopy

Importance of histology:

a basic subject, on which pathology a pathophysiology are built

is used in diagnosis of diseases (department of pathology)

used in farmaceutical industry (for testing drugs and artificial materials used for substitutions of natural organs)

in control of food quality

embryology – its aim and orientation are quite different

the term - of 2 greek words en = in, bryein = to swell

embryology deals with study of individual development of multicellular organisms

individual development /ontogenetic development = ontogeny/ involves

the period from fertilization of the ovum to the death of respective individual

the main reason why histology and embryology are teached together is that all multicellular organisms begin their existence as single cells =generative cells or gametes

model of simultaneous teaching of both disciplines (H + E) is often in countries of middle Europe

Short history of histology

development of histology to a separate branch of science was closely associatedwith 3 preconditions:

the construction of the light microscopethe proclamation of the cell theorythe improving of methods of tissue processing

a) beginning of microscopic examinations dates from the end of sixteenth century

the first simple microscope was constructed by father Hans and his son Zacharias Jansen (1590)R. Hooke and M. Malpighi have employed microscope in studies of structural features of various organisms (the first half of the 17th century)A. Leeuvenhook (between 1673-1716) constructed and used composed microscope and published a series of observation upon protozoa, bacteria, muscle, nerve and other structures (the second half of the 17th century)

a great boom of light microscopy comes in the 19th century when cell nucleus and basic cell organelles (e.g. centrioles, mitochondria, apparatus of Golgi) have been discovered

b) the proclamation of the cell theory - this idea says that the all animal as well as plant organisms consist of cells

in biology the theory was introduced independently by Czech scientist J. E. Purkyně (1837) + two German microscopists M. Schleiden (1838) and T. Schwann (1839)

later, German pathologist R. Virchow (1863) postulated - cells arise only through mitotic division of pre-existing cells

c) to discover and introduce adequate methods of tissue processing for the

light microscopic examinations

- first commercially used microtomes occur during the second half of the 19th century, - to find new fixation and embedding methods,- to finds of methods for visualization of cells and cellular components ( dyes originally prepared for staining of textiles were used and introduced for stainig of cells and tissues)

History of histology and embryology in Brnothe Department of Histology and Embryology was established in 1919

three teachers and scientists achieved international reputation

Prof. MUDr. František Karel Studnička (1870 - 1955)

is founder of department and first head, he became famous

in Europe through the study of connectives tissue and

his exoplasmic theory: the all-intercellular

substances are living and arise as a result of transformation

of cell processes, he prepared and published the first czech

handbook of microscopy (Praktická mikroskopie, 1923)

Prof. MUDr. Jan Florian Prof. MUDr. Jan Florian (1897 (1897 -- 1942)1942)

is pupil of Studnička Florian was outstanding embryologist

in the world literature he has firstly described very young human

embryos aged only 13 to 15 days

his observations are valid to recent time

Prof. MUDr. Karel Mazanec, DrSc. (1922 - 1967)

embryologist and electron microscopist

he introduced TEM - 60 years of the last

century

he explored TEM in study of preimplantation

embryos of mammals

author of book dealing with development of

the human from the zygote to the end of the 3rd week

(published in German language - 1953)

The preparation of tissues and organs for microscopic examination

in general, two ways are used:

- samples (specimens) prepared from living cells and tissues

- fixed samples (specimens) taken from dead organisms

Observation of living cells or tissues

is very valuable as structure and function of cells may be studied simultaneously

is mainly used in unicellular organisms and, occasionally, in free cells of a complex organism,

preparations are not stable

- CELL AND ORGAN CULTURE - cells or fragments of tissue are removed aseptically,

transferred to a physiological medium and kept at a temperature normal for animal from which the sample was taken. The cultures are placed in thin glass vessels or in hanging drops on a coverglas mounted over a hollow slide

- VITAL AND SUPRAVITAL STAINING - in vital staining, dyes are injected into the living animal. The activity of certain cells will result in the selective absorption of the coloring material by these cells. An example of this procedure is the staining by trypan blue of macrophages on the basis of their ability to phagocytose foreign particles. In supravital staining, a dyestuff is added to a medium of cells already removed from the organism. Examples of this technique are the staining of mitochondria in living cells by Janus green, of lysosomes by neutral red and of nerve fibers and cells by methylene blue.

ObservationObservation ofof tissuestissues oror organsorgans fromfrom deaddead organismorganism

is mostly used in histology

it has a great advantage - specimens are permanent and may be stored foryears

tissue processing for microscopic examination involves 6 phases:

sampling (taking or obtaining) of tissue probesfixation of samplesembedding of samplescutting of blocks and affixing of sectionsstaining of sectionsmounting of sections

making of permanent preparations will be demonstrated this week in thefirst practice

CYTOLOGYCYTOLOGYDefinition of cell: an elementary unit of living substance that is capable

basic vital functions (metabolism, autoreproduction (multiplication),

growth, movement , excitability etc.)

minimal unit

prokaryotic cells - with no nucleus and membranous organelles, DNA isnot separated from other cellular components, cells have distinct cell walland hyaloplasm containing plasmids and ribosomes, cell size varies from1- 5 m (bacteria, actinomycets)

eukaryotic cells - form bodies of plants and animals, are larger andshow more complicated organization, contain prominent nucleussurrounded by nuclear envelope, DNA is associated with histones,

eukaryotic (e.) cells utilize membranes as main material for constructionof compartments

e. cells differ in size, shape, number (in multicellularorganisms), and life-span (time)

ShapeShape, , sizesize, , andand lifelife--spanspan ofof cellscells

variable

shape of cells -

environment (free cells

vs tissues) and function

size of cells -

between 10 - 30 m

ovum - 130 - 150 mm

life time of cells -

hours … years (nerve cells)

number of cells –

1013 -1014

Eukaryotic cell as a system

animal cell, isolated or within tissue, shows high level of organization and may be compared toan opened system

it exchanges substances, energy and informations with its enviroment

organization of the system is maintained in the stationary status because all processes in livingcells are directed to a permanent rebuilding and establishing of the balance between them and environment

the middle sized eukaryotic cell consists of molecular units of two kinds:

- macromolecules (of nucleic acids, proteins, lipids, and carbohydrates) - more than 4200 milliards in average

- molecules of inorganic elements (Na, K, Cl, Ca, Mg) + and molecules of water(about 225 billions in average)

molecules associate each other to form

supramolecular complexes - ribosomes, membranes, microtubules

cell organelles - mitochondriae, Golgi apparatus, centriol, endoplasmic reticulum etc.

complex of cell organelles + nucleus + plasmalemma = a cell

main cell components observable with the light microscope have been discovered in the second half of the 19th century

important progress in study of cell components has come with an introduction of electron microscopy in cytology

the all eukaryotic cells consist of:

the nucleus - is the center of cellular activity that plays important role in gene expression,

heredity and cell division. The nucleus consists of several components

the cytoplasm - surrounds the nucleus and is the site of metabolic and synthetic activities of each cell

the cell membrane - plasma membrane, plasmalemma - separates the cell from itsenvironment and maintains its integrity, the membrane is also engaged in creationof the interior cellular environment

the nucleus and cytoplasm differ from each other not only in their physical and chemical properties but also in their structural organization and functionup to date accepted structural organization of both cell compartments in the fixed cell is basedon the use of the electron microscope

Structural components of the fixed animal cell

the nucleus:- nuclear envelope- chromatin- nucleolus- nuclear cytoskeleton(- nuclear inclusions)

the cytoplasm: - cell organelles - are the "little organs" of the cell that posses a distinctive

structure and well established function; they are present in most cells in different number

- mitochondria,- endoplasmic reticulum- ribosomes - Golgi apparatus- lysosomes- peroxisosomes- centrioles

- cell inclusions - are lifeless and have temporary character; in most cases they are of a result of the cell activity

- stored foods (proteins, lipids and carbohydrates), - crystals, - pigments - secretion granules

- hyaloplasm or cytoplasmic ground substance (cytosol) –it is defined as a portion of the cytoplasm that surrounds the cells organelles and inclusionsit seems to be structureless, by electron microscopy and special immunohistochemical methods very fine network within it can be visualized –the cytoskeleton

the cell membrane(plasma membrane, plasmalemma) –

separates cell from the environment and maintains its integrity

Structure of the unit (biological) membrane

membranes are the most frequent supramolecular structuresfound in cells

apart from the plasmalemma, they separate vesicular, tubular nd other cellular configurations

spaces limited by membranes constitute intracellular microcompartments that serve for segregation and concentrationof substrates, products, and other substances in cellular interior

the all membranes in cells display the same characteristictrilaminar structure that has lead to a concept of the unit membrane

the total thickness of the unit membrane is about 7.5 to 10 nm

unit membrane consists of two electron-dense layers (about 2.0 thick) separated by 3.0 nm thick intermediate light layerthe intermediate or central lucent layer is formed of bimolecular layer of lipids with hydrophilic ends of lipids proteins are associated - they form peripheral dense layers - peripheral proteinsintegral or penetrating proteins

the arrangement of lipids and proteins in the unit membrane is very dynamic and is in permanent renewal during the cell life-time

fluid mosaic model of the membrane unit

many intracellular compartments are fabricated of unit membranes (Golgi apparatus, endoplasmic reticulum, wall of the mitochondrion, nuclear envelope etc).

The plasma membrane and its specializations

The plasma membrane (cell membrane) a thin limiting membrane that surrounds cell body against the external environment

plasma membrane is not detectable by the light microscopeas viewed with the electron microscope, the plasma membrane is only 8 to 10 nm thick and shows trilaminar structure of the unit membrane

in many cells, a surface coat composed of protein-polysaccharidescovers the outer membrane surface = glycocalix

functions of the glycocalix:stabilizes the cell membraneparticipates in cell adhesionis responsible for antigenic properties of the cellis also engaged in processes of cell recognizing

Glycocalix visualized using the ruthenium red staining on apices of nonciliated and ciliated cells in the oviduct

Specializations of cell surfaces

in cells, namely polarized epithelial ones, three distinct aspects of the plasma membrane

are distinguished:

- an apical cell surface - it borders a luminal space

- a lateral cell surface - is oriented to the adjacent cell

- a basal cell surface - it is in contact with the basal lamina

ApicalApical cell cell surfacesurface- may be smooth or is provided with microvilli, cilia

(rarely flagellum),

- in addition, it is often involved in processes of cell internalization e.g. cell drinking

or pinocytosis and phagocytosis

Microvilli

finger-like processes, 0.1 m in diameter and 0.1 to 0.5 m in length

projecting from the cell apex either irregularly or regularly and in a great number

numerous microvilli of regular dimensions are found in cells specialized to absorption and correspond with a "brush" or "striated" border of the cell detectable by light microscopy

microvilli greatly increase the surface area of cells and facilitate the absorption of molecules or substances

microvilli „brush border“

Cilia are larger than microvilli and are motile, they reach 1 m in diameter and 5-10 m in length so already well visible by the light microscope

a number of cilia that project from the free apical surface is variable (for instance about 270 cilia are on every ciliated cell in the trachea)cilium is covered with the plasma membrane and has core called the axoneme

the axoneme consists of a sheaf of 9 doublet microtubules enclosing central pair of microtubules

each cilium terminates within the cell at basal body (kinetosome)with cross-striated rootlet

.

cilia beat with frequency 16-20 Hz/min

TEM: SEM:

pseudostratified ciliated columnar epithelium

Flagellum

spermatozoon

Pinocytosis (cell drinking)

is a way of an internalization of colloid fluids surrounding the cellthe process begins by the binding of colloid molecules to the cell membrane,after it the respective membrane parts form small pits or caveolae that then pinch off and give to rise pinocytotic vesiclesvesicles contain internalized colloid molecules and then pass through the cytoplasm to reach the opposite cell aspect where they fuse with the plasma membrane and release their fluid content

exocytosis = release

of content

of pinocytotic vesicles

Phagocytosis

Lateral cell aspectsLateral cell aspectsmay be smooth (rarely) but may also form more or less extensive interdigitationsintercellular space = 10 to 20 nm wide separates plasma membranes of adjacent cells

BasalBasal cell cell aspectaspect

smooth orin cells specialized for transport of ions it is organized in basolateral labyrinth

Intercellular junctions

are local specializations of lateral cell membranes between adjacent cellsin general, they serve to three functions:

increase the cellular attachment - adhering junctionsseal the intercellular space - occluding junctionsserve for cell-to-cell communication - gap or communicating junctions

Adhering junctions: form strong bond between adjacent cells or between basal part of the cell membrane

spot desmosome (macula adherens)

belt desmosome (zonula adherens)

hemidesmosome

proteins as cadherins + desmoplakin andplakoglobin

Spot desmosome (macula adherens) occurs especially in tissues that are

subjected to extreme mechanical stress

about 0.1 um in diameter

intercellular space is 30 nm wide and contains extracellular glycoproteins that

promote adhesion of adjacent cells

on both cytoplasmic sides of the macula adherens, there are 20 nm thick electron dense plaques (containing special proteins - desmoplakins I and II), into which tonofilaments insert

Spot desmosomes

Belt desmosome (zonula adherens)

encircles an epithelial cell completely (zone zonula)

in addition, the intercellular space is about half as wide (15 nm)cytoplasmic plaques are poorly develop

hemidesmosomes = junctions between the plasmalema of basal aspect and laminabasalis spot-like appearance

Occluding junction

= sites where plasma membranes are in such close contact that their integral and peripheral proteins are fused

integral proteins that are shared belong

to family occludins and claudins

are called also tight junctions orzonulae occludentes because they have belt-like structure and encircle epithelial cell completely in a manner similar to that of the belt desmosome

the function of tight junction is to seal

the extracellular space between adjacent epithelial cells

they prevent fluid penetration

they also determine the apical and basolateral domains in cells

Communicating junction or gap junction (nexus)

occur between a variety of excitable and non-excitable cells; serve to passageof ions and electrical impulses in the cardiac and smooth muscle

they usually show form of plaques or spot-like regions (0.5 to 1.0 um in d.) in which membranes of adjacent cells run in close apposition intercellular space is retained and reduced to only 2 to 4 nm

are numerous bridges in extent of each gapjunctionare formed by a special protein, calledconnexin

CELL NUCLEUS AND NUCLEOLUSCELL NUCLEUS AND NUCLEOLUS –– STRUCTURESTRUCTURE

AND FUNCTIONAND FUNCTION

CHROMOSOMESCHROMOSOMES

the nucleus is the center of cellular activity, containing chromosomal

DNA and systems for synthesis and processing that allow the information

in the DNA to be expressed as specific proteins in the cytoplasm

with other words said

THE NUCLEUS PLAYS AN IMPORTANT ROLE IN GENE

EXPRESSION, HEREDITY, AND CELL DIVISION

-- the nucleus of dividing cellthe nucleus of dividing cell-- the nucleus of the nucleus of interphaseinterphase (not dividing) cell(not dividing) cell