LAB EVALUATION OF CELL DISORDERS

1- Cell structure

Ola H. Elgaddar

MBChB, MSc, MD, CPHQ, LGBSS Lecturer of Chemical Pathology

Medical Research Institute Alexandria University

Ola.elgaddar@alexu.edu.eg

Sources:

1. Grey's anatomy, 39th ed, 2008

2. Ganong physiology, 23rd ed,2010

3. Robbins pathology, 7th ed, 2005

4. Kumar clinical medicine,6th ed,2005

5. Molecular cell biology, 5th ed, 2003

-The cell is the basic unit of life.

-Microorganisms such as bacteria, yeast, and amoebae exist as single cells.

-By contrast, the adult human is made up of about 30 trillion cells (1 trillion = 1012) which are mostly organized into collectives called tissues.

TYPES OF CELLS

1) Prokaryotic:

- Bacterial cells are said to be prokaryotic (Greek for “before nucleus”)

- Does not have a complex metabolic structure such as nucleus and mitochondria.

- They have very little visible internal organization so that, for instance, the genetic material is free within the cell.

- They are also small, the vast majority being 1–2 μm in length.

2) Eukaryotic:

- The cells of all other organisms (mammals, fungi & plants) are eukaryotic (Greek for “with a nucleus”).

- These are generally larger (5–100μm, although some eukaryotic cells are large enough to be seen with the naked eye.

- Structurally, they are more complex; contain a variety of specialized structures known collectively as organelles, surrounded by a viscous substance called cytosol.

CELL MEMBRANE

-It is made up of lipids and proteins and is semi permeable, allowing some substances to pass through it and excluding others.

-However, its permeability can also be varied because it contains numerous regulated ion channels and other transport proteins that can change the amounts of substances moving across it.

-The major lipids forming plasma membranes are phospholipids such as phosphatidylcholine and phosphatidylethanolamine.

- The shape of the phospholipid molecule reflects its solubility properties:

• The head of the molecule contains the phosphate portion and is relatively soluble in water (polar, hydrophilic)

• The tails are relatively insoluble (nonpolar, hydrophobic).

[Mosaic model of cell membrane,

lipid bilayer]

1-Cell structure.pptx

-The possession of both hydrophilic and hydrophobic properties make the lipid an amphipathic molecule.

-In the membrane, the hydrophilic ends of the molecules are exposed to the aqueous environment that bathes the exterior of the cells and the aqueous cytoplasm; the hydrophobic ends meet in the water-poor interior of the membrane.

-Many different proteins are embedded in the membrane. They exist as separate globular units and many pass through the membrane (integral proteins), whereas others (peripheral proteins) stud the inside and outside of the membrane.

-The amount of protein varies significantly with the function of the membrane but makes up on average 50% of the mass of the membrane.

The proteins in the membranes carry out many functions:

• Cell adhesion molecules that anchor cells to their neighbors or to basal laminas.

• Pumps, actively transporting ions across the membrane.

• Carriers, transporting substances down electrochemical gradients by facilitated diffusion.

• Ion channels, which, when activated, permit the passage of ions into or out of the cell.

• Receptors that bind ligands, initiating physiologic changes inside the cell.

• Enzymes, catalyzing reactions at the surfaces of the membrane.

The cell coat (glycocalyx)

- The plasma membrane differs structurally from internal membranes in that it possesses an external, diffuse, carbohydrate-rich coat, the cell coat or glycocalyx.

- The cell coat forms an integral part of the plasma membrane, projecting as a diffusely filamentous layer 2-20 nm or more from the lipoprotein surface.

-The precise composition of the glycocalyx varies with cell type: many tissue and cell type-specific antigens are located in the coat, including :

•major histocompatibility antigen systems

•blood group antigens (in the case of RBCs)

•adhesion molecules

- Cells tend to repel each other because of the predominance of negatively charged carbohydrates at cell surfaces. There is consequently a distance of at least 20 nm between the plasma membranes of adjacent cells, other than at specialized junctions.

CYTOPLASM

- This is the fluid component inside the cell membrane and contains many specialized organelles

- It contains a scaffolding or cytoskeleton that regulates the passage and direction in which the interior solutes and storage granules flow.

ENDOPLASMIC RETICULUM

-Endoplasmic reticulum is a system of interconnecting membrane-lined channels within the cytoplasm.

-These channels take various forms, including cisternae (flattened sacs), tubules and vesicles.

-The phospholipid membrane of ER is continuous with the perinuclear space.

Rough endoplasmic reticulum

-The rough endoplasmic reticulum, studded with ribosomes, is a site of protein synthesis.

- Most proteins pass through its membranes and accumulate within its cisternae, although some integral membrane proteins, e.g. plasma membrane receptors, are inserted into the rough endoplasmic reticulum membrane, where they remain.

- After passage from the rER, proteins remain in membrane-bound cytoplasmic organelles such as lysosomes, become incorporated into new plasma membrane, or are secreted by the cell.

- Some carbohydrates are also synthesized by enzymes within the cavities of the rough endoplasmic reticulum and may be attached to newly formed protein (glycosylation).

- Vesicles are budded off from the rough endoplasmic reticulum for transport to the Golgi as part of the protein-targeting mechanism of the cell.

Smooth endoplasmic reticulum - The smooth endoplasmic reticulum is associated with carbohydrate metabolism, detoxification and synthesis of lipids, cholesterol and other steroids.

- The membranes of the sER serve as surfaces for the attachment of many enzyme systems, e.g. the enzyme cytochrome P450, which is involved in important detoxification mechanisms and is thus accessible to its substrates, which are generally lipophilic.

- They also cooperate with the rER and the Golgi apparatus to synthesize new membranes; the protein, carbohydrate and lipid components are added in different structural compartments.

- Highly specialized types of ER are present in some cells. For example, in skeletal muscle cells, the sER (sarcoplasmic reticulum) stores calcium ions, which are released into the cytosol to initiate contraction after stimulation initiated by a motor neuron at the neuromuscular junction.

RIBOSOMES

-Ribosomes are macromolecules that catalyze the synthesis of proteins from amino-acids.

- They are granules 15 nm in diameter, composed of ribosomal RNA (rRNA) molecules assembled into two unequal subunits

-The subunits can be separated by their sedimentation coefficients (S) in an ultracentrifuge, into larger 60S and smaller 40S components.

-These are associated with 73 different proteins (40 in the large subunit and 33 in the small), which have structural and enzymatic functions.

-Three small rRNA strands (28S, 5.8S and 5S) make up the large subunit, and one strand (18S) is in the small subunit.

Title

-A typical cell contains millions of ribosomes.

- They may be solitary, relatively inactive structures, or may form groups (polyribosomes or polysomes), attached to messenger RNA (mRNA), which they translate during protein synthesis.

- In a mature polysome, all the attachment sites of the mRNA are occupied as ribosomes move along it, synthesizing protein according to its nucleic acid sequence. Consequently, the number of ribosomes in a polysome indicates the length of the mRNA molecule and hence the size of the protein being made.

../DATA/Translation.flv

-The two subunits have separate roles in protein synthesis.

-The 40S subunit is the site of attachment and translation of mRNA.

-The 60S subunit is responsible for the release of the new protein and, where appropriate, attachment to the ER via an intermediate docking protein that directs the newly synthesized protein through the membrane into the cisternal space.

GOLGI APPARATUS (GOLGI COMPLEX)

- It is a membranous organelle consisting of a stack of several flattened membranous cisternae, together with clusters of vesicles surrounding its surfaces. Seen in vertical section, it is often cup-shaped.

- The Golgi apparatus forms part of the pathway by which proteins synthesized in the rER undergo post-translational modification and are targeted to the cell surface for secretion or for storage in membranous vesicles.

Small transport vesicles from the rER, generated by a process of budding and pinching off, are received at one face of the Golgi stack, the convex cis-face (entry or forming surface). Here, they deliver their contents to the first cisterna in the series by membrane fusion. From the edges of this cisterna, the protein is transported to the next cisterna by vesicular budding and then fusion, and this process is repeated until the final cisterna at the concave trans face (exit or condensing surface) is reached. Here, larger vesicles are formed for delivery to other parts of the cell.

-The cis-Golgi network is a region of complex membranous channels interposed between the rER and the Golgi cis face, which receives and transmits vesicles in both directions.

-Its function is to select appropriate proteins synthesized on the rough endoplasmic reticulum for delivery by vesicles to the Golgi stack, while inappropriate proteins are shuttled back to the rough endoplasmic reticulum.

-The trans-Golgi network, at the other side of the Golgi stack, is also a region of interconnected membrane channels engaged in protein sorting.

- Here, modified proteins processed in the Golgi cisternae are packaged selectively into vesicles and dispatched to different parts of the cell.

-Within the Golgi stack proper, proteins undergo a series of sequential chemical modifications that started in the rER.

-These include: changes in glycosyl groups, e.g. addition of N-acetyl glucosamine and sialic acid; sulphation of attached glycosaminoglycans; protein phosphorylation.

LYSOSOMES

- These are dense cellular vesicles containing acidic digestive enzymes. They fuse with phagocytotic vesicles from the outer cell membrane, digesting the contents into small biomolecules that can cross the lysosomal lipid bilayer into the cell cytoplasm.

- Lysosomal action is crucial to the function of macrophages and polymorphs in killing and digesting infective agents, tissue remodeling during development and osteoclast remodeling of bone.

PEROXISOMES

-These are dense cellular vesicles, so named because they contain enzymes that catalyze the breakdown of hydrogen peroxide.

- They are involved in the metabolism of bile and fatty acids, and are primarily concerned with detoxification, e.g. D-amino acid oxidase and H2O2 catalase.

MITOCHONDRIA

-The mitochondrion is a membrane-bound organelle.

-It is the principal source of chemical energy in most cells.

-The numbers of mitochondria in a particular cell reflect its general energy requirements

-Cells with few mitochondria generally rely largely on glycolysis for their energy supplies.

-Mitochondria appear in the light microscope as long thin threads, or alternatively as spherical or ellipsoid bodies in the cytoplasm of most cells.

-In the electron microscope, mitochondria appears to have an outer and an inner unit membrane, separated by a variable gap termed the intermembrane space. The lumen is surrounded by the inner membrane and contains the mitochondrial matrix.

- The outer membrane is smooth and sometimes attached to other organelles.

-The inner membrane is deeply folded to form incomplete transverse or longitudinal tubular invaginations, cristae, which create a relatively large surface area of membrane.

-Cristae are most numerous and complex in cells with a high metabolic rate, e.g. cardiac muscles.

-The permeabilities of the two mitochondrial membranes differ considerably:

• The outer membrane is freely permeable to many substances because of the presence of large non-specific channels formed by proteins (porins)

•The inner membrane is permeable to only a narrow range of molecules. The presence of cardiolipin, a phospholipid, in the inner membrane may contribute to this relative impermeability.

-The outer membrane contains many gated receptors responsible for the import of raw materials like pyruvate and ADP, and the export of products such as oxaloacetate and ATP.

-Proteins of the Bd-2/Bax family are incorporated in this outer membrane and can release mitochondrial enzymes that trigger apoptosis

-The inner membrane contains transmembrane enzyme complexes of the electron transport chain, which generate H+ ion gradient.

-This gradient then drives the adjacent transmembrane ATPase complex to form ATP from ADP and P (oxidative phosphorylation)

- The inner matrix contains the enzymes of the Krebs cycle that generate the substrates of both the electron transport chain (FADH2 and NADH) and central metabolism (e.g. succinyl CoA & oxaloacetate).

- The mitochondrial matrix is an aqueous environment. It contains a variety of enzymes and strands of mitochondrial DNA with the capacity for transcription and translation of a unique set of mitochondrial genes

- Mitochondria of the sperm are not generally incorporated into the ovum at fertilization. Thus mitochondria (and mitochondrial genetic variations and mutations) are passed only through the female line.

THE CYTOSKELETON

- This is a complex network of structural proteins which regulates not only the shape of the cell, but also its ability to traffic internal cell organelles and even move in response to external stimuli. The major components are microtubules, intermediate filaments and microfilaments.

•Microtubules:

-These are made up of two protein subunits; “a” and “fi” tubulin.

-They form a 'highway', transporting organelles through the cytoplasm.

-During cell division, microtubules are rearranged by the microtubule-organizing centre (MTOC), which consists of centrosomes containing tubulin and provides a structure on which the daughter chromosomes can separate.

•Some anti cancerous drugs cause cell death by binding to microtubules and stabilizing them so much, thus mitotic spindles cannot form.

•Intermediate filaments:

-These form a network around the nucleus and extend to the periphery of the cell.

-They make cell-to-cell contacts with the adjacent cells via desmosomes, and with basement matrix via hemidesmosomes. Their function appears to be in structural integrity.

•Microfilaments:

-Muscle cells contain a highly ordered structure of actin and myosin filaments, which form the contractile system.

-These filaments are also present throughout the non muscle cells as truncated myosins, in the cytosol and beneath the plasma membrane.

-Cell movement is mediated by the anchorage of actin filaments to the plasma membrane at adherent junctions between cells

NUCLEUS

-Generally, it is the largest intracellular structure, usually spherical or ellipsoid in shape.

- Stains used to identify nuclei in tissue sections mainly detect the acidic molecules DNA

Nuclear membrane: - The nucleus is surrounded by two layers of membrane, each of which is a lipid bilayer, and which together form the nuclear membrane or envelope.

- The outer membrane layer and the lumen between the two layers are continuous with the rER.

- Like the rER, the outer membrane of the nuclear envelope is studded with ribosomes that are active in protein synthesis; the newly synthesized proteins pass into the perinuclear space between the two membrane layers.

-The transport of molecules between the nucleus and the cytoplasm is achieved by specialized nuclear pore structures that perforate the nuclear membrane.

-They act as highly selective directional molecular filters, permitting proteins such as histones and gene regulatory proteins (which are synthesized in the cytoplasm but function in the nucleus) to enter the nucleus, and molecules that are synthesized in the nucleus but destined for the cytoplasm (e.g. ribosomal subunits, transfer RNAs and messenger RNAs), to leave the nucleus.

Chromatin:

-DNA is organized within the nucleus in a DNA-protein complex known as chromatin. The protein constituents of chromatin are the histones and the non-histone proteins.

- Non-histone proteins are an extremely heterogeneous group that includes DNA and RNA polymerases and gene regulatory proteins.

- Histones are the most abundant group of proteins in chromatin, primarily responsible for the packaging of chromosomal DNA into its primary level of organization, the nucleosome.

- There are five histone proteins: H1, H2A, H2B, H3 and H4; the last four combine in equal ratios to form a compact octameric nucleosome core. The DNA molecule (one per chromosome) winds 1.65 times around each nucleosome core, taking up 146 nucleotide pairs (the nucleosome proper)

- However, chromatin rarely exists in this simple form and is usually packaged further into a 30 nm thick fibre, involving a single H1 histone per nucleosome, which interacts with both DNA and protein to impose a higher order of nucleosome packing.

- In a typical interphase nucleus, euchromatin (nuclear regions that appear pale in appropriately stained tissue sections, or relatively electron-lucent in electron micrographs ;) is likely to consist mainly of 30 nm fibres and loops, and contains the transcriptionally active genes.

- Heterochromatin (nuclear regions that appear dark in appropriately stained tissue sections or electron-dense in electron micrographs) is characteristically located mainly around the periphery of the nucleus, except over the nuclear pores, and around the nucleolus.

- Heterochromatin includes non-coding regions of DNA, such as centromeric and telomeric regions, which are known as constitutive heterochromatin.

Nucleolus: - They are the site of most of the synthesis of rRNA and assembly of ribosome subunits.

- Ultrastructurally, the nucleolus appears as a pale fibrillar region (non-transcribed DNA), containing dense fibrillar cores (sites of rRNA gene transcription) and granular regions (sites of ribosome subunit assembly) within a diffuse nucleolar matrix.

- During mitosis the nucleolus breaks down. It reforms after telophase, in a process initiated by the onset of transcription in nucleolar organizing centres on each chromosome.

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