OBJECTIVES

This chapter should help you to:

· Perceive the inseparability of structure and function in living organisms.
· Know the names and functions of the cellular components (organelles and inclusions).
· Know the important subunits of each cellular component and the role of each subunit in the component's function.
· Name the general and specialized functions of cells and the role of each cellular component in each function.
· Recognize a cell's structural components in a light or electron photomicrograph and hence predict the cell's function(s).
· Predict which structures will be present in a cell from its function.
· Predict the functional deficit(s) that would occur in a cell as a result of specific structural aberrations.
· Predict the cell component(s) likely to be involved in a functional deficit.
· Explain and give examples of cell differentiation.

I. GENERAL FEATURES OF CELLS

A. Subunits of Life: Cells are the structural and functional units of life (and of disease processes) in all tissues, organs, and organ systems. Therefore, students of histology must perceive that a cell's capabilities and limitations are implicit in its structure.

B. Prokaryotes and Eukaryotes: There are 2 basic cell types. Prokaryotic cells are typically small, single-celled organisms leg, bacteria) that lack a nuclear envelope, histones, and mem branous organelles. Eukaryotic cells exist primarily as components of multicellular organisms. This chapter covers the basic structural and functional features of eukaryotic cells. Specific human cell types are described in later chapters.

C. CellularComponents: Eukaryotic cells have3 majorcomponents: The cell membranes (II) separate a cell from its environment and form distinct functional compartments (nucleus, organelles) in the cell. The outer cell membrane is called the plasma membrane, or plasmalemma. The cytoplasm (III) surrounds the nucleus and is enclosed by the plasma membrane. It contains the structures and substances needed to decode the instructions of DNA and carry on the activities of the cell. The membrane-bound nucleus (IV) contains a cell's DNA, which encodes the genetic information needed for protein synthesis and thus for all the activities of the cell. It also has components that help determine which parts of the genetic code are used and that deliver coded information to the cytoplasm.

D. Cellular Functions: Three activities basic to living organisms are nourishment, growth and development, and reproduction. Functions directed toward these activities are described in this chapter. More specialized cell functions will receive more detailed treatment in later chapters.

II. CELL MEMBRANES

A. Biochemical Components (Fig 3-1):


1. Lipids are present in cell membranes as phospbolipids (Fig 3-1, E), sphingolipids, and cholesterol (Fig 3-1, A). Phospholipids leg, lecithin) are by far the most abundant. Each phospholipid molecule has a polar (hydrophilic) phosphate-containing head group (Fig 3-1, G) and a nonpolar (hydrophobic) pair of fatty-acid tails (Fig 3-1, Fl. Membrane phospholipids are arranged in a bilayer with their tails directed toward one another at the center of the membrane. In electron micrographs of osmium-stained tissue, a single membrane, or unit membrane, has 2 dark outer lines with a lighter layer between them. This trilaminar appearance may be due to the deposition of reduced osmium on the hydrophilic head groups.
2. Protein may make up more than 50% of membrane weight. Most membrane proteins are globular and belong to one of the following 2 groups: a Integral membrane proteins (Fig 3-1, C and D) are tightly lodged in the lipid bilayer; detergents are required to extract them. They an folded, with their hydrophilic amino acids in contact with the phosphate groups of the membrane phospholipids and their hydrophobic amino acids in contact with the fatty-acid tails. Some protrude from only one membrane surface (Fig 3-1, D), whereas others, called transmembrane proteins (Fig 3-1, C), penetrate the entire membrane and protrude from both sides. Some trans membrane proteins, such as protein3-tetramer, may serve as hydrophilic channels for the passage of water and water-soluble materials through hydrophobic regions. Freeze Fracture preparations often split plasma membranes through the hydrophobic region, between the ends of the phospholipid's fatty-acid tails (Fig 3-1). The integral proteins exposed in the process end up mainly in the side closest to the cytoplasm, termed the P (protoplasmic) face. The membrane half nearest the environment, the E (ectoplasmic) tace, typically has a smoother appearance. b. Rripheral membrane proteins (Fig 3-1, Il) are more loosely associated with the inner or outer membrane surface; some are globular, some filamentous. In erythrocytes, exam ples on the cytoplasmic surface include spectrin, which helps maintain membrane integrity, and ankyrin, which links spectrin to the portion of protein-3-tetramer facing the cytoplasm.
3. Carbohydrates occur on plasma membranes mainly as oligosaccharide moieties of membrane glycoproteins (Fig 3-1, B) and glycolipids. Membrane oligosac charides have a characteristic branching structure and project from the cell's outer surface, forming a superticial coat called the glycocalyx that participates in cell adhesion and recogni tion.

B. Membrane Organization: The widely accepted Buid mosaic model describes biologic mem branes as "protein icebergs in a lipid sea." Integral proteins exhibit lateral mobility and may undergo rearrangement determined by their association with peripheral proteins, cytoskeletal filaments within the cell (III.I), membrane components of adjacent cells, and components of the extracellular matrix. Integral proteins sometimes diffuse to and accumulate in one membrane region, a process termed capping.

C. Membrane Functions:


1. Selective permeability The cell membrane forms an effective seal between a cell or organelle's internal and external environment, preventing intrusion of harmful substances, dispersion of macromolecules, and dilution of enzymes and substrates. Membranes display selective permeability, essential to maintaining the functional steady state, or homeostasis, required for cell survival. Homeostatic mechanisms attributable to the cell membrane main tain optimal intracellular concentrations of ions, water, enzymes, and substrates. Three mechanisms allow passage of selected molecules. a. Passive diffusion. Certain substances leg, water) can cross the membrane in either direction, following a concentration gradient. Passive diffusion does not require energy expenditure. b. Facilitated diffusion. Certain molecules leg, glucose) must be helped across the mem brane by a membrane component. Thisfacilitared diffusion is often unidirectional, but it follows a concentration gradient and requires no energy. c. Active transport. Some nondiffusible molecules can move into or out of a ceil either along or against a concentration gradient. Such movement requires energy, usually as ATP. An example of this active transport is the sodium pump (Naf/Kf-ATPase), which can expel sodium ions from a cell even when the sodium concentration is higher outside than inside.
2. Signal transduction. Receptors with strong binding affinities for exogenous signals such as hormones are located at the cell surface. The signal molecule to which a receptor binds specifically is called its ligand, Once receptors bind their ligands, they may transmit the signal to the cell interior by one of a variety of mechanisms: a. Receptors may transmit the signal through their association with cytoskeletal components at the inner surface of the membrane. b. Receptors may interact with other membrane components to produce second-messenger molecules, which then transmit the message to the cell's interior. c. Signal-receptor complexes may be moved into the region of a coated pit and be endo- cytosed (see below), carrying the signal into the cell. d. The receptor itself may have enzyme activity (stimulated by binding the signal molecule) and transmit the signal by enzymatically altering intracellular proteins.
3. Endocytosis. Cells engulf extracellular substances and bring them into the cytoplasm in membrane-limited vesicles by mechanisms described collectively as endocytosis. a. In phagocytosis ("cell eating"), the cell engulfs insoluble substances, such as large macromolecules or entire bacteria. The vesicles formed are termed phagosomes, b. In pinocytosis ("cell drinking"), the cell engulfs small amounts of Buid, which may contain a variety of solutes. Pinocytotic vesicles are usually smaller than phagosomes. c. In receptor-mediated endocytosis, the cell engulfs foreign substances along with their own surface receptors. Binding of ligand to receptor induces the collection of ligand receptor complexes in coated pits, shallow membrane depressions with clathrin protein coats lining their cytoplasmic surfaces. invagination and pinching off of the pit creates a coated vesicle, which carries the ligand-receptor complexes into the cell. The clathrin coating is released from the vesicle, now termed an endosome, and the ligands dissociate from the receptors. The later endosome, or CURL (compartment of uncoupling of receptor and ligand), becomes more tubular and divides into 2 portions, segregating the receptors from the ligands. The receptors are returned to the plasma membrane, and the ligands ate directed to lysosomes.
4. Exocytosis. Exocytosis removes substances from the cell. Cells use this process both for secretion and for excretion of undigested material. A membrane-limited vesicle or secretory granule fuses with the plasma membrane and releases its contents into the extracellular space, without disrupting the plasma membrane.
5. Compartmentalization. Membranes selectively inhibit the passage of most water-soluble substances. The cytoplasm has many membrane-limited compartments (organelles), each with a different internal environment with respect to concentrations of solutes. This compart mentalization prevents dilution of metabolic intermediates and cofactors in multistep bio chemical reactions, and it protects sensitive reactions from intrusion of extraneous sub stances.
6. Spatiotemporal organization of metabolic processes. Some cellular membranes leg, the inner mitochondrial membrane and the Golgi complex) contain series of enzymes arranged so that intermediates in multistep metabolic processes are passed from one enzyme to the next. The spatial arrangement of enzymes maintains the chronologic order of such processes, and it sets rate limitations by maintaining local concentrations of intermediates.
7. Storage, transport, and secretion. Membrane-limited vesicles isolate certain substances during intracellular processes. Substances in vesicles may be kept for later use (storage), shuttled from one compartment to another for further processing (transport, II.D), or expelled from the cell (secretion, II.C.4).

D. Membrane Flow: The movement of membrane from one organelle to another is called mem brane flow and is a general feature of organelle function. Membranes bud as vesicles from an organelle and fuse with another membrane, allowing the amount of membrane in a particular organelle to change without membrane synthesis or breakdown.

III. CYTOPLASM

Cytoplasmic structures comprise 3 groups. Organelles are membrane-bound, enzyme-containing subcellular compartments leg, mitochondria). Each type of organelle has a distinctive structure and performs unique functions. Cytoplasmic inclusions are structures, membrane-bound or not, that are generally more transient than organelles and less actively involved in cell metabolism leg, lipid droplets). Organelles and inclusions are discussed in sections III.A-III.H. The cytoskeleton (III.I) is a proteinaceous supporting network within the cytoplasm; components of this network (microtubules) also form discrete cytoplasmic structures such as centrioles.

A. Mitochondria: The largest organelles, mitochondria, provide energy for the cell.


1. Structure. Mitochondria are comparable in size to bacteria (usually 2-6 um in length and 0.2 um in diameter but quite variable) and have various shapes: spheric, ovoid, filamentous. Each mitochondrion is bounded by 2 unit membranes. a. The outer mitocbondrial membrane (Fig 3-2, A) has a smooth contour and forms a continuous but relatively porous covering. It is freely permeable to various small mole- cules. b. The inner mitochondrial membrane (Fig 3-2, B) is less porous and thus is semiperme able. It has many infoldings, or cristae (Fig 3-2, C), that project into the mitochondrion's interior. The mitochondrial cristae of most cells are shelflike, but those in steroid secreting cells are typically tubular. The inner surface is covered by inner membrane snbunits (Fig 3-2, H), also called FI subunits (or lollipops, because of their shape); these are sites of mitochondrial ATFase activity. Mitochondrial ribosomes also associate with the inner surface. Intercalated within the inner membrane are components of the electron transport system, including enzymes and cofactors with important roles in mitochondrial function leg, cytochromes, dehydrogenases, flavoproteins). c. The mitochondrial membranes create 2 membrane-limited spaces. The intermembrane space (Fig 3-2, Fl is Located between the inner and outer membranes and is continuous with the intracristal space (Fig 3-2, E), which extends into the cristae. The intercristal space, or matrix space (Fig 3-2, D), is enclosed by the inner membrane and contains the mitochondrial matrix. d. The mitochondrial matrix contains water, solutes, and large matrix granules (Fig 3-2, G), believed to be concerned with mitochondrial calcium ion concentration. It also contains circular DNA and mitochondrial ribosomes similar to those of bacteria. The matrix contains numerous soluble enzymes involved in such specialized mitochondrial functions as the Krebs cycle (tricarboxylic acid cycle), B-oxidation of lipids, and mitochondrial DNA synthesis.
2. Function. Mitochondria provide energy for chemical and mechanical work by storing energy generated from cellular metabolites in the high-energy bonds of ATP. The ATP leaves the mitochondrion and releases its stored energy at a variety of intracellular sites. Mitochondria synthesize their own DNA and some proteins. They grow and reproduce by fission or budding and can undergo rapid movement and shape changes.
3. Location. Mitochondria are found in nearly all eukaryotic cells, and in most they are dispersed throughout the cytoplasm. However, they accumulate in cell types and intracellular regions with higher energy requirements. Cardiac muscle cells are notable for their abundant mitochondria. Epithelial cells lining the kidney tubules have abundant mitochondria inter digitated between basal plasma membrane infoldings where active transport of ions and water occurs.

B. Ribosomes: The ribosomes are protein-synthesizing organelles. There are 2 basic types. Mitochondrial (like prokaryotic) ribosomes are smaller (20 nm) than the cytoplasmic ribosomes of eukaryotes (25 nm).


1. Structure. Each type of ribosome has 2 unequal ribosomal subunits, named for their sedimentation rates during ultracentrifugation (but often called simply "large" and "small"). Mitochondrial ribosomes (70S overall) have a 50S and a 30S subunit; cytoplasmic ribosomes (80S overall) have a 60S and a 40S subunit. Cytoplasmic ribosomes are composed of ribosomal RNA (rRNA) synthesized in the nucleolus and associated proteins synthesized in the cytoplasm. They are intensely basophilic. Light microscopy reveals cytoplasmic accumu lations of ribosomes as basophilic patches, formerly termed ergastoplasm in glandular cells and Nissl bodies in neurons. In electron micrographs, ribosomes appear as small, electron dense cytoplasmic granules.
2. Location and function. Cytoplasmic ribosomes occur in 2 forms. Free ribosomes are individual ribosomes dispersed in cytoplasm. Polyribosomes, or polysomes, are groups of ribosomes distributed along a single strand of messenger RNA (mRNAX an arrangement that permits synthesis of multiple copies of a protein from the same message. Ribosomes read (translate) the mRNA code and thus play a critical role in assembling amino acids into specific proteins. Polysomes occur free in the cytoplasm (free polysomes) and attached to membranes of the rough endoplasmic reticulum. Free polysomes synthesize structural pro teins and enzymes for intracellular use. Polysomes of the rough endoplasmic reticulum synthesize proteins to be secreted or sequestered.

C. Endoplasmic Retlculum: The endoplasmic reticulum (ER) is a complex organelle in volved in the synthesis, packaging, and processing of various cell substances. It is a freely anastomosing network (reticulum) of membranes that form vesicles, or cisternae; these may be elongated, flattened, rounded, or tubular. Transfer vesicles (transitional vesicles) are small, membrane-limited vesicles that bud from the ER and cross the intervening cytoplasm to reach the Golgi complex for further processing or packaging of their contents. In mature cells, ER occurs in 2 forms, called rough and smooth.


1. Rough endoplasmic reticulum. a. Structure. The rough endoplasmic reticulum (RER) also called granular endoplasmic reticulum, is studded with ribosomes. many of them in polysomal clusters. RER cisternae are typically parallel, flattened, and elongated, especially in cells specialized for protein secretion leg, pancreatic acinar cells, plasma cells), in which RER is particularly abundant. The ribosomes give RER basophilic staining properties. The fine structure of RER (membranes and individual ribosomes) is visible only with the electron microscope. Proteins unique to RER membranes include docking protein, which functions as a receptor, and ribophorins i and II, b. Function. RER is mainly concerned with the synthesis of proteins for sequestration from the rest of the cytoplasm, ie, secretory proteins such as collagen, proteins for incorpora tion into cell membranes, and lysosomal enzymes (separated from the rest of the cytoplasm to prevent autolysis). Ribosomes or free polysomes begin reading at the 5' end of mRNAs and move toward the 3' end. The 5' end of mRNAs for secretory and se questered proteins carries the code for a 20- to 25-amino acid signal sequence. The signal sequence is translated first on a free polysome and interacts with a signal recognition particle (SRP) which is 6 polypeptides plus a 7S RNA molecule. SRP inhibits further translation until the SRP-polyribosome complex binds to the RER docking protein; then the SRP is released and translation continues. Ribophorins I and II mediate the attachment of the large ribosomal subunit to the RER membrane. They may also provide a hydrophilic channel for vectorial discharge (unidirectional passage) of nascent protein into the RER lumen, where the signal sequence is cleaved by signal peptidase and the remainder of the nascent protein is folded and modified. One important posttranslational modification is core glycosylation: Oligosaccharides, usually high in mannose residues, are transferred from a lipid carrier leg, dolichol phosphate) to amino acids, especially asparagine. The oligosaccharides may "address" proteins for transport to intracellular destinations. c. Location. The RER is suspended in the cytoplasm and shows continuity at various points with the outer membrane of the nuclear envelope. RER in protein-secreting epithelial cells often lies in the basal cytoplasm, between the basal plasma membrane and the nucleus.
2. Smooth endoplarsmic reticulum a. Structure. The smooth endopiasmic reticulum (SER) lacks ribosomes and thus ap pears smooth in electron micrographs. SER cisternae are more tubular or vesicular than those of the RER. The SER stains poorly, if at all, so with the light microscope it is indistinguishable from the rest of the cytoplasm. b. Function. Because it lacks ribosomes, the SER cannot synthesize proteins. It has many enzymes, important in lipid metabolism, steroid hormone synthesis, glycogen break down (glucose-6-phosphatase). and detoxification. The last occurs via enzymatic con jugation, oxidation, and methylation of potentially toxic substances. c. Location. The SER is suspended in the cytoplasm of many cells and is especially abundant in cells that synthesize steroid hormones leg, cells of the adrenal cortex and the gonads). It is also abundant in liver cells (hepatocytes), where it is involved in glycogen metabolism and drug detoxification. Specialized SER termed sarcoplasmic reticulum is found in striated muscle cells, where it helps to regulate muscle contraction by sequester ing and releasing calcium ions.

D. Golgi Complex: The Golgi complex (Golgi apparatus) participates in many activities, partic ularly those associated with secretion. It has an essential role in coordinating membrane Bow and vesicle traffic among organelles,


1. Structure. This membranous organelle comprises 3 major compartments: (1) a stack of 3-10 discrete, slightly curved, flattened cisternae; (2) numerous small vesicles peripheral to the stack; and (3) a few large condensing vacuoles at the concave surface of the stack. The cis face (convex face, forming face) of the stack is usually closest to adjacent dilated ER cisternae and is surrounded by transfer vesicles. Its cistemae stain more darkly with osmium. The trans face (concave face, maturing face) often harbors several condensing vacuoles and generally faces away from the nucleus,
2. Functions a. Polysaccharide synthesis. The Golgi complex contains glycosyltransferases that initiate, lengthen, or shorten polysaccharide or oligosaccharide chains one sugar at a time. b. Modification of secretory products. Golgi enzymes glycosylate proteins and lipids and sulfate glycosaminogiycans (GAGs). The Golgi complex is thus important in the syn- thesis of secretory glycoproteins, proteoglycans, glycolipids, and sulfated GAGs. c. Packaging of secretory products.  Products synthesized by the ER are packaged in vesicles by the Golgi complex. These secretory vesicles, or secretory granules, are transported to the plasma membrane for exocytosis (II.C.4). d. Concentration and storage of secretory products. The Golgi complexes of some cells concentrate and store secretory products prior to secretion. Such concentration is a major function of the condensing vacuoles on the trans face of the Golgi complex, which also often serve as precursors to secretory granules.
3. Location. The Golgi complex is typically near the nucleus (juxtanuclear) and is often found near centrioles (which may also have an important role in directing vesicle traffic). Golgi complexes are best developed in neurons and glandular cells, which are specialized for secretion.
4. Flow of materials through the Golgi complex. Secretory materials have long been thought to follow a one-way route through the Golgi complex. In this scheme, transfer vesicles bud from the ER and fuse with the forming (cis) face. The vesicle contents are then modified as they pass successively from cisterna to cisterna toward the maturing (trans) face, which buds off the secretory vesicles containing the final product. However, this view is now being challenged as an oversimplification. Recent evidence indicates that Golgi-associated vesicles differ in their source, destination, function, contents, and surface composition. This and evidence that certain nonclathrin, vesicle-coating proteins leg, B-COP) are associated with specific regions of the Golgi complex suggest that various vesicle types can fuse with, and bud from, the cis, trans, and intermediate Golgi membranes. We appear to have much to learn about the complex vesicle traffic patterns associated with this organelle.

E. Phagosomes: Phagosomes are membrane-limited vesicles of various sizes containing mate rial destined for lysosomal digestion. Two major types are known. Heterophagosomes contain the products of heterophagy, ie, material of extracellular origin ingested by phagocytosis. Autophagosomes contain the products of autophagy, ie, material of intracellular origin such as worn or damaged organelles. The digestion of phagosomal contents begins when a phagosome fuses with one or more primary lysosomes to form a secondary lysosome, as described below. (Note: Some authors use the term heterophagosome to refer to secondary lysosomes.)

F. Lysosomes: Lysosomes are spheric, membrane-limited vesicles that may contain more than 50 enzymes each and function as the cellular digestive system. Their characteristic enzyme activities distinguish them from other cellular granules. The enzyme most widely exploited for their identification is acid phosphatase, because it occurs almost exclusively in lysosomes. Other enzymes common in lysosomes are ribonucleases, deoxyribonucleases, cathepsins, sul fatases, P-giucuronidase, and phospholipases and other proteases, glucosidases, and lipases. An inherited deficiency or lack of a particular lysosomal enzyme can result in life-threatening accumulations of its substrate in the cytoplasm. Lysosomal enzymes usually occur as glycopro teins and are most active at an acidic pH. Lysosomes occur in various sizes and electron densities, depending on their level of activity.


1. Primary lysosomes are small (5-8 nm in diameter), with electron-dense contents; they appear as black circles in electron micrographs. They are the storage form of lysosomes. and their enzymes are mostly inactive. Lysosomal enzymes synthesized and core-glycosylated in the RER are transferred to the Golgi complex for further glycosylation and packaging in vesicles. The primary lysosomes disperse through the cytoplasm. They are found in most cells but are most abundant in phagocytic cells leg. macrophages, neutrophils).
2. Secondary lysosomes are larger and less electron-dense and have a mealed appearance in electron micrographs. They are formed by the fusion of one or more primary lysosomes with a phagosome. Their primary function is digesting products of heterophagy and autophagy. When the lysosomal enzymes mix with the phagosome contents, they become active. Diges tion produces metabolites for cell maintenance and growth (small molecules diffuse into the surrounding cytoplasm) and aids in the turnover of organelles. Lysosomal enzymes also catabolize certain products of cell synthesis, thus regulating the quality and quantity of secretory material. Secondary lysosomes occur throughout the cytoplasm in many cells, in numbers that reflect the cell's lysosomal and phagocytic activity.
3. Residual bodies are membrane-limited inclusions of various sizes and electron densities associated with the terminal phases of lysosome function. They contain undigestible mate rials such as pigments, crystals, and certain lipids. Some cells leg, macrophages) expel residual bodies as waste, but long-lived cells leg, nerve, muscle) tend to accumulate them. In the latter, waste-containing residual bodies reflect cellular aging and may be referred to as "wear-and-tear pigment," or lipofuscin granules, These granules appear yellowish brown in light microscopy and as electron-dense particles in electron micrographs.

G. Peroxisomes: Peroxisomes are membrane-limited, enzyme-containing vesicles somewhat larger than primary lysosomes. In some animals (not humans), they are distinguishable from hydrogen peroxide metabolism. They contain urate oxidase, hydroxyacid oxidase, and D-amino acid oxidase, which produce hydrogen peroxide capable of killing bacteria; they also contain catalase, which oxidizes various substrates and uses the hydrogen removed in the process to convert the toxic hydrogen peroxide to water. Peroxisomes also participate in gluconeogenesis by assisting in the B-oxidation of fatty acids. They are found dispersed in the cytoplasm or in association with the SEP.

H. Other Cytoplasmic Inclusions: Prominent among inclusions serving as storage depots are 'i spheric lipid droplets, which differ in appearance depending upon the type of histologic prepa ration. Glycogen granules are inclusions that are PAS-positive in light microscopy and appear in electron micrographs as rosettes of electron-dense particles. Both lipid droplets and glycogen j granules lack a limiting membrane. Melanin is a brownish pigment widely distributed in vertebrates, often found in electron-dense, membrane-limited granules termed melanosomes, It is particularly abundant in epidermal cells and in the pigment layer of the retina.

I. Cytoskeleton: The cytoskeleton, a mesh of filamentous elements called microtubules, microfilaments, and intermediate Blaments, provides structural stability for the maintenance of cell shape. It is important in cell movement and in the rearrangement of cytoplasmic components. In one model of cytoplasmic organization, organelles and cytoplasmic inclusions are embedded in a delicate meshwork termed the microtrabecuiar lattice which incorporates the cytoskeleton, enzymes, and other cytoplasmic constituents previously considered soluble and randomly dispersed in an amorphous cytosol, This hypothetic lattice may provide a framework for coordinated arrays of enzymes involved in multistep reactions, as do some membranes (II.C.6). However, the possibility that the latticelike appearance of the cytoplasm is a fixation artifact has not been ruled out.

1. Microtubules. a. Structure. Microtubules are the thickest cytoskeleton components, with diameters of 24 nm. They are fine tubular structures of variable length, with dense walls (5 nm thick) and a clear internal space (14 nm across). The walls are composed of subunits called tubulin heterodimers, each of which consists of one a-tubulin and one B-tubulin protein molecule. The tubulin heterodimers are arranged in threadlike polymers called protofila ments, Thirteen of these align parallel to one another to form the wall of each micro tubule. Microtubules increase in length by adding new heterodimers to one end, called the nucleation site, This polymerization can be controlled experimentally by regulating calcium ion concentration or by treating cells with antimitotic alkaloids. Colchicine blocks the process by binding to the nucleation site. Vinblastine disrupts microtubules by binding to free tubulin. b. Function. Microtubules have roles in the maintenance of cell shape, axoplasmic trans port in neurons, melanin dispersion in pigment cells, chromosome movements during mitosis, organization of the Golgi complex, and the shuttling of vesicles within the cell. Unlike microfilaments, microtubules are unable to contract. Shortening occurs via de polymerization . c. Location. Microtubules are found throughout the cytoplasm of most cells and in highly organized groupings in centrioles, cilia, flagella, basal bodies, and the mitotic spindle apparatus.
2. Centrioles. a. Structure. A centriole is a cylindric group of microtubules, 150 nm in overall diameter and 350-500 nm long, containing 9 microtubule triplets in a pinwheel array. Each microtubule in a triplet shares a portion of the wall of the neighbor ing microtubule. An interphase (nondividing) cell has a pair of adjacent centrioles with Perpendicular long axes, each surrounded by several electron dense satellites, or pericentriolar bodies. Other cytoplasmic microtubules radiate from the pericentriolar bodies into the cytoplasm. b. Function. Centrioles are the structural organizers of the cell. Centriole duplica tion is a prerequisite for cell division, and during mitosis the centrioles organize the microtubules of the mitotic spindle. Even in vitro, isolated centrioles can control microtubule polymerization; in the cell, centrioles may transmit un known physical organizing forces via the microtubules radiating from the peri centriolar bodies. Through their effects on microtubules, centrioles may have some control over organelle, vesicle, and granule traffic within the cell. Centrioles give rise to basal bodies (see below). c. Location. Between cell divisions, centrioles are near the nucleus, often sur rounded by Golgi complexes. The centrioles and associated Golgi complexes constitute the cell cytocenter, which appears as a clear zone near the nucleus. During the S phase of interphase, each centriole duplicates by giving rise to a procentriole that grows at right angles to the original. During mitosis, the new centriole pairs migrate to opposite cell poles to organize the spindle.
3. Basal bodies. In cells bearing cilia or flagella, centrioles migrate to the apical plasma membrane and give rise to basal bodies in a manner similar to centriole self duplication. Basal bodies are structurally similar to centrioles, with 9 microtubule triplets. They occur in the cytoplasm, one at the base of each cilium or flagellum, and serve as the anchoring points and microtubule organizers for these structures. (3) Cilia. A ciliated cell usually has hundreds of cilia, motile 5-10 pm long, 0.2 um wide, cell surface evaginations covered by plasma membrane. Each cilium contains a core, or axoneme, composed of 9 peripheral microtubule doublets surrounding a pair of unjoined microtubules (the "9 + 2" arrangement). Partners in a doublet ate called subfibers A and B. Subfiber A is a complete microtubule, having 13 tubulin pro tofilaments, Subfiber B has 10 or 11 protofilaments and is completed by sharing part of its partner's wall. A pair of arms made of dynein (a protein with ATPase activity) extends from the wall of each subfiber A toward the adjacent doublet. Protein bridges called nexins link adjacent doublets, and radial spokes link the doublets to the sheath surrounding the central microtubule pair. Each axoneme is organized by and anchored in a basal body. (4) Flagella, A flagellum is like a cilium, but it is longer and there is usually only one or 2 flagella on a cell. In humans and other mammals, Bagella occur only in the tails of spermatozoa, which are typically 50-55 Clm long and 0.2-0.5 pm thick along most of their length. The axoneme of a tlagellum is identical to that of a cilium. (5) Mitotic spindle apparatus, The spindle-shaped microtubule array called the mitotic spindle apparatus occurs between 2 pairs of centrioles at opposite poles of mitotic cells. Some spindle microtubules (continuous fibers) extend from centriole to centriole. Others (chromosomal fibers) extend from one centriole to the centromere of a chromosome. The spindle apparatus is crucial for chromosome separation during mitosis.
4.Microfilaments a. Structure, Microfilaments are the thinnest cytoskeletal components (5-7 nm wide). They are usually composed of one of several types of actin protein. In striated muscle cells, actin filaments form a stable paracrystalline array in association with filaments of myosin. Actin filaments in other cells are less stable and can dissociate and reassemble. These changes are regulated in part by calcium ions and cyclic AMP and by actin binding proteins in the cytoplasm. b. Function. Microfilaments are contractile, but to contract they usually must interact with myosin. In muscle cells, myosin forms thick tilaments. In nonmuscle cells, it exists in soluble form. Treatment with cytochalasins disrupts microfilament organization and interferes with the following functions: endocytosis; exocytosis; contraction of microvilli; cell movement; movement of organelles, vesicles, and granules; cytoplasmic streaming; maintenance of cell shape; and equatorial constriction of dividing cells. c. Location, In nonmuscle cells, microfilaments are generally distributed as an irregular mesh throughout the cytoplasm. Local accumulations may be present as a thin sheath beneath the plasma membrane called the terminal web, as parallel strands in cores of microvilli, in the cytoplasm at the leading edge of various types of pseudopods, in association with organelles or othercytoplasmic components, or as a belt ("purse string") around the equator of dividing cells.
5. Intermediate filaments a. Structure, Intermediate filaments are intermediate in thickness (10-12 nm) between microtubules and microfilaments. They are composed of proteins that are structurally related to nuclear lamins and differ depending on the cell type. Examples: cytokeratins in epithelial cells, vimentin in mesenchymally derived cells leg, fibroblasts. chondrocytes), desmin in muscle cells, glial fibrillary acidic protein in glial cells, neurofilaments (intermediate filament bundles) in neurons. b. Function. Intermediate filament function is currently being investigated. They are proba bly involved m maintaining cell shape, possibly as components of the microtrabecular lattice. The significance of their similarity to nuclear lamins is not clear. c. Location. Most intermediate filament types are distributed throughout the cytoplasm. Their distribution may represent a highly ordered arrangement that is not yet understood. One example of an ordered arrangement of intermediate filaments is the cytokeratin containing tonofilaments of desmosomes, discussed in Chapter 4.

IV. NUCLEUS

A. General Considerations: Nuclei vary in appearance from tissue to tissue and cell to cell, but each generally has a nuclear envelope, chromatin, nucleoplasm, and one to several nucleoli. Although certain mature cells leg, erythrocytes) lack nuclei, at least one nucleus is present at some stage in all eukaryotic cells. The nucleus contains a linear code (DNA) for the synthesis of cell components and products, conferring upon the cell a range of adaptability to changing environmental conditions and to extrinsic signals such as hormones. The microscopic appear ance of the nucleus is important in identifying and classifying both normal and diseased cells and tissues. Nuclei display wide variations in (l) size, both absolute and relative to the amount of cytoplasm (nucleocytoplasmic ratio); (2) number per cell, allowing classification of cells as enucleate, mononucieate, binucleate. or multinucleate; (3) chromatin pattern, ie, the amount and distribution of heterochromatin; and (4) location, eg, basal, central, eccentric.

B. Nuclear Envelope: The nuclear contents are set apart from the cytoplasm by a double membrane called the nuclear envelope and a narrow (40-70 nm) intermembrane space called the perinuclear cisterna, or perinuclear space. The nuclear envelope is often considered an exten sion of the RER, because its outer surface is often peppered with ribosomes and shows occa sional continuities with the RER. The inside of the inner membrane is lined with the fibrous lamina, a layer consisting of proteins called lamins, The envelope is perforated by many nuclear pores, each of which has a diameter of about 70 nm and is bounded by 8 globular subunits called annular proteins, which present an octagonal appearance in some preparations. Each pore is covered by a thin, proteinaceous diapbragm, The pores provide a channel for the movement of important molecules between the nucleus and cytoplasm, including nucleic acids synthesized in the nucleus and used in the cytoplasm (mRNA. rRNA. tRNA) and proteins synthesized in the cytoplasm and used in the nucleus (histones, polymerases).

C. Chromatin: Nuclear chromatin is intensely basophilic and consists of DNA and associated histone and nonhistone proteins.

1. Nucleosomes. Isolated chromatin appears in electron micrographs as thin strands studded with beadlike particles at regular intervals. Each strand is a double-helical molecule of DNA, and the particles are the repeating structural subunits of chromatin termed nucleosomes. Each nucleosome is composed of 166 base pairs of the DNA strand coiled around a core of 8 histones (2 copies each of H2A, H2B, H3, and H4). The portion of the strand between 2 nucleosomes contains an additional 48 base pairs and is called the linker region. Another histone (usually H1) is bound to the outside of the nucleosome and to the linker. The beaded strand coils into a superhelix with 6 nucleosomes per turn (selenoid) to form the condensed form of chromatin, ie, heterochromatin.
2. Chromatin types. Nuclei containing highly coiled chromatin, termed heterochromatin, stain darkly with basic dyes. Because the DNA of chromatin must uncoil to be transcribed, cells with dark-staining (heterochromatic) nuclei ate less active in DNA transcription and use a smaller portion of their total genome than other cells. Uncoiled chromatin, termed eu chromatin, stains poorly and is difficult to distinguish even by EM. Large, pale-staining (euchromatic) nuclei usually indicate greater transcriptional activity and faster cell division.
3. Chromatin pattern. The amount and distribution of nuclear chromatin are often used to identify cell types, especially in cells with no characteristic cytoplasmic staining properties. Even in mostly euchromatic nuclei, a rim of heterochromatin is often found on the inner surface of the nuclear envelope in association with the fibrous lamina. This envelope associated heterochromatin allows resolution of the nuclear boundary with the light micro- scope.
4. Chromosomes, the most highly condensed form of chromatin, are visible during mitosis. To form chromosomes, selenoids fold further and wind on a central nonhistone protein scaffold. Of the 46 chromosomes present in human cells, 44 (the somatic chromosomes) occur in 22 pairs of structurally similar chromosomes. The other pair (sex chromosomes) consists of dissimilar chromosomes (XY) in males and similar ones (XX) in females. In females, only one X chromosome (either of the two) is used by each cell; the inactive X chromosome is often visible as a clump of heterochromatin termed sex chromatin, or the Barr body In most cells, the Ban body is attached to the inner surface of the nuclear envelope. In a neutrophilic leukocyte, it may appear as a drumstick-shaped appendage of the lobulated nucleus.
5. Karyotyping, A cell's karyotype is its chromosome inventory or a picture of its chrome somes arranged by chromosome type. Preparing such a picture is called karyotyping. Cells in culture are stimulated to enter mitosis with phytohemagglutinin (a plant-derived mitogen). The dividing cells are treated with colchicine to arrest them in metaphase, when the chrome somes are highly coiled and visible. Lysis of the cells with a hypotonic solution causes the chromosomes to spread out on the slide with little or no overlapping. The chromosome spread is photographed, and pictures of the chromosomes are cut out, paired, and assembled into a specific sequence. Karyotyping allows cataloging of chromosomes for detection of structural abnormalities and deleted or excess chromosomes.

D. Nucleolus: During interphase (between mitoses), each nucleus usually has at least one in tensely basophilic body called a nucleolus. Nucleoli are the synthesis sites for most ribosomal RNA (rRNA). They are usually distinguishable from heterochromatin but may be obscured in very dark nuclei. They are largest and most numerous in embryonic cells, in cells actively synthesizing proteins, and in rapidly growing malignant tumor cells. A small amount of hetero chromatin is attached to the nucleolus; the significance of this nucleolus-associated chromatin is unknown. The nucleolus disappears in preparation for mitosis and reappears after mitosis is completed. Distinct nucleolar components can be seen with the electron microscope.


1. The pars amorpha is the pale-staining nucleolar region containing nucleolar organizer DNA, which carries the code for rRNA. Five chromosome pairs have nucleolar organizer regions in humans. Thus, up to 10 nucleoli per cell are possible, but fusion of the organizers into fewer, larger nucleoli is more common. Newly synthesized rRNA appears first in this region.
2. The term nucleolonema is used by light microscopists to refer to a threadlike basophilic substructure of the nucleolus. The nucleolonema contains Z rRNA-rich components distin guishable by EM. a. The pars fibrosa consists of densely packed ribonucleoprotein fibers, 5- 10 nm in diameter. These fibers consist of the newly synthesized primary transcripts of the rRNA genes and associated proteins. Newly synthesized rRNA appears second in this region. b. The pars granulosa contains dense granules, 15-20 nm in diameter, that represent maturing ribosomal subunits during assembly for export to the cytoplasm. Newly synthe sized rRNA appears third in this region.

E. Nucleoplasm: The nucleoplasm is the matrix in which the other intranuclear components are embedded. It is composed of enzymatic and nonenzymatic proteins, metabolites, ions, and water. It includes the nuclear matrix--a fibrillar "nucleoskeletal" structure that appears to bind certain hormone receptors-and newly synthesized DNA.

V. CELL FUNCTIONS

Cells are both empowered and constrained by their available resources. The amounts and types of energy and raw materials at their disposal, the information encoded in their genes, and the intrinsic and extrinsic factors that control their access to that information are the major determinants of cell function. Cells in tissues undergoing growth or repair use a large part of their resources in preparing for, and carrying out, cell division. Fully differentiated cells typically concentrate on more specialized functions such as secretion and contraction. Maintaining a constant internal environment (homeostasis) even in apparently quiescent cells, requires the expenditure of significant amounts of energy and other resources.

A. Cellular Reproduction: The reproductive cycle of a cell is termed the cell cycle. Each complete cycle ends with cell division (mitosis) and results in the production of 2 daughter cells, each about half the size of the parent.


1. Mitosis and interphase. Early views of cellular reproduction focused on easily detected structural changes during mitosis. The apparently inactive phase between successive mitoses seemed a resting period and was dubbed interphase. Yet, even in rapidly dividing cells, the time spent in mitosis is very brief compared with that spent in interphase. We now Lnow that cells cany out many important activities during interphase, including those needed to recover from the previous, and to prepare for the next, mitosis. Both mitosis and interphase are now viewed as complex and important components of the cell cycle and have each been divided into a series of steps to facilitate our understanding of cellular reproduction.
2. The steps in cell division (mitosis). Mitosis is a brief, continuous process. Structural changes observed during this complex process have been used to divide it into 4 successive phases: prophase, metaphase, anaphase, and telophase. a. During prophase, the chromatin coils extensively to form chromosomes. As the nucleolar organizer DNA is coiled into its respective chromosomes, the nucleoli disperse and begin to disappear. The nuclear membrane remains intact. The 2 pairs of centrioles migrate to opposite poles of the cell, and the mitotic spindle apparatus begins to assemble between the centriole pairs. b. During metaphase, the nucleolus and the nuclear envelope disappear. The chromosomes line up at the cell equator between the centriole pairs, and each chromosome splits lengthwise to form a pair of sister chromatids, Each chromosome has a centromere (kinetochore) to which microtubules of the spindle apparatus attach. c. During anaphase, sister chromatids separate and move to opposite poles of the now elliptical cell along the mitotic spindle. The centromere leads, with the chromatin drag ging behind, often in a V shape. The separation mechanism is not fully understood, but the chromatids appear to be translocated by the spindle microtubules. d. During telophase, the chromosomes begin to uncoil. Nucleoli and nuclear envelopes appear as components of 2 separate nuclei at opposite ends of the cell. A purse-string constriction, formed by bands of microfilaments beneath the plasma membrane, appears at the equator. Tightening of the constriction eventually divides the cytoplasm and organelles between the daughter cells.

B. The Cell Cycle: The cell cycle model takes into account important but less visible changes in the cell between divisions. It retains the 4 phases of mitosis but focuses on the timing of DNA synthesis and divides interphase into 3 phases; G,, S, and G,. 1. The G, (gap 1) phase of interphase follows telophase of mitosis. A gap is a period during which no DNA synthesis occurs, as shown by the fact that no radiolabeled thymidine (3H thymidine) is incorporated into the cell's DNA. RNA and protein syntheses do occur during the gap phases, and each daughter cell grows to about the size of the parent. G, usually the longest phase of the cycle, is also the most variable in length among different cell types. In rapidly dividing cells, eg, embryonic and neoplastic cells, G, is short and the transition to subsequent phases is continuous. More highly differentiated cells may pull out of the cycle and enter a phase called G,, in which preparations for mitosis are suspended in favor of specialized functions. G, cells unable to reenter the cycle leg, muscle, nerve) are terminally differentiated. Other cells in G, leg, hepatocytes, fibroblasts) can reenter the cell cycle in response to injury.

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