Structure & Chemistry of Connective Tissue Fibers Lecture Notes PDF

8 Structure & Chemistry of C.T. Fibers ILOs By the end of this lecture, students will be able to 1. Describe distribution and function of CT fibers. 2. Explain synthesis and degradation of collagen. 3. Deduce the structure-function relationship of Collagen and...

8 Structure & Chemistry of C.T. Fibers ILOs By the end of this lecture, students will be able to 1. Describe distribution and function of CT fibers. 2. Explain synthesis and degradation of collagen. 3. Deduce the structure-function relationship of Collagen and Elastin 4. Interpret how biochemical defects in collagen and elastin can impair morphology. Types of CT fibers: Collagen and elastic fibers, the two major fibrous proteins of connective tissue, have distinctive biochemical and mechanical properties as a consequence of their structural characteristics. I- Collagen (Structure and Function)  Collagen is the most abundant protein in the human body representing about 25% of the protein in the body. It is found in tissues with tensile strength and rigidity, where collagen resists tensile forces (stretching or pulling forces) thus providing rigidity to the connective tissue.  It is a long triple helix of peptide chains, known as α-chains. Each individual collagen polypeptide is an α-chain of about 1400 residues. Each α-chain is coded by a separate messenger ribonucleic acid (mRNA).  Every third amino acid is glycine (-Gly-X-Y-) with a very high proportion of proline and lysine in the other two positions.  Proline and glycine, are both important in the formation of the triple-stranded helix. Proline facilitates the formation of the helical conformation of each α chain because its ring structure causes “kinks” in the peptide chain. Glycine, the smallest amino acid, is found in every third position of the polypeptide chain. It fits into the restricted spaces Figure 1: Structure of Collagen where the three chains of the helix come together.  Many proline and lysine residues are hydroxylated to hydroxyproline and hydroxylysine after synthesis of the α-chain (post-translational modification). The extent of lysine hydroxylation of collagen is highly variable in comparison with proline hydroxylation.  Hydroxyproline residues are crucial for collagen folding and stability. Approximately 50% of proline residues in collagen are hydroxylated in various types of collagen in different tissues, and it does not significantly change under physiological conditions. Page 1 of 5  The extent of lysine hydroxylation, however, can vary from 15 to 90% depending on the collagen types and, even within the same type, it varies significantly from tissue to tissue and under the physiological/pathological condition of the tissue. Synthesis and Degradation: (Figure5) - Collagen will spontaneously assemble into fibrils. To avoid premature assembly of fibers within the cell, precursor forms are first synthesized. - The α-chain (preprocollagen) is first targeted to the endoplasmic reticulum (ER) with a signal sequence that is immediately removed in the ER. - Selected proline and lysine residues are then hydroxylated in the ER to form hydroxyproline and hydroxylysine residues. These hydroxylation reactions require molecular oxygen, ferrous iron (Fe2+), and the reducing agent vitamin C (ascorbic acid), without which the hydroxylating enzymes, prolyl hydroxylase and lysyl hydroxylase, are unable to function. - The pro-α-chains spontaneously assemble into procollagen triple helices within the ER. The resulting molecule has propeptide extensions on both ends (carboxy and amino ends), still preventing spontaneous assembly into collagen fibrils. - The procollagen is translocated from the ER into the Golgi apparatus and packaged in secretory vesicles. It is then secreted into the extracellular matrix by exocytosis (fusion with the plasma membrane), and procollagen peptidases remove the propeptide ends - Procollagen then forms units called tropocollagen, which spontaneously assemble into collagen fibrils. Figure 5: Collagen Synthesis Page 2 of 5 - Collagen fibrils are packed regularly to form bundles that appear white glistening in the fresh state. When stained with hematoxylin and eosin, they appear as long, wavy, pink fiber bundles. (Fig 6) - The collagen fibrils are strengthened further by crosslinking between adjacent lysine side chains by the enzyme lysyl oxidase. This is a slow, continuous process throughout an individual’s life. - Collagen can be remodeled by with metalloproteinases. Figure 6: Collagen H&E stained - The action of these digestive enzymes degrade collagen and is balanced by a tissue inhibitor of metalloproteinases (TIMP). - Cross-linking permits scar tissue to strengthen long after a wound has healed, but it also contributes to the decline in collagen quality with aging causing collagen to stiffen, contributing to the decline in vascular elasticity, weakening of the cartilage and the visible signs on the skin, which becomes less firm and supple with age. Clinical Implications: - In the case of ascorbic acid deficiency (and, therefore, a lack of proline and lysine hydroxylation), interchain H-bond formation is impaired, as is formation of a stable triple helix. Additionally, collagen fibrils cannot be cross-linked, greatly decreasing the tensile strength of the assembled fiber. The resulting deficiency disease is known as scurvy. Patients with scurvy often show ecchymoses (bruise-like discolorations) on the limbs as a result of subcutaneous extravasation (leakage) of blood due to capillary fragility - Collagenopathies: Defects in any one of the many steps in collagen fiber synthesis 1- Ehlers-Danlos syndrome:(EDS) is a connective tissue disorder caused by a deficiency of collagen- processing enzymes (as lysyl hydroxylase).The classic form of EDS, is characterized by skin extensibility and joint hypermobility. The vascular form, is the most serious form because it is associated with potentially lethal arterial rupture. 2- Osteogenesis imperfecta: This syndrome, known as “brittle bone disease,” is a genetic disorder characterized by bones that fracture easily, with minor or no trauma. The most common mutations cause the replacement of glycine by amino acids with bulky side chains. The resultant structurally abnormal α chains prevent the formation of the required triple-helical conformation. Phenotypic severity ranges from mild to lethal. Elastic fibers  In contrast to collagen, which forms fibers that are tough and have high tensile strength, elastin is a connective tissue fibrous protein with rubber-like properties.  Elastic fibers provide elasticity to the connective tissue, thus most abundant in areas of the body subjected to volume or pressure changes as lungs, the walls of large arteries, and elastic ligaments. They are highly accommodating and may be stretched one and a half Page 3 of 5  times their resting length without breaking. When the force is released, elastic fibers return to their resting length.  It is rich in proline and lysine but contains scant hydroxyproline and hydroxylysine.  Structural organization; These fibers are usually slender, long, and branching in loose connective tissue (Fig 7A) or form coarser bundles in ligaments and fenestrated sheets (Fig 7B) (discussed later in types of CT).  The core of elastic fibers is composed of elastin and is surrounded by a sheath of microfibrils; each microfibril is about 10 nm in diameter and is composed of the glycoprotein fibrillin. During the formation of elastic fibers, the microfibrils are elaborated first, and the elastin is then deposited in the space surrounded by the microfibrils.   Elastin is an extremely durable and does not turn over appreciably in healthy tissue. It is estimated to have a half-life of about 70 years. However our neutrophils secrete elastase enzyme a powerful protease that is released into the extracellular space and degrades elastin of alveolar walls as well as other structural proteins in a variety of tissues. To counteract this enzyme the liver secretes α1-antitrypsin (AAT), which inhibits a number of proteolytic enzymes including elastase  Patients with inherited defects in α1-antitrypsin are at serious risk for emphysema; intravenous administration of α1-antitrypsin is an effective treatment.  Smoking also can cause emphysema, since a methionine residue in α1-antitrypsin essential for binding to elastase is vulnerable to oxidization by cigarette smoke.  Clinical hint: The integrity of elastic fibers depends on the presence of microfibrils. Patients with Marfan syndrome have a defect in the gene on chromosome 15 that codes for fibrillin; therefore, their elastic fibers do not develop normally. Typically, patients present with tall stature and aortic root dilatation. People who are severely affected with this condition are predisposed to fatal rupture of the aorta. Figure 7. Elastic fibers A B Page 4 of 5  16 Types of immune response: Type I & II hypersensitivity reactions ILOs By the end of this lecture, students will be able to  Distinguish etiology, morphology and pathogenesis of types I and II of hypersensitivity reactions.  Apply types I and II reactions to corresponding clinical conditions as anaphylaxis, allergy, and blood groups incompatibility. Introduction; Abnormal immunological responses comprise the following; o Hypersensitivity Reactions o Autoimmune reaction o Immunodeficiency Hypersensitivity Reactions, A “hyper” or exaggerated response to what should be considered harmless antigens.  Hypersensitivity implies an excessive or harmful reaction to an antigen attributable to imbalance between effector and regulatory mechanisms. It affects individuals who have been previously exposed to an antigen and manifest detectable reaction to that antigen and are therefore said to be sensitized.  Hypersensitivity reactions can be elicited by exogenous environmental antigens (microbial and nonmicrobial) or endogenous self-antigens. The development of hypersensitivity diseases (both allergic and autoimmune) is often associated with the inheritance of particular susceptibility genes.  Hypersensitivity reaction Types I, II, and III are immediate \chemical reactions occurring within 24 hours, whereas, Type IV\ cellular reaction develops over several days.  Classification of Hypersensitivity Reactions Type I hypersensitivity reaction (atopy/ Allergy): - It is a rapidly developing [ immediate type] immunologic reaction occurring after the combination of an antigen with immunoglobulin E (Ig E) bound to mast cells in individuals previously sensitized to the antigen. It is also known as anaphylactic reaction or allergy. - (Allergy): an abnormal adaptive immune response that may or may not involve antigen-specific IgE. 1 - Genetically determined susceptibility to allergic reaction, is called (Atopy), affected patients show increased titer of serum IgE and increased IL-4 secreting TH2 lymphocytes. Patient is usually susceptible to allergic diseases such as allergic rhinitis, asthma, atopic dermatitis and others. Atopy is typically associated with heightened immune responses to common allergens, especially inhaled allergens and food allergens. It also has familial association. - Antigen; Exogenous environmental antigens(allergens) present on the outer surface of epithelial surfaces or skin, such as pollen grains, dandruff, dusts, food components etc. - Antibody; Immunoglobulin E, which is dependent upon the Th-2 helper cells stimuli. - Effector cells; Mast cells in the tissue and basophil in the blood. - Effector chemical mediators; histamine, leukotrienes, prostaglandins, eosinophils and platelets activating factors. Mechanism of Type I hypersensitivity reaction: the reaction occurs in an early and a late phases. Early phase reaction; develop 5–30 minutes after exposure to antigens, initiated by the introduction of an allergen, which stimulates T helper-2 and IgE production in genetically susceptible individuals. IgE binds to Fc receptors on mast cells, this reaction has no clinical manifestation. On subsequent exposure to the allergen, binding of antigen to IgE activates the mast cells to degranulate and secrete the primary mediators [shown in the figure] that are responsible for the pathologic clinical manifestations of immediate hypersensitivity as redness, swelling, edema, increased secretion and smooth muscle contractions. Late-phase reaction; develop 4–12 hours after the early phase and are mediated by eosinophils, neutrophils, and lymphocytes that have been recruited by chemotactic factors released from mast cells. TH2 cytokine IL-5 is the most potent eosinophil-activating cytokine. They cause tissue damage and late- phase inflammatory reaction. Clinical examples: 2  i. Systemic anaphylactic reaction; Anaphylactic shock: a fatal state. Occur in sensitized individuals following injection of foreign proteins (e.g., antisera), hormones, enzymes, polysaccharides, and drugs (e.g., the antibiotic penicillin), exposure to food allergens (e.g., peanuts, shellfish) or insect toxins (e.g., those in bee venom) - Clinically presentation; Itching, hives, and skin erythema, followed shortly by a striking contraction of respiratory bronchioles and respiratory distress, Laryngeal edema results in hoarseness and further compromises breathing. ii. Localized hypersensitivity: often involving epithelial surface at the site of allergen entry; Hay fever (Allergic rhinitis; redness, sneezing, runny nose), bronchial asthma and bronchospasm (allergic or intrinsic), Food allergy (colic and diarrhea), and atopic dermatitis (eczema, urticaria), and allergic conjunctivitis (redness, itching and tearing. N.B; type I reaction plays a defensive role against parasitic infestations. Type II Hypersensitivity Reaction\ cytotoxic hypersensitivity; Antibodies react with antigens present on cell surfaces or in the extracellular matrix cause disease by destroying these cells, triggering inflammation, or interfering with normal functions. Antigen may be: 1. Exogenous : Microbes, parasites, drugs. 2. Endogenous: Autoimmune diseases, and these are self-Ag. Antibody: This is mainly IgG and occasionally IgM. The effector cells are macrophages, neutrophils, eosinophils, and NK (natural killer) cells. Mechanism of Type II hypersensitivity reaction: Binding of antibody (IgM or IgG) to the antigen on the cell surface or other tissue component leading to either 1) cell destruction or 2) altered function Three mechanisms are involved: 1. Opsonization and phagocytosis 2. Complement- and Fc receptor-mediated inflammation 3. Antibody-mediated cellular dysfunction A-Opsonization and phagocytosis; Antibodies coat exogenous\endogenous cell-surface molecules such as antigens associated with blood typing found on red blood cells (RBCs). Coating of the RBCs by antibodies (opsonization), lead to activation of the complement cascade, and complement-mediated lysis of RBCs, as well as opsonization of RBCs for phagocytosis. 3 Clinical Examples; 1. Hemolytic transfusion reaction associated with mismatched blood transfusion reaction. 2. Hemolytic disease of the newborn (erythroblastosis fetalis), in which there is an antigenic difference between the mother (RH -ve) and the fetus (RH+), and IgG anti-erythrocyte antibodies from the mother cross the placenta and cause destruction and hemolysis of fetal red cells. 3. Autoimmune anemia (against RBCs). 4. Drug reaction B- Complement- and Fc receptor-mediated inflammation; antigen-antibody binding leads to activation of complements, recruitment of neutrophils and macrophages mediating an acute inflammatory reaction through the release of proteases and reactive oxygen species. Clinical Examples; transplant organ rejection reaction, skin rash due to drug reaction. C-Antibody-mediated cellular dysfunction; Autoantibodies bind to cell-surface receptors [ recognized as foreign by immune system] to produce an abnormal activation/blockade of the signaling process. Clinical Examples; abnormal activation (Grave’s disease), abnormal blokage : Myasthenia Gravis. 4 Page 5 of 2