Lecture 13 - BioE10: Lecture 13 Professor Irina Conboy...

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Unformatted text preview: BioE10: Lecture 13 Professor Irina Conboy Tissue engineering/Regenerative Medicine Immune System, Part I Objectives: Biomaterials (whether containing cells or not) will be recognized by the immune system of the host: ideal biomaterial does not cause an immune response to any of the components. p y p Biological fundamentals: Immune response, tolerance, antibody, T cell, B cell, Antigen presenting cells; MHC How to deliver Gene X and its functionality to the wounded tissue? Smart biomaterials: Combination of matrix, cells and biologically active factors (gene X, DNA, siRNA, etc.) Engineered cells of future tissue Biomaterial with a combination of protein factors, plasmid, viral vectors, etc. t t Orchestrated tissue repair and maintenance, Neo-organogenesis. Considerations: immune-rejection, inflammation, blood-clotting (structural components and cells); timing/duration and place or the response; (need to overlap with tissue regeneration in space and time) Synthetic biomaterials are typically: N-(2N (2 hydroxypropyl)-methacrylamide (HPMA) or PEG or gel-like interpenetrating polymeric network (IPN) of poly(acrylamide) and poly(ethylene glycol general name is hydrogels, hydrogels since they have high and variable water content) . [–CH2–CH2–O–]–monomer for PEG; Natural structural biomaterial are ECM proteins, which surround cells and are secreted by cells, e.g.: laminin, collagen, y g g fibronectin. Bioreactor Liquid N2 Syngeneic engineered cells expressing chimeric receptor X could b generated hi i t ld be t d and banked for everyone (and used as needed) The Immune System = tolerating self and destroying non-self: Considerations for Tissue Engineering and Regenerative Medicine •People who acquire bacterial or viral infections and recover often develop immunity to that disease—they do not contract the same disease in the future. Immunization is the conferring of immunity to a particular disease. •In the late 1700s Edward Jenner had a key insight 1700s, about why milkmaids often avoided smallpox infections. Because cows suffered from a smallpox-like disease called cowpox, Jenner reasoned th t milkmaids were ll d J d that ilk id immune (resistant) to smallpox because they had been p p exposed to cowpox. To test his hypothesis, he injected a boy with fluid taken from a cowpox pustule and then later injected the same child with fluid from a smallpox pustule—a procedure called inoculation. As predicted, the boy did not contract smallpo Jenner’s technique was q ickl adopted thro gho t smallpox. techniq e as quickly throughout Europe and was later named vaccination. • Innate immunity refers to immune system cells that are ready to respond to foreign invaders at all y p g times. The innate immune system is nonspecific and responds in the same way to all antigens. •Cells in the innate system have proteins on their plasma membranes that bind to certain bacteriabacteria specific compounds (bacteria cell wall parts, such as LPS). These proteins are known as patternrecognition receptors. When they are activated=bound by antigens, the leukocytes respond. d • An antigen is any foreign molecule that can initiate an immune response response. Hematopoietic lineage Passague, et al HSCs can be divided into LT-HSCs, highly self-renewing cells that reconstitute an animal for its entire life span, or ST-HSCs, which reconstitute the animal for a limited period. ST-HSCs diff tit t th i lf li it d i d ST HSC differentiate into ti t i t MPPs, which do not or briefly self-renew, and have the ability to differentiate into oligolineage-restricted progenitorsthat ultimately give rise to d e e t ated p oge y t oug functionally irreversible maturation differentiated progeny through u ct o a y e e s b e atu at o steps. The CLPs give rise to T lymphocytes, B lymphocytes, and natural killer (NK) cells. The CMPs give rise to GMPs, which then differentiate into monocytes macrophages and granulocytes, and to megakaryoticerythroid progenitors (MEP), which produce megakaryocytesplatelets and (MEP) erythrocytes. Both CMPs and CLPs can give rise to dendritic cells. All of these stem and progenitor populations are separable as pure populations by using cell surface markers. y g Passague, et al g Environment of a degenerating organ is likely to be inflammatory. Innate immune response Q: Could this response occur to HPMA or other synthetic material? Innate immune response Pathogen-associated molecular patterns (LPS, high mannose) ; apoptotic cell-associated molecular patterns (phosphatidil-serine on the outer side of plasma membrane) provide “eat me” signals to promote the safe disposal of l b ) id “ t ” i l t t th f di l f the intruders by professional and amateur phagocytes. Pattern recognition receptors)control phagocytosisand associated inflammatory response depending on the meaning of these signals. Importantly, in order to avoid p g g g p y, excessive collateral damage of surrounding cells, it is increasingly evident that self-associated molecular patterns (CD55) signal a robust antiinflammatory response and promote tissue repair. Molecular Immunology:40 (2003) 85-94 Q: will this system target a biomaterial of an engineered transplant? Q: how would a bioengineer use (A) “eat me” molecule and (B) ”do not eat me” molecule as building me modules of advanced biomaterials? The acquired immune responses are based on interactions between specific immune system cells and specific pieces of antigens. The cells involved in the acquired immune response are called lymphocytes. T cells come in two major types that are differentiated by distinctive proteins on their plasma membranes. These membrane proteins are called CD4 (secrete cytokines, such as TNF-alpha)and CD8 (make holes in membranes of other cells). B cells can be naïve (before antigen encounter), .plasma (secrete antibodies) or memory (after antigen encounter) Natural Killer (NK) cells recognize and destroy cells wihtout MHC molecules, for example, cells infected by a virus that down-regulated the expression of MHC genes. Clonal selection theory:1. Each lymphocyte formed in the bone marrow or thymus recognizes one antigen.2. The lymphocyte is activated when it binds to its specific antigen.3. Some of the cloned cells descended from an activated lymphocyte persist after the pathogen is eliminated f ti t d l h t i t ft th th i li i t d and allow a rapid response if the infection recurs. •The B-cell receptor (or Ig or antibody) has two identical light chains (a protein with a molecular weight of about 25,000 daltons), two identical heavy chains that are about twice the size of the light chains, and a transmembrane domain domain— a structure that anchors the protein in the plasma membrane of the B cell. • The T-cell receptor (TCR) binds to antigens only after they’ve been processed by other immune system cells and presented (displayed) on their cell membranes. The TCR is composed of a single alpha chain and a single beta chain and has a shape similar to the arm of the BCR. Q: Mice were immunized with proteins from cancer cells and the activated B and T cells were p isolated. How would you identify the variable BCR and TCR regions? How would you use this knowledge to organize an immune response against tumor? Structures of BCR and TCR genes differ in immature vs. mature cells Orange-Rag proteins; Triangles-RRS=recombination recognition sequences; Extra E t nucleotides are added at the l tid dd d t th ends of coding regions-> more variability.SCID=no Rag activity; Q: What is the genomic organization of V(D)J regions in neurons, muscle and other adult tissues and why? How to prove that Rag proteins are necessary and sufficient for the recombination of V(D)J genes? Somatic hypermutation (in lymph nodes after antigen encounter) Memory B cells encounter). cells. After repetitive introduction of a biomaterial a much enhanced immune response will occur. •Antibodies, BCRs, and TCRs do not bind to the entire antigen but to a g selected region of the antigen called an epitope. Figure illustrates a protein called hemagglutinin, which is found on the surface of the influenza virus. This antigen has six epitopes, each of which is recognized by a particular antibody, B-cell receptor, or T-cell receptor. p , p Epitopes for BCR/Ig can be parts of proteins, DNA, small molecules (chemicals), lipids, sugars, etc.=virtually any compound in the Universe; BCR/Ig recognizes 3D and denatured proteins; epitopes for TCR are parts of proteins and a processed by APC 3D structure is recognized. recognized Western blot, immunochemistry and Chip assays are based on the Ig-protein recognition. Circulating and newly formed antibodies will recognize most components of most biomaterials as foreign antigens, thus initiating an immune response. MHC Class I – intrinsic cellproduced peptides MHC Class II – extrinsic processed peptides The human major histocompatibility complex lies on chromosome 6. The MHC contains more than 200 genes (encoding proteins involved in binding and presentation of the peptide degradation products of proteins to the T cell antigen receptor) receptor). Q: Will viral proteins from an infected cell be presented by MHC class I or MHC class II? Will parts of hydrogels be presented by MHC class I or MHC class II? 3. Complement system activation (making holes in membranes of cells) Q: What would happen if viruses or bacteria are used as nano-tools in vivo? How would you modify such viruses and bacteria for the in vivo applications? What would happen to artificial tissue implants containing progenitor cells in vivo? What materials are best to choose for tissue engineering: synthetic=plastic, chemical or metal-based or natural extracellular matrix-based and why? Q: Why is constant immuno-suppression required for long-tern or especially secondary tissue and organ transplants? Why is this relevant for tissue engineering? Self-tolerance or avoiding self-destruction Multiple sclerosis is an autoimmune disease: T cells specific for myelin basic protein; Type I diabetes is an autoimmune disease: T cells specific for pancreatic beta cell epitopes •If a B-cell receptor or T-cell receptor responded to a self molecule—a molecule belonging to the host—the receptor would trigger an immune response. Immune system cells would begin to destroy parts of the host's own body. An anti-self reaction such as this is known as autoimmunity. At the periphery, prolonged/hyper-active TCR signaling leads to functional inactivation of T-cells (anergy) and cancer cells are known to induce such a state of immune inactivity in lymphocytes that could potentially eliminate cancer. cancer Q: Can HLA(MHC) matched tissue or cells cause an immune response? If yes, by what mechanism? Natural killer cells: do you have an MHC class I? Q: Can we eliminate immune response by mutating/inactivating MHC? Conlusions: Immune response will likely occur to engineered tissues: both to cells and biomaterials; Optimal O ti l engineered tissue would be compatible with i d ti ld b tibl ith physiological tissue with respect to the cellular components and biochemistry and would not induce an p y immune response. Check your understanding: You should be able to describe the innate and adaptive immune responses and the generation of TCR and BCR repertoire t i You should be able to describe a mechanism of central tolerance and give examples of autoimmune diseases You should understand the fundamental principles of the immune response to bacteria, viruses and to non-self components of engineered tissues. ...
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This note was uploaded on 04/21/2010 for the course BIOE 10 taught by Professor Conboy during the Fall '09 term at University of California, Berkeley.

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