Types of Tissue Engineered Implants in The Body Synthetic | BMES 212, Study notes of Biology

Download Types of Tissue Engineered Implants in The Body Synthetic | BMES 212 and more Study notes Biology in PDF only on Docsity! Tissue Engineering Goal: Replace, repair, or enhance biological function at the tissue or organ level Method: Transplantation of cells in a suitable polymeric matrix or scaffold Implant type: Dictated by the function which the implant is designed to replace Types of Tissue Engineered Implants Type Mode of action Examples Physical Graft performs primarily Bone a biomechanical role Blood vessels Physiological Graft performs a Liver physiological role kidney Chemical Use of cells to produce Pancreatic soluble, diffusible Islets components Combination Graft performs more Skin than one role Autologous Cells  Simplest, by patient biopsy, always compatible  Limited supply  Takes time to harvest, expand, construct implant  Very costly  Variable patient to patient  First application keratinocytes, sheet expanded 2000 fold  Now also chondrocytes for atricular cartilage Cells Recruited from Host  Implant scaffold to promote cell infiltration and differentiation Eventual replacement of scaffold with host-derived ECM Can include growth factors in scaffold (e.g., BMF, bone morphogenic factor) Usually use natural scaffold (e.g., nerves, meniscous, blood vessels Allogenic Cells No supply problems Can be grown up and preserved Reproducible Consistent More cost effective Can produce more complex tissues Need to do extensive quality control General Safety Concerns Autologous  Screen and validate to guard against changes in cells  Prevent introduction of pathogens Allogenic Minimize transmission of infectious agent  Prevent phenotype changes Xenogenic  Prevent introduction of new animal virus Interesting site on TE skin manufacture: http://biomed.brown.edu/Courses/BI108/BI108_2005_Groups/08/index.html Dermagraft Manufacturing Process Maternal Serum Donor Cells HIV-1 &: -2 Mycoplasma HTLV-I EIA USP Sterility CMV and ALT Adenovirus HSV-1 & -2 non-A &: non-B, HIV-1 & -2 Hepatitis Bc HTLV-1 BsAg &: C, Karyology RPR Growth characteristics CMV CMV HSV I & II Tumorigenicity HTLV I & II Hepatitis B&C HIV 1&2 and EBV and HHV-6 Apligraf Manufacturing Process Table 3. Cell Bank Testing” Sterility EBV Wycoplasma CMV In vivo Virus Human papillama In vitro virus HHV 6 and 7 Virus by EM In vivo tumorigenicity Reverse transcriptase Porcine parvovirus HIV 7, cacultivation WIAP test (HEP only) Hiv 1 and 2, PGR Extended bovine assay HTLV 7 and 2 Bovine polyoma Hepatitis A, B, and C-PCR Karyology and isoenzyme * Courtesy of W. Parenteau, Grganogenesis inc, Canton, Mass. EM indicates électran microscopy; AI human immunodeficiency virus; PER, polymerase chain reaction; HTLV human T-cell lymphotrapie virus; EBV Epstein-Barr virus; CMV. cytomegalovirus; HNV human herpesvirus; MAP murine antibody production: and HEP human epidermal cell,  The pluripotent stem cells undergo further specialization into stem cells that are committed to give rise to cells that have a particular function. Examples of this include blood stem cells which give rise to red blood cells, white blood cells and platelets; and skin stem cells that give rise to the various types of skin cells. These more specialized stem cells are called multipotent. Multipotent Stem Cells Extracellular Matrix (ECM) Extarcellular gel-like substance which provides 3D organization to cells and means of communication, control of proliferation, cell migration, attachment, differentiation and repair Major Macromolecular Components Glycosaminoglycans (GAG’s) - form a gel Collagen - provides strength and organization Elastin - provides resilience of elasticity Fibronectin - adhesion of fibroblasts and other cells Laminin - adhesion of cells to basal lamina 3D Tissue Fabrication • Often just scaffold for support + cells is not enough, need to: – Mimic micro architecture of tissue – Mimic microenvironment – Repeating units on scale of microns • Apply CAD-based manufacturing technologies • Fabricate vascular beds which allow for larger tissue constructs no limited by diffusion • Customs build to fit defect using imaging data with CAD Drug Del. Rev. 6:1635 2004. FDA Regulations • Quality program, which would evaluate all aspects of the firm's operations, to ensure compliance with GTP. • Adequate organizational structure and sufficient personnel. • Standard operating procedures for all significant steps in manufacturing. • Facilities, equipment and the environment with control and validation of manufacturing processes. • Adequate and appropriate storage. • Record keeping and management. • Complaint file. • Procedures for tracking the product from donor to recipient, and from recipient to donor. Time Line for Development • 1993: first regulations specifically addressing human tissues - regulations required that tissue donors be tested for certain communicable diseases such as HIV and hepatitis, and screened for behavioral risk factors. • 1997: Proposed Approach to the Regulation of Cellular and Tissue-based Products • 2001: Establishment Registration and Listing for Manufacturers of Human Cellular and Tissue-Based Products requires tissue facilities to register with the FDA and list their products • 2004: Suitability Determination for Donors of Human Cellular and Tissue-Based Products focuses on donor screening and testing requires reporting certain adverse reactions and HCT/P deviations, accurate and complete information in labeling, and to allow FDA inspections to ensure compliance with regulations. • 2005: Current Good Tissue Practice for Human Cell, Tissue, and Cellular and Tissue-Based Establishments; Inspection and Enforcement - requires manufacturers to recover, process, store, label, package, and distribute HCT/Ps in a way that prevents the introduction, transmission, or spread of communicable diseases. Tissue Engineered Constructs  Rapidly evolving field  Only one FDA approved (skin) Current literature good source of the latest advances (e.g., Tissue Engineering, Biomaterials research, Science, Nature) Will give select few examples to show what has been achieved and the complexity of the task Epidermal Component Dermal Component Apligraf Organogenesis Canton, MA Human mature keratinocytes (Stratum Corneum) Human fibroblasts in bovine collagen ge l Dermagraft Smith & Nephew London, U K None Human fibroblasts in PLGA scaffold Orcel Ortec International New York, NY Immature keratinocytes (not confluent, not cornified ) Human fibroblast in bovine collagen spong e Commercial Ventures Dermagraft was acquired in 2006 in the US by Advanced BioHealing Skin (FDA approved product) Dermagraft: newborn human dermal fibroblasts grown on 3D degradable scaffold Cell source human foreskin  Support: 5 x 7.5 cm knitted lactic acid/glycolic acid copolymer Medium, Dulbeco’s modified Eagle medium + bovine calf serum, glutamine ascorbate and non- essential amino acids  Stored frozen at –70oC  Sheet is 100 microns thick Achieved complete healing of diabetic foot ulcers in 12 weeks. Also used for burn victims Histological Comparisons Natural Skin Dermagraft Apligraft Orcel Cartilage (developing)  Chondrocytes extracted from distal femoral joints of immature donors (rabbit and sheep)  Seeded onto 3D porous scaffold (PGA fibers) Grown in convective-flow bioreactor Implanted in medial patellar grooves Wu et al. Annals of New York Academy of Science (1999) 875 405-411 Monolayer of muscle cells detaching from substrate and rolling into a cylinder. Requires several days. Dish diameter 35 mm Approximately 3 days after delamination of the monolayer, the cells have self organized into a cylinder Representative cross section of a myooid, stained with 1% Toluidine blue. scale bar is 100 µm Tissue Engineered Muscle Bundle (myooid) Robert G. Dennis, U. Mich Extracorporial Artificial Liver  70 kg adult has 1.75 kg liver( 2.5% body weight) (1.2 kg of hepatocytes (liver cells) Liver failure in 1989, 30000 deaths, 2160 transplants  Two types of support: Passive and Bioreactors  Passive systems remove toxins, e.g., hemodialysis, hemoperfusion, plasma exchange No improvement, still get hepatic encephalopathy (HE) which leads to hepatic failure (HF) Bioreactors: cell based, Liver Assist Devices (LAD’s) Liver  Damage often progresses in a zonal fashion Regeneration is possible in vivo from hepatocytes at the portal space (candidates for LAD?) Zonal hepatocyte necrosis stimulates fibrous scaring, disrupts blood supply, leads to cirrhosis Blood is shunted to vena cava (can not pass through liver) and toxins build up Functions of Replacement Liver  Regulation of plasma protein production Metabolism and storage of vitamins Biotransformation and Detoxification of drugs (first pass effect) Lipoprotein and cholesterol synthesis Maintenance of glycogenesis and glyconolysis Conversion of heme to bilirubin, and biliverdin Why Do Livers Fail?  Excessive alcohol consumption (major cause)  Hepatitis (B and C produce chronichepatitis)  Cancer  Drugs which give toxic metabolites with P450 enzymes Genetic LAD Configurations  Provide maximum environment for cell viability and function Good transport of nutrients, oxygen toxins and metabolites Various designs (Table 30.2) HepatAssist2000™ Microporous (0.15 micron) polysulfone hollow fibers  Inoculated with rabbit hepatocytes  Perfused with whole blood Clinical evaluation ongoing of the next generation device , LAS, liver assist system (Figure 30.3) Apheresis system separates plasma from blood, feeds plasma to the device containing porcine hepatocytes Also have a charcoal column and an oxygenator HepatAssist The HepatAssist System is an extracorporeal, bioartificial liver support system incorporating primary hepatocytes (pig liver cells), and is designed to treat patients with acute liver failure by temporarily providing essential liver functions. The HepatAssist System includes a hollow fiber bioreactor containing hepatocytes, two charcoal columns, a membrane oxygenator, and a pump (see Figure 2 below). The HepatAssist System is used in combination with a commercially available plasma separation machine, which diffuses plasma through the HepatAssist circuit before it is reconstituted with the blood cells and returned to the patient. Each HepatAssist treatment lasts 6 hours. The hepatocytes are isolated, processed, and cryopreserved in compliance with Good Manufacturing Practices (cGMP). Cryopreserved cells are shipped to and stored at the clinical sites, where the cells are thawed just prior to a patient treatment. http://www.circebio.com/technology/hepatassist.html Current Status • 1997 Dolly born from cloned adult somatic cell • Possible targets for cure – Congenital abnormalities – Cancer – Trauma or organ failure (e.g., kidney) – Infection – Inflammation – Iatrogenic (injuries caused by physician during treatment eg infection) – Age related Advances in Organ Transplantation • Great shortage of organs • First transplant 1955 (kidney) • 1960’s first allogenic transplant (unrelated recipient) • Revolution in transplant surgery and immunosuppressive therapy, 2001 over 23,000 patients received transplants in US alone, but 80,000 are waiting for an organ and 6000 died awaiting an organ Time Line • Cloned frogs 1962 • Lambs 1996 from differentiated epithelial cells • Cattle 1998 • Goats 1999 • Mice 1998 • Pigs 2000 Cloned Kidney! Advanced Cell Technology, Boston Generation of Histocompatible Tissues Using Nuclear Transplantation: Nature Biotechnology, 20 (7), 689-696, 2002. A. Unseeded control B. Seeded with alogenic control cells C. Seeded with cloned cells D. 12 weeks after implantation Renal cells were extracted from 56 day old bovine clones, expanded and seeded onto collagen-coated polycarbonate membranes, and renal constructs created by attaching tubes. Constructs were re-implanted sub Q. MARS MARS®FLUX DIALYZER = diaMARS®ADSORPTION diaFLUX DIALYZER COLUMNS PATIENT BLOOD CIRCUIT MARS®ALBUMIN CIRCUIT DIALYSATE CIRCUIT VitaGen Inc.’s Extracorporeal Liver Assist Device (ELAD®) http://bms.brown.edu/curriculum/b108/liver/vitagenpg.htm Uses HUMAN hepatocytes  Modular Extracorporeal Liver System (MELS) http://cito.charite.net/PressRoom/photos/mels/FrameSet.htm    http://cito.charite.net/PressRoom/photos/hollowfiber/FrameSet.h MELS Diagram taken from: http://cito.charite.net/projects/PROJ8.html Thin hollow fiber membranes, human hepatocytes or liver cells Autologous Bladder • Homologous Decellularised Bladder Submucosa Scaffold • Biodegradable Composite Scaffold – Made of collagen and PGA – Shaped into a bladder with polyglycolic sutures Atala A., Bauer S., Soker S.: Tissue-engineering autologus bladders for patients needing cystoplasty. Lancet 376: 1241-46, 2006 Fig. 3. Electron micrographs of BALB/C mouse fibroblast after 1 day in culture shows cellular spreading across the fiber (left magnification—800, 10 mm bar and right magnification—1500, 10mm bar). Fig. 4. Electron micrographs of rabbit ACL cells after 1 day in culture shows cell migration and attachment along the fibers (left and right magnification—250 and 100 mm bar). Fig. 5. Electron micrographs of BALB/C mouse fibroblast after 8 days in culture shows large cellular networks with cells proliferating with and without the underlying scaffold (left magnification—250, 100 mm bar and right magnification—500, 10mm bar). Fig. 7. Electron micrographs of rabbit ACL cells after 8 days in culture shows cell response to 3-D circular braid, the cells did not cover the whole scaffold but did continue to follow the underlying fibrous geometry (left magnification— 05, 10mm bar and right magnification—1000, 10mm bar).