Tissue engineering in orthopaedics

Tissue Engineering in Orthopaedics

Alexander M. Tatara, BS; Antonios G. Mikos, PhD

J Bone Joint Surg Am, 2016 Jul 06; 98 (13): 1132 -1139 . http://dx.doi.org/10.2106/JBJS.16.00299

Investigation performed at the Department of Bioengineering, Rice University, Houston, Texas

A Journal Club presentation at Amala Institute of Medical Sciences by

Dr. Libin Thomas Manathara

Topics

• Principles of Orthopaedic Tissue Engineering• Scaffolds

• Signals

• Cells

• Applications• Fracture Nonunion

• Osteonecrosis

• Chondral and Osteochondral Defects

• Future Directions and Challenges

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Tissue Enigineering

• Tissue engineering is the use of a combination of cells, engineeringand materials methods, and suitable biochemical andphysicochemical factors to improve or replace biological tissues

• Tissue engineering involves the use of a scaffold for the formation ofnew viable tissue for a medical purpose

• The term regenerative medicine is often used synonymously withtissue engineering

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Principles of Orthopaedic Tissue Engineering

• The tissue engineering paradigm consists of scaffolds, signals, andcells

• These 3 elements can be combined or used independently to attemptto generate tissues in a limitless number of arrangements

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The orthopaedic tissue engineering paradigm.

Alexander M. Tatara, and Antonios G. Mikos J Bone Joint Surg Am 2016;98:1132-1139

©2016 by The Journal of Bone and Joint Surgery, Inc.6

Scaffolds

Scaffolds

• Tissue engineering scaffolds are cytocompatible biomaterials thatcells can adhere to and/or replace with extracellular matrix toproduce native tissues

• Scaffolds can be as simple as morselized autologous bone or ascomplex as injectable, thermally responsive, synthetic hydrogelscapable of mineralizing in situ

• morselize- To break up or divide into small portions

• hydrogel- a gel in which the liquid component is water

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Scaffolds

• On the basis of material composition, scaffolds can be divided into 3basic classes: metals, ceramics, and polymers

• Scaffolds can be further divided by their source (naturally derivedversus synthetically fabricated) and ability to degrade (nonresorbableversus resorbable)

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Overview of the classes of scaffolds.

Alexander M. Tatara, and Antonios G. Mikos J Bone Joint Surg Am 2016;98:1132-1139

©2016 by The Journal of Bone and Joint Surgery, Inc.10

Scaffolds

• Although they are stable, nonresorbable scaffolds and deliverysystems cannot be replaced by native tissues and may elicit a chronicforeign-body reaction detrimental to tissue healing

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Scaffolds

• Naturally derived scaffolds (such as those made from collagen,chitosan, and hyaluronan) are generally all resorbable in situ andoften already possess adhesion ligands for cellular attachment

• However, naturally derived scaffolds typically have a narrow range ofavailable physical properties, such as mechanical strength anddegradation rate

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Scaffolds

• Synthetic scaffolds can be tuned to have a wide variety of propertiesby altering synthesis components and parameters

• Not all synthetic scaffolds are biodegradable, and cell adhesion motifsmay need to be added in order to promote biocompatibility

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Scaffolds

• Different scaffold materials can also be combined to create compositescaffolds that have novel properties not observed in either materialused alone

• For orthopaedics, mechanical properties and durability areparamount to a successful device.

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Scaffolds

• While a ceramic scaffold may have appropriate compressive strengthin a femoral nonunion defect, its high stiffness and weak tensileproperties would be inappropriate for use in a cartilage defect orrepairing a rotator cuff.

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Scaffolds

• Another important factor to consider is the compatibility of the rateof scaffold resorption with the rate of native tissue replacement.

• If a scaffold resorbs too rapidly, it may not be able to support thegrowth of new tissues.

• If a scaffold resorbs too slowly, it may not fully integrate withsurrounding tissues and may pose risks associated with chronicforeign bodies.

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Scaffolds

• Scaffolds may be osteoconductive (i.e., permit the growth of bone,such as the calcium sulfates) or osteoinductive (i.e., actively promotebone growth in a defect that otherwise would not heal, such asdemineralized bone matrix).

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Signals

Signals

• rhBMP-2

• Platelet rich plasma (PRP)

• Antibiotics

• Mechanical cues

• Electrical cues

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Signals

• Signals are internally or externally derived environmental factors thatcan influence the regeneration of tissues

• As in the case of scaffolds, these signals can be further broken downinto subcategories including biological, chemical, mechanical, andelectrical cues

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Signals

• In orthopaedics, the most commonly utilized biological signal isrhBMP-2, a potent osteogenic growth factor

• Has been approved for specific uses by US FDA

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Signals

• However, it appears that rhBMP-2 is associated with greater risksthan originally reported

• There remain concerns about the use of growth factors in light oftheir association with malignancies such as osteosarcoma

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Signals

• Another common source of biological signals used in tissueengineering strategies is platelet rich plasma (PRP) and its differentvariants

• While use of PRP is appealing because of the ease of autologousobtainment, its actual efficacy in the regeneration of musculoskeletaltissues currently remains uncertain

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Signals

• Antibiotics are another example of chemical cues often delivered inconjunction with tissue engineering strategies

• In an infected orthopaedic defect, encapsulating antibiotics for localdrug delivery into the scaffold may be required to stimulate healing

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Signals

• Mechanical cues have long been used to stimulate bone formation,such as in distraction osteogenesis

• Recent studies have demonstrated that passive mechanical signalsprovided by scaffolds (such as substrate stiffness, roughness, andporosity) can influence the differentiation of stem cells towardspecific lineages

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Signals

• Electrical cues have been demonstrated to be important in generatingfunctional skeletal muscle tissue as well as innervation of neotissuesbut have not been explored as thoroughly relative to other cellularsignals for orthopaedic tissue engineering applications

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Cells

• In order to create living tissues, as well as integrate living engineeredtissue with native host tissues, cells must be present

• Cells can be recruited into an implanted scaffold by methods such as• the release of chemokines

• attachment of cell ligands to the scaffold

• osteoconduction

• osteoinduction

• scaffolds containing cells can be implanted into a defect

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Cells

• Unlike in other tissue engineering fields, there is little controversyregarding stem cell type in orthopaedic tissue engineering

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Cells

• By far, the most commonly utilized cell type is the mesenchymal stemcell (MSC)

• Depending on their environment, MSCs have the ability todifferentiate into

• osteoblasts

• chondroblasts

• myoblasts

• tenocytes

• as well as other adult cells

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Cells

• MSCs are relatively easy to harvest from an autologous host

• The current gold standard is MSCs harvested from bone marrowaspirate

• However, adipose-derived MSCs are gaining more traction in the fieldbecause of their

• increased availability

• lower harvesting costs

• ease of expansion

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Cells

• Amniotic fluid-derived MSCs are another intriguing source that hasrecently been shown to be capable of osteogenesis andchondrogenesis in small animal models

• Other sources of MSCs include skin, periosteum, and umbilical cordblood

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Cells

• However, ease of collection should not be the only consideration inMSC harvesting

• MSCs from different sources have different potential fordifferentiation and may affect the quality of tissue repair

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Cells

• Initially, it was believed that the primary mechanism of action ofimplanted MSCs within an orthopaedic defect was structural

• It was assumed that the implanted MSCs themselves wouldproliferate, differentiate, and generate the extracellular matrixrequired to repair the defect

• However, recent studies have revealed the tremendous pleiotropiceffects of MSCs

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Cells

• Beyond their ability to terminally differentiate into adultmusculoskeletal cells, MSCs secrete a variety of cytokines andmodulate inflammatory and immune response pathways

• Because of these immunomodulatory effects, implantation ofallogeneic MSCs carries minimal risk of rejection by the host andcommercially available MSCs are being explored in a number ofclinical trials for a multitude of autoimmune diseases

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Cells

• From a product development and regulatory standpoint, acellularstrategies present advantages over cell-containing products, whichcarry a potential risk of rejection or disease transmission, haveheterogeneous cell populations, etc

• For example, it is clear that rhBMP-2 combined with a collagenscaffold has ample regenerative capacity for spinal fusion without theaddition of any exogenous MSCs

• However, for complex diseases such as osteonecrosis, the pleiotropicand immunomodulatory capacities of MSCs may be required fortreatment

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Orthopaedic Tissue Engineering Elements

Applied in Recent Clinical Experiences

Fracture Nonunion

Fracture Nonunion

• While there is currently no standardized definition, fracture nonunionhas been defined by the FDA and others as incomplete healing at 9months after injury and the absence of healing progression over thefollowing 3 consecutive months

• Despite advances in surgical techniques, fracture nonunions continueto present clinical challenges

• As disrupted vascularity is one of the major contributors to nonunion,strategies to enhance angiogenesis, including delivery ofhematopoietic stem cells, are being explored

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Fracture Nonunion

• In one study, autologous bone marrow-derived hematopoietic stemcells were delivered on an atelocollagen scaffold into tibial andfemoral nonunion defects in 7 patients

• This combination of cells and scaffold resulted in fracture-healing in 5(71%) of 7 patients at 12 weeks

• While there was no control arm, the threshold of healing achievedwas 18% (2 of 11 patients)

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Atellocollagen

• Collagen is a major connective tissue protein that plays an importantrole in the extracellular matrix in animals.

• As such, collagen possesses good biocompatibility with animal bodytissues.

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Atellocollagen

• Atelocollagen is a type of soluble collagen produced fromtropocollagen, the collagen molecule that makes up collagen fibrils,via the elimination of the telopeptide moieties, which are consideredto account for most of collagen's antigenicity.

• Thus, atelocollagen is considered to have little immunogenicity, whichmakes it a safe biomaterial.

• It is widely used for implantable medical and plastic surgical products

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Fracture Nonunion

• In a retrospective review of 52 patients treated for forearm nonuniondefects at a single center, patients were categorized as being treatedwith a single tissue engineering element (MSCs, a scaffold, or BMP,i.e., “monotherapy”) or with all 3 elements of the tissue engineeringparadigm (“polytherapy”)

• With a minimum follow-up time of 1 year, patients receiving all 3elements had significantly improved radiographic healing, clinicalsuccess criteria, and rapidity of healing compared with patientstreated with monotherapy

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Fracture Nonunion

• While that study lends support to the synergistic nature of combiningcells, scaffolds, and signals to treat severe orthopaedic defects, arandomized prospective study would have provided strongerevidence by minimizing bias

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Osteonecrosis

Osteonecrosis

• Given their necrotic nature, the lesions associated with osteonecrosishave poor innate regenerative capacity with few or no viable MSCs

• In addition, it has been reported that, in corticosteroid-inducedosteonecrosis, there is a global decrease in available MSCs

• In theory, MSC therapy may be promising, given theimmunomodulatory properties of MSCs as well as their ability tosecrete angiogenic growth factors and recruit local vasculature

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Osteonecrosis

• In a recent clinical study of 5 femoral heads in 4 patients, autologousbone marrow-derived MSCs were seeded onto morselized allogeneicbone as scaffold and were implanted in the defect after coredecompression

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Osteonecrosis

• Unfortunately, in the patient who received treatment bilaterally,disease progressed in both femoral heads, leading to bilateral totalhip replacement (after 13 and 19 months, respectively)

• Microcomputed tomography revealed that the treated zone wasgreater in opacity than trabecular bone

• The tissue in the treated zone was histologically and mechanicallyindistinguishable from the patient’s healthy trabecular bone

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Osteonecrosis

• The other 3 patients had no more disease progression after treatment(22 to 44 months of follow-up)

• As the authors noted, “Further clinical trials are necessary, includingcomparison to concurrent therapies”

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Osteonecrosis

• In a similar study in which cells and scaffolds were utilized in diseasetreatment, 10 patients with osteonecrosis were treated withvascularized bone grafts augmented with autologous bone marrow-derived MSCs seeded on a β-tricalcium phosphate scaffold

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Osteonecrosis

• At 2 years of follow-up, 9 patients had completed the protocol and 7had no further disease progression

• In the 2 patients who had disease progression, the authors postulatedthat “an imbalance between bone resorption and formation” mayhave contributed to the cystic lesions observed in their femoral heads1 year after surgery

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Osteonecrosis

• Given the degradation rate of β-tricalcium phosphate (weeks tomonths) and the low innate regenerative potential of the femoralhead in osteonecrosis, a mismatch in the rate of scaffold degradationand native tissue regeneration may have resulted in collapse of thelesion

• While a prospective randomized study is required for conclusiveevidence on treatment superiority, these studies in sum demonstratehow selection of scaffold properties may impact the clinical outcome

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Chondral and Osteochondral Defects


• Cell monotherapy for the treatment of cartilage defects is morecommon than in other orthopaedic pathologies

• Despite the popularity of these monotherapies, the optimal amountof MSCs required for efficacy is unclear

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• In a recent study, autologous adipose-derived MSCs in 3 differentdoses were injected without scaffold or exogenous signals intoosteoarthritic knees in 18 patients

• While no patient experienced treatment-related adverse events, theeffects of dose are more difficult to ascertain as the low and mediumdosage groups had only 3 patients each

• However, patients in the high dosage group demonstratedsignificantly improved clinical, radiographic, and arthroscopicmeasurements compared with baseline, and these improvementswere not reflected in the low and medium dosage groups

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• Increasing in complexity, allogeneic chondrocytes that have beengenetically modified to express transforming growth factor-beta 1(TGF-β1) by retroviral modification ex vivo were injected at 2 differentconcentrations in 27 patients with osteoarthritis

• While the 2 groups did not show significant differences in healingcompared with one another, both groups demonstrated significantlyimproved clinical scores (including reduced pain and stiffness andincreased physical function) compared with baseline values

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• This recent work demonstrates that cells can be programmed todeliver signals themselves to mitigate disease in human patients,rather than requiring exogenous delivery of the signals through non-living vehicles such as collagen sponges or microparticles

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• A composite scaffold, constructed using collagen and hydroxyapatitenanoparticles in a gradient-based fashion, was implemented in 27patients

• Again, patients showed significant improvement compared withbaseline values at both 2 and 5 years following treatment, which ishistorically not the case with osteochondral defects

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• While this improvement could be due to the gradient-based nature ofthe acellular scaffold, without a nongradient-based control, it isdifficult to draw any definitive conclusions regarding the effects ofacellular gradient-based scaffolds on the treatment of osteochondraldefects in human disease

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Future Directions and Challenges


• Although the future for orthopaedic tissue engineering is bright, thereis much work to be done

• While the small number of clinical studies reviewed in the presentwork may not necessarily be a representative sample of the field as awhole, they demonstrate that small patient numbers, lack ofrandomization, and absence of control groups consisting of currentclinical treatment standards can make it difficult to determine theefficacy of the various elements of the tissue engineering paradigm inorthopaedics

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• New manufacturing techniques, such as the advent of high-resolutionbioprinting are allowing for the rapid creation of personalized devicesto an extent that was not previously possible

• Infections, which often plagued implantation of foreign materials likescaffolds, are being mitigated by advances in anti-infective strategiessuch as biofilm-repulsing surfaces and antibiotic-delivering materials

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• Synthetic biology and genetic engineering techniques are beingutilized to reprogram cells to optimize healing

• Ultimately, products such as INFUSE (Medtronic) and Carticel havedemonstrated that the field of orthopaedics is willing to adopt tissueengineering strategies into clinical practice

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• In the past, the benchmark for successful treatment of conditionssuch as osteoarthritis was lack of disease progression

• With the new therapies inspired by tissue engineering, the newbenchmark may be the absence of disease

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THANK YOU

Tissue engineering in orthopaedics

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