Br Med Bull 2011 Sakabe Bmb Ldr025

Musculoskeletal diseasestendon

Tomoya Sakabe, and Takao Sakai,*

Department of Biomedical Engineering, Lerner Research Institute, Cleveland Clinic, Cleveland, OH44195, USA, and Orthopaedic and Rheumatologic Research Center, Cleveland Clinic, Cleveland,OH 44195, USA

Introduction: Tendons establish specific connections between muscles and the

skeleton by transferring contraction forces from skeletal muscle to bone thereby

allowing body movement. Tendon physiology and pathology are heavily

dependent on mechanical stimuli. Tendon injuries clinically represent a serious

and still unresolved problem since damaged tendon tissues heal very slowly

and no surgical treatment can restore a damaged tendon to its normal structural

integrity and mechanical strength. Understanding how mechanical stimuli

regulate tendon tissue homeostasis and regeneration will improve the treatment

of adult tendon injuries that still pose a great challenge in todays medicine.

Source of data: This review summarizes the current status of tendon treatment

and discusses new directions from the point of view of cell-based therapy and

regenerative medicine approach. We searched the available literature using

PubMed for relevant original articles and reviews.

Growing points: Identification of tendon cell markers has enabled us to study

precisely tendon healing and homeostasis. Clinically, tissue engineering for

tendon injuries is an emerging technology comprising elements from the fields

of cellular source, scaffold materials, growth factors/cytokines and gene

delivering systems.

Areas timely for developing research: The clinical settings to establish

appropriate microenvironment for injured tendons with the combination of

these novel cellular- and molecular-based scaffolds will be critical for the

treatment.

Keywords: tendon injury/tissue engineering/regenerative medicine/stem cells/scleraxis/mechanical force

Accepted: May 3, 2011

British Medical Bulletin 2011; 115

DOI:10.1093/bmb/ldr025

& The Author 2011. Published by Oxford University Press.

All rights reserved. For permissions, please e-mail: [email protected]

*Correspondence address:

Department of

Biomedical Engineering,

Lerner Research Institute,

Cleveland Clinic, ND20,

9500 Euclid Avenue,

Cleveland, OH 44195,

USA. E-mail: sakait@ccf.

org

British Medical Bulletin Advance Access published July 4, 2011 by guest on A

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Tendon physiology

Tendon, a fibrous connective tissue made of specialized fibroblastscalled tenocytes and an abundant collagenous extracellular matrix(ECM), is a tissue whose physiology and pathology is heavily depen-dent on mechanical stimuli.1 Tendons establish specific connectionsbetween muscles and the skeleton by transferring contraction forcesfrom skeletal muscle to bone, thereby allowing body movement.2

Tendons exhibit high mechanical strength, good flexibility and anoptimal level of elasticity to perform their unique role. The tensilestrength of a tendon is related to its thickness and collagen content: forexample, a tendon with an area of 1 cm2 is capable of bearing 5001000 kg.3 Tendons have relatively few blood vessels and function at alow metabolic rate. Tendons receive oxygen and nutrients from threemain sources: internally via the myotendinous junction and osteotendi-nous junctions, and externally through the paratenon or the synovialsheath.4

Tendon development and adult homeostasis

During embryonic development, tenocytes originate from mesodermalcompartments, as do skeletal myoblasts, chondrocytes and osteoblasts.5

Some of the multipotent mesenchymal progenitor cells that arise fromthese compartments express the basic helix-loop-helix transcriptionfactor scleraxis. However, once they are committed to become cellsmaking up a specific tissue, only tendon progenitor cells and tenocytesretain the ability to express scleraxis. Therefore, scleraxis is a highlyspecific marker of tenogenic cells and mature differentiated teno-cytes.6,7 The scleraxis gene is thus the first master gene found to beessential for establishing the tendon lineage during development.8

Tenomodulin is a type II transmembrane glycoprotein. Its robustexpression is induced in mouse tendons in a late (embryonic day [E]17.5) developmental phase and is also observed in adult tendonsshowing that tenomodulin is a marker of mature (differentiated) teno-cytes.9 In vitro experimental evidence shows that the genes composedof tendon-specific ECM are tightly regulated in tenocytes by mechan-ical forces.2 Very recently, tendon stem/progenitor cells have been dis-covered in human and mouse tendon, and the proteoglycans biglycanand fibromodulin have been identified as essential elements in a micro-environment to keep phenotypes and differentiation of stem/progenitorcells.10 However, the roles of these stem/progenitor cells in adulttendon homeostasis and/or wound healing remains unknown.

T. Sakabe and T. Sakai

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Mechanical force and tenocytes

Since tendon tissues are constantly exposed to variable transmittalforces from skeletal muscles, our laboratory examined the functionallinks between mechanical forces and tenocyte phenotypes using scler-axis as a tenocyte marker. We utilized a transgenic mouse strain, whichexpresses the green fluorescent protein (GFP) marker driven by thescleraxis gene in a way that GFP is produced in a pattern that mimicsits expression in the body6 (Scleraxis-GFP transgenic mice were kindlyprovided by Dr Ronen Schweitzer, Research Division, ShrinersHospital for Children, Portland, OR, USA). The vast majority of teno-cytes show robust expression of scleraxis-GFP in adult tendon tissuesunder the fluorescence microscope (Fig. 1a). Strikingly, the suddeninterruption of continuous tensile loading, such as by complete transec-tion tendon injury, leads to a decreased expression of scleraxis andtenocyte death (Fig. 1b). Thus, these findings indicate a critical role formechanical forces in adult tendon homeostasis and strongly suggestnew directions for therapy following tendon injuries.11

Tendon injury

Incidence of tendon injury

Soft-tissue injuries, including injury to tendon, ligament or meniscus,can induce abnormal joint motions and altered loading in the shortterm and they could contribute to degenerative joint disease and osteo-arthritis in the long term.12. These injuries can be acute or chronic andare caused by intrinsic or extrinsic factors, either alone or in

Fig. 1 Tensile loading plays a crucial role in tenocytes. (a) Achilles tendons in adultScleraxis-GFP transgenic mice. Left panel: under fluorescence stereomicroscope; rightpanel: under microscope with GFP/ultraviolet (UV) filters to show scleraxis-GFP (green) andnucleus [406-diamidino-2-phenylinodole (DAPI) blue]. AT, Achilles tendon. Bar 100 mm. (b)Analysis of cell death at 2 h after complete transection of adult Achilles tendon inScleraxis-GFP transgenic mice. Arrows indicate the transection edge of tendons. Note thatthe expression of scleraxis-GFP (green) is diminished and cells positive by terminal deoxy-nucleotidyl transferase dUTP nick-end labeling assay (TUNEL assay: a marker for cell death,red) are evident in the transected Achilles tendon. Bar 100 mm.

Strategies for treatment in tendon injury

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combination.4 Acute tendon injury interrupts tendon continuity withconsequent disruption of ECM architecture and dramatic loss of trans-mittal forces from skeletal muscle.4 Tendon injuries represent a seriousand still unresolved problem. More than 130 000 patients per yearundergo tendon-related surgery in the USA.13 The tendons most fre-quently affected are shoulder rotator cuff (51 000 cases), Achillestendon (44 000 cases) and patellar tendon (42 000 cases).13 Injuries toAchilles tendon, patellar tendon, hand flexor tendon and shoulderrotator cuff have clinical importance since they can lead to loss ofmuscle function, significant disability, joint instability and secondaryosteoarthritis, adversely affecting a patients activities of daily livingand quality of life. The incidence of tendon injury has increased inrecent years as the number of aging adults continues to grow.14 Thealtered activity of mechanical loading, and vasculature and angiogen-esis are suggested to play a significant role in degenerative tendondiseases.15,16

Tendon healing

Tendon wound healing involves regeneration of tenocytes and recon-struction of dense collagen fibrils, and the tendon repair process intransected experimental animal tendons is known to involve three over-lapping phases, as for other organs/tissues.4,13. An initial, inflammatoryphase occurs until Day 2 after injury. It involves extensive cell death inthe injured area and subsequent inflammatory cell infiltration. Asecond, proliferative phase starts at Day 3. It involves cell migrationinto the injured area, extensive proliferation and production of collagenfibrils. A third, remodeling phase occurs from 6 weeks on. This phasecan be divided into a consolidation stage, from 6 to 10 weeks afterinjury, and a maturation stage, after 10 weeks. It is characterized bydecreased cellularity and collagen synthesis, and the alignment of teno-cytes and collagen fibrils in the direction of stress. ECM-remodelingduring tendon wound healing follows in general the same processes asin other tissues, i.e. in an early stage, provisional matrix formation bythe plasma proteins fibrinogen and fibronectin, followed by replace-ment of the provisional matrix by collagen fibrils.2,4

In the inflammatory phase, vasoactive and chemotactic factors suchas cytokines and growth factors are released and lead to an increasedvascular permeability, initiation of angiogenesis and stimulation oftenocyte proliferation. In particular, various growth factors/cytokinesplay a number of important roles, including stimulation of tenocyteproliferation, cell migration to the wound and synthesis of the newECM during tendon healing.17,18 In the proliferation phase, two





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mechanisms, intrinsic and extrinsic mechanisms, are likely to contrib-ute to the healing process. The intrinsic mechanism involves the pro-liferation of tenocytes from the tendon and epitenon. These tenocytescontribute to synthesize the new ECM, which consists largely of col-lagens and glycosaminoglycans. Such intrinsic healing results inimproved biomechanics and fewer post-injury complications. In con-trast, the extrinsic mechanism involves the migration of inflammatorycells and fibroblasts from the overlying sheath and synovium into thewounded site.4,19 This often causes scar tissue and results in adhesionformation, which disrupts tendon gliding. Although tendon healingfollows the same process as that in various connective tissues, includingskin and muscle, tendon tissues heal more slowly than other connectivetissues, probably because of the low metabolic rate of tendons andtheir dense and hypocellular composition (only 5% of the volume isoccupied by cells).4,5 Furthermore, ECM remodeling such as increasingdiameters and cross-linking of collagen fibrils occurs over a period ofup to 2 years following tendon injury.3 Despite such remodeling, thebiochemical and mechanical properties of healed tendons never matchthose of intact ones.3 The final tensile strength in the healed tendons iseventually reduced to 30% the strength of intact tendons.20

Therapies in tendon in injury

General concepts in therapies in tendon injury

Clinically, two main strategies for treating injured tendons are fol-lowed:21 (i) leave them untreated to heal naturally or (ii) performsurgery for primary repair of injured tendons using various techniques(Table 1). These treatments are determined in general according toclinical factors, such as age, tendon location, mode of injury and sizeof defect. Surgical treatment of an acute Achilles tendon rupture hasbeen considered superior to non-operative treatment to prevent the riskof re-rupture.22,23 However, investigators in a recent multicenter ran-domized trial (NCT00284648 in ClinicalTrials.gov) reported thatsimilar clinical outcomes are observed between (i) non-operativegroups with accelerated functional rehabilitation and (ii) operativegroups.24 Thus, the treatment for acute Achilles tendon ruptureremains inconclusive.22,24 Furthermore, for injuries to the hand flexortendon or shoulder rotator cuff, long-term clinical outcomes ofprimary repair remain unsatisfactory.13,25 Despite improved surgicaltechniques and advances in postoperative therapy regimens, these treat-ments still have potential complications such as re-rupture of the repairsite or the formation of restrictive adhesions.25,26 Recently, the





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biological grafts, such as autologous fascia, porcine small intestinalsubmucosa or synthetic materials, have been developed and used intendon graft procedures to fill large tissue defects (Table 2). However,no graft method exists to restore a damaged tendon to its normal func-tion. Grafting poses several potential complications, includingincreased inflammatory responses, antigenic reactions, failure at thefixation sites and deficiencies in long-term biocompatibility.25,26 Thus,current knowledge of the biology in tendon healing has yet to lead toclinically successful strategies for treatment.19 New treatment modal-ities are required to promote the regeneration of tendon tissues.

Table 1 Characteristics for treatment in tendon injury.

Treatment type Indication Advantages Disadvantages

Immobilization Acute Achilles tendon

rupture

No surgical complication Failure to heal

Plaster Tendinopathy Rerupture

Brace Muscle atrophy

Tendon atrophy

Limitation of range of motion

Decreasing of tensile strength

Primary repair Most acute tendon

injury

First clinical choice Suture failure

Chronic tear of

rotator cuff

Recovery of muscle

contraction

Rerupture

Inability to achieve functional

results

Postoperative adhesion

Infection

Tendon transfer Large tendon defect Filling of tissue defect Suture failure

Primary repair failure Recovery of muscle

contraction

Rerupture

Chronic tendon injury Inability to achieve functional

results

Postoperative adhesion

Infection

Altered biomechanics of joint

Tendon graft Large tendon defect Filling of tissue defect Suture failure

Autograft Primary repair failure Recovery of muscle

contraction

Rerupture

Allograft Chronic tendon injury Inability to achieve functional

results

Xenograft Destructive disease Postoperative adhesion

Synthetic

material

Congenital disorder Infection

Limited availability of

material

Altered biomechanics of joint

Deficiency of biocompatibility

Immunologic rejection

Antigenic reaction

Disease transmission





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New treatment strategies for tendon healing

To design efficient strategies to enhance tendon repair following injury,it is essential that scientists and clinicians understand the cellular andmolecular mechanisms responsible for the development, homeostasis,regeneration and repair of tendons. To date, however, the molecularmechanisms underlying tendon repair remain largely unknown. Thetarget molecules for tendon repair are based on knowledge fromgeneral wound healing, not on results generated specifically fromanalysis of tendon healing.27

Tissue engineering is an emerging technology that offers a novelapproach for treating tendon injuries and restoring tissue and jointfunction.28 The most common tissue-engineering principles are (i) theuse of healthy multipotent cells that are non-immunogenic, easy toisolate and highly responsive to distinct environmental cues; (ii) thedevelopment of carrier scaffolds that provide short-term mechanicalstability of the transplant as well as a template for spatial growth ofthe regenerating tissue; and (iii) the delivery of growth factors thatdrive the process of cell differentiation and maturation.25

Orthopedic tissue engineering comprises elements from the fields ofcell biology, materials science, mechanical engineering and surgery.12

Various types of scaffolds, such as naturally occurring ECMs as well ascell-based strategies have been developed12,25,26,2933 (Table 2).Mesenchymal stem cells (MSCs) are becoming a subject of increasinginterest because of their potential utility in tissue-engineering appli-cations34,35 (Table 3). MSCs exist in adult bone marrow and can beinduced to form different mesenchymal tissue lineage cells such as

Table 2 Scaffold materials for tendon injury.

Biologic (naturally occurring)

Human tissue Dermis

Dura mater

Animal tissue Porcine small intestinal submucosa

Porcine dermis

Biopolymers Type I collagen

Synthetic (manufactured)

Resorbable Polyethylene

Polyglycolic acid

Polylactic acid

Poly-N-acetyl-D-glucosamine

Non-resorbable Carbon fibers

Dacronw

Nylon

Polyacrylamide

Silicone

Teflonw

Silk





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chondrocytes, adipocytes or osteoblasts. In fact, MSCs-based scaffoldshave been tried in animal models of tendon wound healing.

Cellular scaffold-based therapy

Scaffolds

The underlying concept for tissue engineering technologies has beenchanging. Traditionally, a graft was composed of some material (suchas nylon or silk) meant solely to fill the tissue defect. Nowadays,implants are expected to serve as biocompatible scaffolds (natural orsynthetic materials that can be replaced by host tissues without undesir-able responses). These biocompatible scaffolds are suitable as vehiclesfor implanted cells, the delivery of growth factors, or the transfer of

Table 3 New modalities for treatment in tendon injury.

Treatment type Methods of delivery Advantages Disadvantages

Scaffolds

Biologic material Direct implantation Abundant supply (type I

collagen)

Limited clinical

applications (autograft)

Limited recovery of

strength

Relatively low

complications (type I

collagen)

Potential complications as

in Table 1

Synthetic

material

Direct implantation Abundant supply Limited recovery of

strength

Limited regeneration

Potential complications as

in Table 1

Cell therapy

Tenocyte Direct implantation

with collagen materials

Differentiated cell Limited cellular source

Low proliferative ability

Low morbidity

Mesenchymal

stem cell

Direct implantation

with collagen materials

Excellent regenerative

capacity

Unclearness of

mechanisms in

differentiation

High proliferative ability

Low morbidity

Invasive procurement

procedure

Low yielding

Growth factors Direct administration Easy administration Unclearness of growth

factor stability

Unclearness of effective

concentration

Gene therapy

Viral method Direct viral infection High transduction

efficiency

Inflammatory response

Transient expression Non-specific infection

Non-viral

method

Direct administration

with liposomes

Low pathogenic response Relatively low

transduction efficiency





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genes25,26. Both biologic and synthetic materials are used to create scaf-folds for tendon reconstruction with a three-dimensional biocompatibleconstruct that serves as a temporary or permanent implant26. Asdescribed, injured tendons have very limited spontaneous healing capa-bilities. Thus, ideal scaffold materials need to play at least two essentialroles: to stimulate regeneration (including proliferation and differen-tiation of cells) at implanted sites and to establish the specific compo-sition and structure of an ECM that can then provide an appropriatemicroenvironment for regenerating cells. The major ECM componentin tendons is type I collagen. The advantages of using type I collagenfor tendon reconstruction include its strength, capacity to resorb andability to induce the alignment of host connective tissues.26. Scaffoldsof type I collagen cross-linked with glutaraldehyde or carbodiimide areused in research to regenerate tendon tissue because of the low antige-nicity and strength.26,36 Indeed, they have improved graft strength in arabbit Achilles tendon model.36 Synthetic non-resorbable materials,including nylon, silk and carbon, are not biocompatible because ofhost foreign body responses and late mechanical failure.26 To circum-vent these problems, synthetic resorbable materials have been devel-oped using polyglycolic acid or polylactic acid.25 They can befabricated into three-dimensional scaffolds of variable structure andporosity with a correspondingly wide range of mechanical and degra-dation properties.25 Unfortunately, some synthetic resorbable scaffoldsalter the mechanical properties of the repaired tendon, lose strengthand integrity over time, limit tendon ingrowth, cause abrasions of sur-rounding tissues, enhance the inflammatory response and cause unde-sirable scar formation around the repair site.26 A study in a goatshoulder rotator-cuff repair model indicates that the polylactic acid-scaffold does not show significant increase in the load-to-failurestrength, even though the polylactic acid patch is occupied by cellularfibrous tissues.37 Therefore, despite their potential roles in tendonreconstruction, further investigation will be necessary to find analternative to natural materials.

Cell-based therapyCell-based therapy is also a novel technique to improve the compo-sition, structure and biomechanical properties of new tendon tissue:cells are initially seeded onto scaffolds, and then they are delivered tothe injured sites as cell- and scaffold-combined materials.26 To date,several different combinations of cell types and biomaterial scaffoldshave been used in experimental animal models (including MSCs-type Icollagen gel, MSCs-knitted polylactide-co-glycolide matrix,tenocytes-non-woven polyglycolic acid fibers), and they have thecapacity to enhance tendon formations.3033,38 In these biomaterial





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scaffolds, a plenty of materials such as collagen gel or synthetic biode-gradable polymers are commercially available. On the other hand, cellsseeded on such a scaffold need to proliferate rapidly in vitro to provideadequate numbers for in vivo implantation.25 An important prerequi-site for cell-based therapy is the successful isolation and selection ofappropriate cells.25 A tenocyte-based method is one of the potentialcell-based therapies, but a number of concerns still limit the practicalityof its use: (i) a limited availability of donor sites tenocytes from whichtenocytes may be obtained for implantation, (ii) the time requirementsfor lengthy in vitro culture to expand the number of cells and (iii) themorbidity of tenocytes themselves.39 To circumvent the negativeimpact of this tenocyte-based method, MSCs have been investigated asan alternative source for tendon engineering. MSCs, which show anexcellent capability for regeneration and rapid proliferation, have thepotential to differentiate into a spectrum of specialized mesenchymaltissues, tendon, ligament, bone, cartilage, muscle, fat and marrowstroma.25 In addition, MSCs can be relatively easily isolated from bonemarrow, but they are also found in muscle, adipose tissue, skin andaround blood vessels.40 The ability of MSCs for tendinogenic differen-tiation has been documented in several studies.3133 In fact, recruit-ment of MSCs to accelerate repair and tissue regeneration was shownin vivo in a rabbit tendon tissue model.32 However, no significantdifferences were observed in mechanical properties betweenMSC-transplanted and non-transplanted repaired tissues. Moreover,28% of MSC-treated tendons developed foci of ectopic bone, whereasno bone formed in naturally healing contralateral controls.29,41 Thesestudies clearly indicate that the determination of an appropriate MSCmicroenvironment for tenocyte differentiation is a critical issue thatneeds further investigation. We also need to take into considerationseveral more issues relating to the clinical application of MSC-basedtherapy: long-term safety of the patient, large-scale culture and storageof cells, ideal scaffold materials, optimal cell seeding conditions and analternative mode of applying MSC-material composite to the injuredsite.4,25

Molecular-based therapy

Growth factors and cytokines

Growth factors/cytokines represent one of the largest molecularfamilies involved in the wound healing process, and a considerablenumber of studies have been undertaken in an effort to elucidate theirmany functions and behaviours during healing progression.17 Severalmolecules have been identified as key factors during the repair processof tendons, including transforming growth factor-b (TGF-b), insulin-like growth factor 1 (IGF-1), platelet-derived growth factor (PDGF),





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vascular endothelial growth factor (VEGF), basic fibroblast growthfactor (bFGF) and growth and differentiation factor (GDF)-5through 7.26 Since TGF-b regulates a wide variety of cellular processes,including the expression of scleraxis during tendon formation inembryonic development,42 such multifunctional aspects of TGF-b havebeen extensively studied in relation to adult tendon injury and homeo-stasis. The expression levels of TGF-b in adult tendons are dramaticallyupregulated in a short time after injury, and TGF-b initiates an inflam-matory response to tissue damage.17 In contrast, TGF-b upregulatesthe production of ECMs, which results in excessive scar formation.Indeed, the local administration of a neutralizing antibody of TGF-bcan diminish excessive production of ECM and improve the postopera-tive range of motion in a rabbit model of complete transection of thehand flexor tendon.43 Thus, such contradictory functional aspects ofTGF-b make it difficult to rely on TGF-b for clinical use in tendonhealing.3 IGF-1 stimulates synthesis of DNA, collagen and proteogly-cans, as well as tenocyte proliferation and migration in vitro.44 IGF-1also acts synergistically with PDGF to stimulate tenocyte migration.44

A study in a rat Achilles tendon transection model indicates that theinjection of IGF-1 at injured sites accelerates functional recovery ofAchilles tendon.45 GDF-5, -6 and -7 (members of the TGF-b superfam-ily that are related to bone morphogenetic proteins) can induce neoten-don formation, as assessed by histochemical analysis when injected atsubcutaneous sites in rats.18 Another study shows that the injection ofGDF-5, -6 or -7 into injured Achilles tendons in rats results in a signifi-cant dose-related increase of mechanical properties in rat Achillestendon.46

Some success has been achieved utilizing single growth factors astherapeutics.17 Direct injection of a growth factor at the injured site maygive a temporary boost of a single healing signal but has only limitedeffect on the final outcome.17 The combination of patients own growthfactors to promote healing in injured tissues is a potentially very fruitfularea of research.17 Platelet-rich plasma (PRP), easily harvested fromwhole blood by a few centrifugation steps, contains autologous growthfactors such as PDGF, TGF-b, IGF-1 and -2 and bFGF.47 Postoperativedirect injection of PRP significantly improves mechanical strength andstiffness in a rat Achilles tendon repair model.48 Recently, there has beenincreasing interest in the field of sports medicine to facilitate healing andearlier return to activity after tendon and ligament injury.49 Severalclinical trials investigating the efficacy of PRP treatment have been per-formed for Achilles tendon rupture (NCT00731068 in ClinicalTrials.gov) and rotator cuff injury (NCT01000935; NCT01152658;NCT01170312 in ClinicalTrials.gov). However, recent randomized





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clinical trials indicate that PRP treatment has no significant effect on thehealing of tendon and ligament injuries.49,50

Gene therapy

A variety of gene transfer methods have been used both in vitro and invivo to induce local production of growth factors such as GDF-7 orPDGF by means of viral (e.g. adenovirus, retrovirus) or non-viralvectors (e.g. liposomes) to accelerate the tendon repair process.51,52

The use of a non-viral vector-mediated delivery is less pathogenicbecause of the absence of viral proteins but is also less efficient thanthe use of viral vectors.25 The expression of a transgene is transient butgenerally manipulated for up to 810 weeks: this method is more ben-eficial than repeating local injection of growth factors/cytokines.3,25 Todate, the recombinant viral system is well suited for a study of itspotential as a therapy of tendon injuries using experimental animalmodels. Two main strategies for gene transfer using vectors can beenvisioned: (i) in vivo transfer with a vector that is applied directly tothe relevant tissue (in vivo direct gene transfer method) and (ii) theremoval of cells from the body, transfer of the gene in vitro, and thenreintroduction into the target site in the body (ex vivo indirect genetransfer method).25 The in vivo direct gene transfer method is less inva-sive and technically easier than transfer of cells in vitro. However, thedisadvantages of in vivo transfer are potential inflammatory responsesand non-specific infections at the injury site. The ex vivo indirect genetransfer method can ensure the collection of only selected/targeted cellsin vitro that express the transgene at high concentrations to the injurysite with less chance of contamination of viral DNA and proteins.Therefore, the ex vivo indirect gene transfer method could be prefer-able for the treatment of a large and degenerative tendon injury such asthat to the rotator cuff, rather than an acute tendon injury.

Future directions for tendon injury treatment

Tendon injuries clinically represent the serious and still unresolvedproblem of how best to restore a damaged tendon to more nearlynormal structural integrity and mechanical strength. Considering theunique feature of tendons with their constant and high mechanicalloading, new treatment modalities are required to promote the regener-ation of tendon tissues. Tissue engineering offers many approaches fortreating tendon injuries and restoring tissues and joint functions.Indeed, a variety of synthetic and biologic scaffolds have been devel-oped. The combination of several scaffolds, including syntheticmaterials, cells such as MSCs or tendon progenitor cells and growth





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factors/cytokines, could be an attractive way to generate an appropriatemicroenvironment for both transplanted and tendon cells at the injuredsite. We also foresee many other novel cellular sources and technol-ogies, induced pluripotent stem (iPS) cells or tendon stem/progenitorcells and nanomedicine scaffolds. The emerging interventions usingthose orthopaedic tissue engineering technologies have shown a thera-peutic effect in experimental animal models of tendon injury.However, their efficacy and safety in humans remains to be elucidatedin the clinical setting. Unfortunately, to date, the development of newtreatment strategies for injured tendons has been hindered because ofour limited understanding of basic tendon biology. Nevertheless, thetranslation of basic research into improved therapeutic approaches isan essential step in the management of patients with tendon injuries,having the potential for significant benefit to public health.

Acknowledgements

We wish to acknowledge many outstanding contributions of investi-gators in the field whose work could not be cited because of spaceconstraints.

Funding

This work was supported in part by National Institutes of Health,grant no. R01 DK074538, The Cleveland Clinic and the SumitomoFoundation, Japan (to T. Sakai).

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