Effect of single intralesional treatment of surgically induced equine superficial digital flexor tendon core lesions with adipose-derived mesenchymal stromal cells: a controlled experimental trial.

BACKGROUND: Adipose tissue is a promising source of mesenchymal stromal cells (MSCs) for the treatment of tendon disease. The goal of this study was to assess the effect of a single intralesional implantation of adipose tissue-derived mesenchymal stromal cells (AT-MSCs) on artificial lesions in equine superficial digital flexor tendons (SDFTs). METHODS: During this randomized, controlled, blinded experimental study, either autologous cultured AT-MSCs suspended in autologous inactivated serum (AT-MSC-serum) or autologous inactivated serum (serum) were injected intralesionally 2 weeks after surgical creation of centrally located SDFT lesions in both forelimbs of nine horses. Healing was assessed clinically and with ultrasound (standard B-mode and ultrasound tissue characterization) at regular intervals over 24 weeks. After euthanasia of the horses the SDFTs were examined histologically, biochemically and by means of biomechanical testing. RESULTS: AT-MSC implantation did not substantially influence clinical and ultrasonographic parameters. Histology, biochemical and biomechanical characteristics of the repair tissue did not differ significantly between treatment modalities after 24 weeks. Compared with macroscopically normal tendon tissue, the content of the mature collagen crosslink hydroxylysylpyridinoline did not differ after AT-MSC-serum treatment (p = 0.074) while it was significantly lower (p = 0.027) in lesions treated with serum alone. Stress at failure (p = 0.048) and the modulus of elasticity (p = 0.001) were significantly lower after AT-MSC-serum treatment than in normal tendon tissue. CONCLUSIONS: The effect of a single intralesional injection of cultured AT-MSCs suspended in autologous inactivated serum was not superior to treatment of surgically created SDFT lesions with autologous inactivated serum alone in a surgical model of tendinopathy over an observation period of 22 weeks. AT-MSC treatment might have a positive influence on collagen crosslinking of remodelling scar tissue. Controlled long-term studies including naturally occurring tendinopathies are necessary to verify the effects of AT-MSCs on tendon disease.

Effect of intralesional platelet-rich plasma (PRP) treatment on clinical and ultrasonographic parameters in equine naturally occurring superficial digital flexor tendinopathies - a randomized prospective controlled clinical trial.

BACKGROUND: Regenerative and anti-inflammatory effects on tendinopathies have been attributed to blood-derived biologicals. To date the evidence for the efficacy of autologous platelet-rich plasma (PRP) treatment of naturally occurring equine tendinopathies is limited. The purpose of this placebo-controlled clinical trial was to describe the effect of a single treatment of equine superficial digital flexor tendon (SDFT) disease with PRP on clinical and ultrasonographic parameters. Twenty horses with naturally occurring tendinopathies of forelimb SDFTs were randomly assigned to the PRP-treated group (n = 10) or control group (n = 10) after clinical and ultrasonographic examination. The SDFTs received an intralesional treatment with autologous PRP or were injected with saline, respectively (day 0). All horses participated in a standardized exercise programme and were re-examined clinically, with B-mode ultrasonography (5 times at regular intervals) and ultrasound tissue characterization (week 12 and 24 after treatment) until week 24. Long-term performance was estimated via telephone inquiry. RESULTS: Compared to day 0, lameness decreased significantly by week 8 after treatment with PRP and by week 12 in the control group. Ultrasonographically there was no difference in the summarized cross sectional area between the groups at any time point. Ultrasound tissue characterization showed that echo types representing disorganized matrix decreased significantly throughout the observation period in the PRP-treated group. Echo type II, representing discontinuous fascicles, not yet aligned into lines of stress was significantly higher 24 weeks after PRP treatment. Eighty percent of the PRP treated horses reached their previous or a higher level of performance after 12 months compared to 50 % in the CG. After 24 months these proportions were 60 % and 50 %, respectively. CONCLUSIONS: A single intralesional treatment with PRP up to 8 weeks after onset of clinical signs of tendinopathy contributes to an earlier reduction of lameness compared to saline treatment and to an advanced organization of repair tissue as the fibrillar matrix is getting organized into fascicles while remodelling continues. Long term, PRP treatment has the potential to increase the number of horses reaching their previous level of performance. Earlier treatment of tendinopathy with PRP should be considered to enhance these effects.

Tracking of autologous adipose tissue-derived mesenchymal stromal cells with in vivo magnetic resonance imaging and histology after intralesional treatment of artificial equine tendon lesions--a pilot study.

BACKGROUND: Adipose tissue-derived mesenchymal stromal cells (AT-MSCs) are frequently used to treat equine tendinopathies. Up to now, knowledge about the fate of autologous AT-MSCs after intralesional injection into equine superficial digital flexor tendons (SDFTs) is very limited. The purpose of this study was to monitor the presence of intralesionally injected autologous AT-MSCs labelled with superparamagnetic iron oxide (SPIO) nanoparticles and green fluorescent protein (GFP) over a staggered period of 3 to 9 weeks with standing magnetic resonance imaging (MRI) and histology. METHODS: Four adult warmblood horses received a unilateral injection of 10 × 10(6) autologous AT-MSCs into surgically created front-limb SDFT lesions. Administered AT-MSCs expressed lentivirally transduced reporter genes for GFP and were co-labelled with SPIO particles in three horses. The presence of AT-MSCs in SDFTs was evaluated by repeated examinations with standing low-field MRI in two horses and post-mortem in all horses with Prussian blue staining, fluorescence microscopy and with immunofluorescence and immunohistochemistry using anti-GFP antibodies at 3, 5, 7 and 9 weeks after treatment. RESULTS: AT-MSCs labelled with SPIO particles were detectable in treated SDFTs during each MRI in T2*- and T1-weighted sequences until the end of the observation period. Post-mortem examinations revealed that all treated tendons contained high numbers of SPIO- and GFP-labelled cells. CONCLUSIONS: Standing low-field MRI has the potential to track SPIO-labelled AT-MSCs successfully. Histology, fluorescence microscopy, immunofluorescence and immunohistochemistry are efficient tools to detect labelled AT-MSCs after intralesional injection into surgically created equine SDFT lesions. Intralesional injection of 10 × 10(6) AT-MSCs leads to the presence of high numbers of AT-MSCs in and around surgically created tendon lesions for up to 9 weeks. Integration of injected AT-MSCs into healing tendon tissue is an essential pathway after intralesional administration. Injection techniques have to be chosen deliberately to avoid reflux of the cell substrate injected. In vivo low-field MRI may be used as a non-invasive tool to monitor homing and engraftment of AT-MSCs in horses with tendinopathy of the SDFT.

Effect of a single injection of autologous conditioned serum (ACS) on tendon healing in equine naturally occurring tendinopathies.

INTRODUCTION: Autologous blood-derived biologicals, including autologous conditioned serum (ACS), are frequently used to treat tendinopathies in horses despite limited evidence for their efficacy. The purpose of this study was to describe the effect of a single intralesional injection of ACS in naturally occurring tendinopathies of the equine superficial digital flexor tendon (SDFT) on clinical, ultrasonographic, and histological parameters. METHODS: Fifteen horses with 17 naturally occurring tendinopathies of forelimb SDFTs were examined clinically and ultrasonographically (day 0). Injured tendons were randomly assigned to the ACS-treated group (n = 10) receiving a single intralesional ACS injection or included as controls (n = 7) which were either untreated or injected with saline on day 1. All horses participated in a gradually increasing exercise programme and were re-examined nine times at regular intervals until day 190. Needle biopsies were taken from the SDFTs on days 0, 36 and 190 and examined histologically and for the expression of collagen types I and III by immunohistochemistry. RESULTS: In ACS-treated limbs lameness decreased significantly until day 10 after treatment. Swelling (scores) of the SDFT region decreased within the ACS group between 50 and 78 days after treatment. Ultrasonographically, the percentage of the lesion in the tendon was significantly lower and the echogenicity of the lesion (total echo score) was significantly higher 78 and 106 days after intralesional ACS injection compared to controls. Histology revealed that, compared to controls, tenocyte nuclei were more spindle-shaped 36 days after ACS injection. Immunohistochemistry showed that collagen type I expression significantly increased between days 36 and 190 after ACS injection. CONCLUSIONS: Single intralesional ACS injection of equine SDFTs with clinical signs of acute tendinopathy contributes to an early significant reduction of lameness and leads to temporary improvement of ultrasonographic parameters of repair tissue. Intralesional ACS treatment might decrease proliferation of tenocytes 5 weeks after treatment and increase their differentiation as demonstrated by elevated collagen type I expression in the remodelling phase. Potential enhancement of these effects by repeated injections should be tested in future controlled clinical investigations.

Comparison of the diagnostic value of ultrasonography and standing radiography for pelvic-femoral disorders in horses.

OBJECTIVE: To assess agreement between ultrasonography (transcutaneous and transrectal) and standing radiography in horses with fractures in the pelvic region and disorders of the coxofemoral joint. STUDY DESIGN: Case series. ANIMALS: Warmblood horses (n=23) and 2 ponies. METHODS: Medical records (1999-2008) of equids with pelvic or coxofemoral disorders that had pelvic radiography and ultrasonography were retrieved and results of both techniques compared. RESULTS: Radiography and ultrasonography each identified equal numbers of fractures of the tuber coxa (n=4), ilial shaft (2), ischium (3), femoral neck (2), and osteoarthritis/osis of the coxofemoral joint (6). Fractures of the ilial wing (4) were only identified by ultrasonography not by standing radiography. Of 9 acetabular fractures, 3 were identified on radiographs only, 5 were identified with both modalities. One pubic fracture was identified using ultrasonography and radiography. One acetabular and 1 pubic fracture were only diagnosed on necropsy. CONCLUSIONS: We found reasonable agreement (73%; 24/33) between ultrasonography and standing radiography for diagnosis of pelvic-femoral disorders. Ultrasonography was more useful for ilial wing fractures and radiography for acetabular fractures. CLINICAL RELEVANCE: Ultrasonography is a rapid, safe imaging technique for detecting disorders of the pelvic region with a high diagnostic yield and is a preferred initial approach in horses with severe hindlimb lameness.