Extracorporeal perfusion of isolated organs of large animals - Bridging the gap between in vitro and in vivo studies

Since the early 20th century extracorporeal perfusion of large animal organs has been used to investigate a broad variety of research questions, thereby overcoming common draw backs of in vitro studies without suffering from ethical concerns associated with live animal research. The technique is in accordance with the 3R principles and represents an excellent opportunity to investigate in detail the physiology of organs under standardised conditions. It is also suitable for the translation of basic pre-clinical research into a more relevant arena prior to or avoiding altogether live animal research. Furthermore, organ perfusion has also been an important tool in developing methods of organ preservation for transplantation surgery. Yet due to the nature of the experiments only short term observations can be made and while cells are still exposed to their regional secretome, the whole organ itself is isolated from the body and correlations between organ-systems cannot be taken into consideration. This review gives an overview over the history of extracorporeal perfusion of large animal organs and limbs highlighting major achievements in the field and discussing different experimental set-ups. Advantages and limitations of this technique are presented. Prospective future research perspectives, which might include tracking of specific cell types and study of their distinct behaviour towards different stimuli, are given to illustrate the relevance of this method.</p

The effect of exercise on cytokine concentration in equine autologous conditioned serum

Background: Autologous conditioned serum (ACS) is a commonly administered intra-articular treatment for the management of osteoarthritis in athletic horses. Objectives: To investigate the influence of exercise on the concentration of cytokines in a non-commercial method of ACS production. Study design: Non-randomised cross over design. Methods: Whole blood was obtained from eight healthy Standardbred horses immediately prior to, 1 h and 24 h following a single bout of exhaustive exercise. Blood was processed using a non-commercial method of ACS production. Fluorescent microsphere immunoassay (FMIA) analysis was performed to quantify Interleukin 1 receptor antagonist (IL-1Ra), Interleukin-10 (IL-10), Interleukin 1β (IL-1β) and tumour necrosis factor α (TNF-α) concentrations at each time point. Mixed effect repeated measures analysis of variance (ANOVA) was used to compare the pre-exercise and post-exercise cytokine concentrations. Significance was set at P &lt; 0.05. Results: A reduced concentration of IL-1Ra (median 584.4, IQR 81.9–5098 pg/ml, p = 0.004) and an increased concentration of TNF-α (11.92, 9.28–39.75 pg/ml, P =.05) at 1 h post-exercise were observed when compared with baseline values (IL-Ra 7349, 1272–10 760 pg/ml; TNF TNF-α 11.16, 8.36–32.74 pg/ml). No difference in cytokine concentrations of IL-10 or IL-1β were found between any of the time points. Main limitations: The large biological variability and small sample size represents limitations of this study. Conclusions: These results suggest that a single bout of intense exercise can reduce the concentration of the anti-inflammatory cytokine IL-1Ra and increase the concentration of the pro-inflammatory cytokine TNF-α, reducing the 'anti-inflammatory' cytokine composition of ACS. Our findings suggest that collection of blood for ACS production should be performed no sooner than 24 h following a single episode of intense exercise.</p

Comparison of interfragmentary compression across simulated condylar fractures repaired using four techniques

IntroductionEquine condylar fractures are commonly repaired using cortex screws applied in lag fashion. Inadequate interfragmentary compression can lead to post-operative complications.MethodsLateral condylar fractures were simulated in 21 cadaver limbs (8 third metatarsals, 13 third metacarpals). In each limb, pressure-sensitive film (Prescale®, Fuji Photo Film Co.) was placed in each osteotomy prior to repair with 4.5 mm diameter cortex screws placed in lag fashion. Screws were placed in linear (L), triangular (T), linear plus a washer (LW) and sequentially tightened triangular configurations (TD1). All screws were tightened to a torque of 4 Nm. Pressure prints obtained were scanned using dedicated software (Fuji FPD-8010E, Fuji Photo Film Co.). A Bayesian Network (BN) model was developed to investigate the impact and interrelationship of each factor on interfragmentary compression. Sixty-three repairs (20*L, 24* T, 11*TD1, and 8*LW) performed on 21 limbs were included in the analysis.ResultsThe BN predicted mean contact area (±s.d.) for pressures within the operating range of the prescale film [≥2.5 Megapascals (MPa) ≤ 10 MPa] by L, T, TD1 and LW repairs were 403mm2 ± (140), 411 mm2 ± (120), 403 mm2 ± (120), and 366mm2 ± (70). The mean contact area (± s.d.) created by L, T, TD1 and LW repairs at pressures &gt;10 MPa were 112 mm2 ± (48), 167 mm2 ± (67), 142 mm2 ± (50), and 100mm2 ± (27). When pressures ≥2.5 MPA to ≤10 MPa were considered, the construct (T or L), washer and screw tightening sequence variables had a very low effect on interfragmentary contact area. At pressures &gt;10 MPa BN sensitivity findings were 16.3, 5.03, and 0.133% for construct, washer and screw tightening sequence. The BN model indicated that triangular repair configuration had a weak influence in the ≥2.5 MPa ≤ 10 MPa range and a moderate influence in the &lt;10 MPa range, on interfragmentary compression. The addition of a washer and the screw tightening sequence had a weak influence on interfragmentary compression at all pressure ranges.DiscussionThe results show that triangular repairs create larger interfragmentary contact areas at greater interfragmentary pressure in simulated condylar fractures, however it is unknown if this results in improved repair stability in the clinical scenario

Ex vivo effect of gold nanoparticles on porcine synovial membrane

Gold nanoparticles (AuNPs) have great potential as carriers for local drug delivery and as a primary therapeutic for treatment of inflammation. Here we report on the AuNP-synovium interaction in an ex vivo model of intra-articular application for treatment of joint inflammation. Sheets of porcine femoropatellar synovium were obtained post mortem and each side of the tissue samples was maintained in a separate fluid environment. Permeability to AuNPs of different sizes (5−52 nm) and biomarker levels of inflammation were determined to characterize the ex vivo particle interaction with the synovium. Lipopolysaccharide or recombinant human interleukin-1β were added to fluid environments to assess the ex vivo effect of pro-inflammatory factors on permeability and biomarker levels. The synovium showed size selective permeability with only 5 nm AuNPs effectively permeating the entire tissues’ width. This process was further governed by particle stability in the fluid environment. AuNPs reduced matrix metalloproteinase and lactate dehydrogenase activity and hyaluronic acid concentrations but had no effect on prostaglandin E(2) levels. Exposure to pro-inflammatory factors did not significantly affect AuNP permeation or biomarker levels in this model. Results with ex vivo tissue modeling of porcine synovium support an anti-inflammatory effect of AuNPs warranting further investigation

Intratumoural tigilanol tiglate in the multicentre treatment of equine sarcoids and cutaneous melanomas

BACKGROUND: Intralesional chemotherapeutic administration represents an important treatment option for equine cutaneous neoplasia. Tigilanol-tiglate (TT), a novel molecule extracted from Fontainea picrosperma, an Australian rainforest plant, is registered for intratumoural treatment of canine MCT, leading to rapid oncosis and tumour slough. Evidence from horses is limited but suggests that efficacy may be similar. OBJECTIVES: To evaluate the response to intratumoural TT treatment in horses with sarcoids (fibroblastic/nodular) and cutaneous melanomas. STUDY DESIGN: Two noncontrolled prospective multicentre clinical trials, one for each of sarcoids and melanomas. METHODS: Cases were enrolled across multiple sites and treated by the same site-specific clinician with intralesional TT (sarcoids: 0.35 mg/cm3; melanomas: 0.2 mg/cm3 of tumour volume - Tvol; max dose 2 mg). Quantitative (Tvol regression) and qualitative outcomes (likely tumour free (LTF) per expert opinion) were recorded, and potential determinants of efficacy were assessed using random effects logistic models. A full clinical response was complete Tvol regression and a LTF treatment site. RESULTS: Forty-one sarcoids and 97 melanomas were enrolled and treated. 73/74% of treated sarcoids/melanomas showed complete Tvol regression. 64/61% (sarcoids/melanomas) showed a full clinical response at medians of 546/247 days post final treatment. For both tumour types, this response was dependent on initial tumour volume (Psarcoids = 0.006; Pmelanomas <0.001). The predicted probability of a full clinical response was 6 times greater for initially small sarcoids (Tvol = 1 cm3) than for the maximum study volume (Tvol = 6 cm3). For melanomas in the perineal region, this was 11 times greater for Tvol ≤0.3 cm3 than for tumours ≥2.0 cm3. For melanomas, tumour location further affected treatment efficacy = 0.005). In total, 5 adverse events were reported. MAIN LIMITATIONS: Lack of treatment control and histologic/biomolecular follow-up data. CONCLUSIONS: The observed therapeutic efficacy of TT supports clinical use as well as early interventions in horses. Successful use necessitates knowledge of the drug's mode of action and management of associated local site responses

A Rare Case of Deep Digital Flexor Tendinopathy Following Centesis of the Navicular Bursa

Navicular bursa (NB) centesis is a common diagnostic and therapeutic procedure in equine practice. This case report documents the clinical, diagnostic imaging and histological findings in a horse with a suspected iatrogenic deep digital flexor tendon (DDFT) injury following centesis of the NB via a modified distal plantar approach (placement of two needles in a weight bearing position). Although it cannot be proven with absolute certainty, the authors believe that this is the first reported case where NB centesis is the likely cause of a DDFT lesion, and with magnetic resonance imaging performed both pre- and post-centesis. With this potential, though rare, complication of the procedure, alternative tendon sparing injection techniques should be considered prior to NB centesis in certain cases

Radiographic assessment of the femorotibial joint of the CCLT rabbit experimental model of osteoarthritis

<p>Abstract</p> <p>Background</p> <p>The purposes of the study were to determine the relevance and validity of in vivo non-invasive radiographic assessment of the CCLT (Cranial Cruciate Ligament Transection) rabbit model of osteoarthritis (OA) and to estimate the pertinence, reliability and reproducibility of a radiographic OA (ROA) grading scale and associated radiographic atlas.</p> <p>Methods</p> <p>In vivo non-invasive extended non weight-bearing radiography of the rabbit femorotibial joint was standardized. Two hundred and fifty radiographs from control and CCLT rabbits up to five months after surgery were reviewed by three readers. They subsequently constructed an original semi-quantitative grading scale as well as an illustrative atlas of individual ROA feature for the medial compartment. To measure agreements, five readers independently scored the same radiographic sample using this atlas and three of them performed a second reading. To evaluate the pertinence of the ROA grading scale, ROA results were compared with gross examination in forty operated and ten control rabbits.</p> <p>Results</p> <p>Radiographic osteophytes of medial femoral condyles and medial tibial condyles were scored on a four point scale and dichotomously for osteophytes of medial fabella. Medial joint space width was scored as normal, reduced or absent. Each ROA features was well correlated with gross examination (p < 0.001). ICCs of each ROA features demonstrated excellent agreement between readers and within reading. Global ROA score gave the highest ICCs value for between (ICC 0.93; CI 0.90-0.96) and within (ICC ranged from 0.94 to 0.96) observer agreements. Among all individual ROA features, medial joint space width scoring gave the highest overall reliability and reproducibility and was correlated with both meniscal and cartilage macroscopic lesions (r_s= 0.68 and r_s= 0.58, p < 0.001 respectively). Radiographic osteophytes of the medial femoral condyle gave the lowest agreements while being well correlated with the macroscopic osteophytes (r_s= 0.64, p < 0.001).</p> <p>Conclusion</p> <p>Non-invasive in vivo radiography of the rabbit femorotibial joint is feasible, relevant and allows a reproducible grading of experimentally induced OA lesion. The radiographic grading scale and atlas presented could be used as a template for in vivo non invasive grading of ROA in preclinical studies and could allow future comparisons between studies.</p