P. René van Weeren

Effects of platelet-rich plasma on the quality of repair of mechanically induced core lesions in equine superficial digital flexor tendons: A placebo-controlled experimental study

Tendon injuries are notorious for their slow and functionally inferior healing. Intratendinous application of platelet‐rich plasma (PRP) has been reported to stimulate the repair process of tendon injuries, but there is little conclusive evidence for its effectiveness. A placebo‐controlled experimental trial was performed to test the hypothesis that a single intratendinous PRP treatment enhances the quality of tendon repair, as evidenced by improved biochemical, biomechanical, and histological tissue properties. In six horses, tendon lesions were created surgically in the Superficial Digital Flexor Tendons (SDFT) of both front limbs, one of which was treated with PRP and the other with saline. After 24 weeks, the tendons were harvested for biochemical, biomechanical, and histological evaluations. Collagen, glycosaminoglycan, and DNA content (cellularity) was higher in PRP‐treated tendons (p = 0.039, 0.038, and 0.034, respectively). The repair tissue in the PRP group showed a higher strength at failure (p = 0.021) and Elastic Modulus (p = 0.019). Histologically, PRP‐treated tendons featured better organization of the collagen network (p = 0.031) and signs of increased metabolic activity (p = 0.031). It was concluded that PRP increases metabolic activity and seems to advance maturation of repair tissue over nontreated experimentally induced tendon lesions, which suggests that PRP might be beneficial in the treatment of clinical tendon injuries.

Hyaluronic acid and dextran-based semi-IPN hydrogels as biomaterials for bioprinting.

Bioprinting is a recent technology in tissue engineering used for the design of porous constructs through layer-by-layer deposition of cell-laden material. This technology would benefit from new biomaterials that can fulfill specific requirements for the fabrication of well-defined 3D constructs, such as the preservation of cell viability and adequate mechanical properties. We evaluated the suitability of a novel semi-interpenetrating network (semi-IPN), based on hyaluronic acid and hydroxyethyl-methacrylate-derivatized dextran (dex-HEMA), to form 3D hydrogel bioprinted constructs. The rheological properties of the solutions allowed proper handling during bioprinting, whereas photopolymerization led to stable constructs of which their mechanical properties matched the wide range of mechanical strengths of natural tissues. Importantly, excellent viability was observed for encapsulated chondrocytes. The results demonstrate the suitability of hyaluronic acid/dex-HEMA semi-IPNs to manufacture bioprinted constructs for tissue engineering.

Endochondral bone formation in gelatin methacrylamide hydrogel with embedded cartilage-derived matrix particles

The natural process of endochondral bone formation in the growing skeletal system is increasingly inspiring the field of bone tissue engineering. However, in order to create relevant-size bone grafts, a cell carrier is required that ensures a high diffusion rate and facilitates matrix formation, balanced by its degradation. Therefore, we set out to engineer endochondral bone in gelatin methacrylamide (GelMA) hydrogels with embedded multipotent stromal cells (MSCs) and cartilage-derived matrix (CDM) particles. CDM particles were found to stimulate the formation of a cartilage template by MSCs in the GelMA hydrogel in vitro. In a subcutaneous rat model, this template was subsequently remodeled into mineralized bone tissue, including bone-marrow cavities. The GelMA was almost fully degraded during this process. There was no significant difference in the degree of calcification in GelMA with or without CDM particles: 42.5 ± 2.5% vs. 39.5 ± 8.3% (mean ± standard deviation), respectively. Interestingly, in an osteochondral setting, the presence of chondrocytes in one half of the constructs fully impeded bone formation in the other half by MSCs. This work offers a new avenue for the engineering of relevant-size bone grafts, by the formation of endochondral bone within a degradable hydrogel.

Of Mice, Men and Elephants: The Relation between Articular Cartilage Thickness and Body Mass

Mammalian articular cartilage serves diverse functions, including shock absorption, force transmission and enabling low-friction joint motion. These challenging requirements are met by the tissue’s thickness combined with its highly specific extracellular matrix, consisting of a glycosaminoglycan-interspersed collagen fiber network that provides a unique combination of resilience and high compressive and shear resistance. It is unknown how this critical tissue deals with the challenges posed by increases in body mass. For this study, osteochondral cores were harvested post-mortem from the central sites of both medial and lateral femoral condyles of 58 different mammalian species ranging from 25 g (mouse) to 4000 kg (African elephant). Joint size and cartilage thickness were measured and biochemical composition (glycosaminoclycan, collagen and DNA content) and collagen cross-links densities were analyzed. Here, we show that cartilage thickness at the femoral condyle in the mammalian species investigated varies between 90 µm and 3000 µm and bears a negative allometric relationship to body mass, unlike the isometric scaling of the skeleton. Cellular density (as determined by DNA content) decreases with increasing body mass, but gross biochemical composition is remarkably constant. This however need not affect life-long performance of the tissue in heavier mammals, due to relatively constant static compressive stresses, the zonal organization of the tissue and additional compensation by joint congruence, posture and activity pattern of larger mammals. These findings provide insight in the scaling of articular cartilage thickness with body weight, as well as in cartilage biochemical composition and cellularity across mammalian species. They underscore the need for the use of appropriate in vivo models in translational research aiming at human applications.

Zonal Chondrocyte Subpopulations Reacquire Zone-Specific Characteristics During In Vitro Redifferentiation

Background If chondrocytes from the superficial, middle, and deep zones of articular cartilage could maintain or regain their characteristic properties during in vitro culture, it would be feasible to create constructs comprising these distinctive zones. Hypothesis Zone-specific characteristics of zonal cell populations will disappear during 2-dimensional expansion but will reappear after 3-dimensional redifferentiation, independent of the culture technique used (alginate beads versus pellet culture). Study Design Controlled laboratory study. Methods Equine articular chondrocytes from the 3 zones were expanded in monolayer culture (8 donors) and subsequently redifferentiated in pellet and alginate bead cultures for up to 4 weeks. Glycosaminoglycans and DNA were quantified, along with immunohistochemical assessment of the expression of various zonal markers, including cartilage oligomeric protein (marking cells from the deeper zones) and clusterin (specifically expressed by superficial chondrocytes). Results Cell yield varied between zones, but proliferation rates did not show significant differences. Expression of all evaluated zonal markers was lost during expansion. Compared to the alginate bead cultures, pellet cultures showed a higher amount of glycosaminoglycans produced per DNA after redifferentiation. In contrast to cells in pellet cultures, cells in alginate beads regained zonal differences, as evidenced by zone-specific reappearance of cartilage oligomeric protein and clusterin, as well as significantly higher glycosaminoglycans production by cells from the deep zone compared to the superficial zone. Conclusion Chondrocytes isolated from the 3 zones of equine cartilage can restore their zone-specific matrix expression when cultured in alginate after in vitro expansion. Clinical Relevance Appreciation of the zonal differences can lead to important advances in cartilage tissue engineering. Findings support the use of hydrogels such as alginate for engineering zonal cartilage constructs.

Pain in Osteoarthritis

This article focuses on pain associated with osteoarthritis (OA). It first describes the basic biology of articular cartilage and other joint structures and the defining features of the osteoarthritic disease process. Subsequently, the possible origins of pain in OA are discussed before embarking on how to manage this clinical entity. The emphasis is on the pharmacologic management of joint pain, and attention is paid to systemic therapeutic strategies as well as to local (intra-articular) treatment modalities. Nonmedical ways of modulating joint pain are briefly mentioned, but not extensively discussed, as these are outside the scope of this article.

Monitoring of the repair process of surgically created lesions in equine superficial digital flexor tendons by use of computerized ultrasonography.

OBJECTIVE To evaluate quantitative ultrasonography for objective monitoring of the healing process and prognostication of repair quality in equine superficial digital flexor (SDF) tendons. ANIMALS 6 horses with standardized surgical lesions in SDF tendons of both forelimbs. PROCEDURES Healing was monitored for 20 weeks after surgery by use of computerized ultrasonography. Pixels were categorized as C (intact fasciculi), B (incomplete fasciculi), E (accumulations of cells and fibrils), or N (homogenous fluid or cells). Four scars with the best quality of repair (repair group) and 4 scars with the lowest quality (inferior repair group) were identified histologically. Ratios for C, B, E, and N in both groups were compared. RESULTS During 4 weeks after surgery, lesions increased 2- to 4-fold in length and 10-fold in volume. Until week 3 or 4, structure-related C and B ratios decreased sharply, whereas E and N ratios increased. After week 4, C and B ratios increased with gradually decreasing E and N ratios. At week 12, C and B ratios were equivalent. After week 12, C ratio increased slowly, but B ratio more rapidly. At week 20, C ratio remained constant, B ratio was substantially increased, and E and N ratios decreased. Values for the inferior repair group were most aberrant from normal. Ratios for C differed significantly between repair and inferior repair groups at weeks 16 and 18 and for B beginning at 14 weeks. CONCLUSIONS AND CLINICAL RELEVANCE Computerized ultrasonography provided an excellent tool for objective monitoring of healing tendons in horses and reliable prognostication of repair quality.

Urolithiasis in small ruminants. I. A retrospective evaluation of urethrostomy.

A retrospective study on the results of urethrostomy in 28 small ruminants was made. Ten animals were euthanised during surgery. In the 18 remaining animals short-term results were good. Recurrence of symptoms occurred in 8 animals (45%). Five animals were operated a second time, leading to 4 recurrences. Two animals were operated a third time and remained free of symptoms.

Effect of loading on the organization of the collagen fibril network in juvenile equine articular cartilage

We investigated the effects of exercise‐induced loading on the collagen network of equine articular cartilage. Collagen fibril architecture at a site (1) subjected to intermittent high‐intensity loading was compared with that of an adjacent site (2) sustaining continuous low‐level load. From horses exposed to forced exercise (CONDEX group) or not (PASTEX group), the spatial parallelism of fibrils and the orientation angle between fibrils and the surface at depths 9 µm apart through cartilage from surface to tidemark were determined using polarized light microscopy, and expressed as parallelism index (PI) and orientation index (OI). PI was significantly higher in site 2 than 1 in CONDEX and PASTEX groups. PI was significantly higher in forced exercised horses at site 2 but not site 1. OI was significantly greater (more perpendicular to the surface) in the superficial and deep cartilage of site 2 than 1 in both CONDEX and PASTEX groups. Superficial zone OI was higher in exercised horses at site 1 but not at site 2. Exercise increased collagen parallelism and affected orientation. The site differences in OI indicate that Benninghoffs classic predominantly perpendicular arcades appear not to be a consistent architectural feature, but adapt to local forces sustained.

Computerised analysis of standardised ultrasonographic images to monitor the repair of surgically created core lesions in equine superficial digital flexor tendons following treatment with intratendinous platelet rich plasma or placebo.

The effectiveness of new therapies to treat tendon injuries is difficult to determine and is often based on semi-quantitative methods, such as grey level analysis of ultrasonographic images or subjective pain scores. The alternatives are costly and long-lasting end-stage studies using experimental animals. In this study, a method of ultrasonographic tissue characterisation (UTC), using mathematical analysis of contiguous transverse ultrasonographic images, was used for intra-vital monitoring of the healing trajectory of standardised tendon lesions treated with platelet rich plasma (PRP) or placebo. Using UTC it was possible to detect significant differences between the groups in the various phases of repair. At end stage, over 80% of pixels showed correct alignment in the PRP group, compared with just over 60% in the placebo group (P<0.05). UTC also showed significant differences in the course of the healing process between PRP treated and placebo treated animals throughout the experiment. It was concluded that computerised analysis of ultrasonographic images is an excellent tool for objective longitudinal monitoring of the effects of treatments for superficial digital flexor tendon lesions in horses.