Cellular, molecular and genetical overview of avian tibial dyschondroplasia.

Tibial dyschondroplasia (TD) is an intractable avian bone disease that causes severe poultry economic losses. The pathogenicity of TD is unknown. Therefore, TD disease has not been evacuated yet. Based on continuous research findings, we have gone through the molecular and cellular insight into the TD and proposed possible pathogenicity for future studies. Immunity and angiogenesis-related genes expressed in the erythrocytes of chicken, influenced the apoptosis of chicken chondrocytes to cause TD. TD could be defined as the irregular, unmineralized and un-vascularized mass of cartilage, which is caused by apoptosis, degeneration and insufficient blood supply at the site of the chicken growth plate. The failure of angiogenesis attributed improper nutrients supply to the chondrocytes; ultimately, bone development stopped, poor calcification of cartilage matrix, and apoptosis of chondrocytes occurred. Recent studies explore potential signaling pathways that regulated TD in broiler chickens, including parathyroid hormone-related peptide (PTHrP), transforming growth factor ß (TGF- ß)/bone morphogenic proteins (BMPs), and hypoxia-inducible factor (HIF). Several studies have reported many medicines to treat TD. However, recently, rGSTA3 protein (50 µg·kg-1) is considered the most proper TD treatment. The present review has summarized the molecular and cellular insight into the TD, which will help researchers in medicine development to evacuate TD completely.

Tibial growth plate vascularization is inhibited by the dithiocarbamate pesticide thiram in chickens: potential relationship to peripheral platelet counts alteration.

The widespread use of thiram has raised concerns for health and its toxic effects, but the underlying toxicity mechanism on platelets and bones is poorly defined. Here, we found a significant increase in the number of platelets in chickens with the thiram intake, due to the increased expression of thrombopoietin mRNA in the dysfunction liver. Furthermore, the decreased vascular distribution and cell death of chondrocytes in the tibial growth plates (TGPs) were observed, resulting in bone growth inhibition, which is associated with the abnormal activation of platelets leading to the extraordinary decrease of vascular endothelial growth factor A (VEGFA) and angiopoietin-1 protein were released and their corresponding receptors VEGFR2 and Tie-2 expressions were also reduced in the TGPs. Taken together, these findings revealed that thiram has an adverse effect on bones and platelets, which may have a high risk of thrombosis and osteoarthritis.

Disrupted type II collagenolysis impairs angiogenesis, delays endochondral ossification and initiates aberrant ossification in mouse limbs.

Cartilage remodelling and chondrocyte differentiation are tightly linked to angiogenesis during bone development and endochondral ossification. To investigate whether collagenase-mediated cleavage of the major cartilage collagen (collagen II) plays a role in this process, we generated a knockin mouse in which the mandatory collagenase cleavage site at PQG775↓776LAG, was mutated to PPG775↓776MPG (Col2a1Bailey). This approach blocked collagen II cleavage, and the production of putative collagen II matrikines derived from this site, without modifying matrix metalloproteinase expression or activity. We report here that this mouse (Bailey) is viable. It has a significantly expanded growth plate and exhibits delayed and abnormal angiogenic invasion into the growth plate. Deeper electron microscopy analyses revealed that, at around five weeks of age, a small number of blood vessel(s) penetrate into the growth plate, leading to its abrupt shrinking and the formation of a bony bridge. Our results from in vitro and ex vivo studies suggest that collagen II matrikines stimulate the normal branching of endothelial cells and promote blood vessel invasion at the chondro-osseous junction. The results further suggest that failed collagenolysis in Bailey leads to expansion of the hypertrophic zone and formation of a unique post-hypertrophic zone populated with chondrocytes that re-enter the cell cycle and proliferate. The biological rescue of this in vivo phenotype features the loss of a substantial portion of the growth plate through aberrant ossification, and narrowing of the remaining portion that leads to limb deformation. Together, these data suggest that collagen II matrikines stimulate angiogenesis in skeletal growth and development, revealing novel strategies for stimulating angiogenesis in other contexts such as fracture healing and surgical applications.

Evaluation of the Suitability of Miniature Pigs as an Animal Model of Juvenile Osteochondritis Dissecans.

Juvenile osteochondritis dissecans (JOCD) is a developmental disease characterized by formation of intra-articular (osteo)chondral flaps or fragments. Evidence-based treatment guidelines for JOCD are currently lacking. An animal model would facilitate study of JOCD and evaluation of diagnostic and treatment approaches. The purpose of this study was to assess the suitability of miniature pigs as a model of JOCD at the distal femur. First, stifle (knee) joints harvested from three juvenile miniature pigs underwent magnetic resonance imaging (MRI) to establish the vascular architecture of the distal femoral epiphyseal cartilage. Second, vessels supplying the axial or abaxial aspects of the medial femoral condyle were surgically interrupted in four additional juvenile miniature pigs, and the developing epiphyseal cartilage lesions were monitored using three consecutive MRI examinations over nine weeks. The miniature pigs were then euthanized, and their distal femora were harvested for histological evaluation. Vascular architecture of the distal femoral epiphyseal cartilage in the miniature pigs was found to be nearly identical to that of juvenile human subjects, characterized by separate vascular beds supplying the axial and abaxial aspects of the condyles. Surgical interruption of the vascular supply to the abaxial aspect of the medial femoral condyle resulted in ischemic cartilage necrosis (a precursor lesion of JOCD) in 75% (3/4) of the miniature pigs. Cartilage lesions were identified during the first MRI performed 3 weeks post-operatively. No clinically apparent JOCD-like lesions developed. In conclusion, miniature pigs are suitable for modeling JOCD precursor lesions. Further investigation of the model is warranted to assess induction of clinically apparent JOCD lesions. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:2130-2137, 2019.

Osteochondrosis in the Distal Femoral Physis of Pigs Starts With Vascular Failure.

Articular osteochondrosis (OC) arises due to vascular failure and ischemic chondronecrosis. The aim of the study was to describe the histological and computed tomographic (CT) characteristics of changes in the distal femoral physis of pigs, to determine if they represented OC lesions and if the pathogenesis was the same as for articular OC. The material included 19 male Landrace pigs bred for predisposition to OC. One or 2 pigs were euthanized and CT-scanned at 2-week intervals from 82 to 180 days of age. Material from 10 pigs was available for histological validation. The CT scans revealed 31 lesions confirmed in 3 planes and 1 additional macroscopically visible lesion confirmed in 2 CT planes. Twelve of the lesions were histologically validated. All lesions were compatible with OC. Cartilage canal and eosinophilic streak morphological changes corresponded to failure of end arteries coursing from the epiphysis, toward the metaphysis. The location of lesions was compatible with failure at the point of vessel incorporation into bone. Vascular failure was associated with retention of viable hypertrophic chondrocytes and delayed ossification but not cartilage necrosis. Lesion width ranged from 1.1% to 45.6% of the physis. Several lesions were expected to resolve due to small size and evidence of CT-identifiable, reparative ossification. Angular limb deformity was not detected in any pig. The pathogenesis of physeal OC started with vascular failure that was morphologically identical to articular OC. The heritable predisposition may therefore be the same. The association between lesions and limb deformity should be studied further in older pigs in future.

Role of Angiopoietin-like 4 on Bone Vascularization in Chickens Exposed to High-altitude Hypoxia.

The aim of this study was to investigate the role and expression of a novel angiogenic factor (angiopoietin-like 4, ANGPTL4) in tibial growth plates of broiler chickens exposed to high-altitude hypoxia. One-day-old healthy broiler chickens (n = 120) were transported from lowland to a high-altitude hypoxic region (nearly 3,000 m above sea level) and were reared under hypoxic- (natural lower oxygen content) and normoxic conditions (nearly 21% oxygen content) for 14 days. The effect of hypoxia on angiogenesis in the tibial growth plates and hypoxia-inducible factor (HIF)-1α and ANGPTL4 expressions were determined by histological examination, quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), western blot and enzyme-linked immunosorbent assay (ELISA) techniques. The increase in vascular distribution to the hypertrophic chondrocyte zone of tibial growth plates contributed to promoting growth and development of the tibia under hypoxic conditions, which was highly correlated with the upregulation of ANGPTL4 at both the mRNA and protein levels together with activation of HIF-1α under hypoxic conditions. These findings demonstrate that angiogenic factor ANGPTL4 upregulation is involved in tibial growth plate angiogenesis to promote the development of the tibia in broiler chickens under hypoxic conditions. They also suggest that ANGPTL4 may serve as a new molecular therapeutic target for ameliorating tibial dyschondroplasia chicken bone vascularization.

Role and regulation of growth plate vascularization during coupling with osteogenesis in tibial dyschondroplasia of chickens.

Tibial dyschondroplasia (TD) is the most-prevalent leg disorder in fast-growing chickens; it is intractable and characterized by abnormal endochondral bone formation of proximal tibial growth-plates (TGPs). Previous studies have shown that bone is a highly vascularized tissue dependent on the coordinated coupling between angiogenesis and osteogenesis, but the underlying mechanisms of bone formation and bone remodeling are poorly defined in TD chickens. Here, we observed that inhibition of vasculogenesis and angiogenesis remarkably impaired vascular invasion in the hypertrophic chondrocyte zone of the TGPs, resulting in the massive death of chondrocytes due to a shortage of blood vessels and nutrients. Moreover, the balance of the OPG (osteoprotegerin)/RANKL (receptor activator of nuclear factor-kB ligand) system is also severely disrupted during the osteogenesis process while coupling with angiogenesis, both of which eventually lead to abnormal endochondral bone formation in TD chickens. Thus, the process of vascular formation in endochondral bone appears to initiate the pathological changes in TD, and improvement of this process during coupling with osteogenesis may be a potential therapeutic approach to treat this intractable disease.

Vessel architecture in human knee cartilage in children: an in vivo susceptibility-weighted imaging study at 7 T.

OBJECTIVES: To evaluate the clinical feasibility of ultrahigh field 7-T SWI to visualize vessels and assess their density in the immature epiphyseal cartilage of human knee joints. METHODS: 7-T SWI of 12 knees (six healthy volunteers, six patients with osteochondral abnormalities; mean age 10.7 years; 3 female, 9 male) were analysed by two readers, classifying intracartilaginous vessel densities (IVD) in three grades (no vessels, low IVD and high IVD) in defined femoral, tibial and patellar zones. Differences between patients and volunteers, IVDs in different anatomic locations, differences between cartilage overlying osteochondral abnormalities and corresponding normal zones, and differences in age groups were analysed. RESULTS: Interrater reliability showed moderate agreement between the two readers (κ = 0.58, p < 0.001). The comparison of IVDs between patients and volunteers revealed no significant difference (p = 0.706). The difference between zones in the cartilage overlying osteochondral abnormalities to corresponding normal zones showed no significant difference (p = 0.564). IVDs were related to anatomic location, with decreased IVDs in loading areas (p = 0.003). IVD was age dependent, with more vessels present in the younger participants (p = 0.001). CONCLUSIONS: The use of SWI in conjunction with ultrahigh field MRI makes the in vivo visualization of vessels in the growing cartilage of humans feasible, providing insights into the role of the vessel network in acquired disturbances. KEY POINTS: • SWI facilitates in vivo visualization of vessels in the growing human cartilage. • Interrater reliability of the intracartilaginous vessel grading was moderate. • Intracartilaginous vessel densities are dependent on anatomical location and age.

Assessment of pharmacokinetics for microvessel proliferation by DCE-MRI for early detection of physeal bone bridge formation in an animal model.

OBJECTIVES: Bone bridge formation occurs after physeal lesions and can lead to growth arrest if not reversed. Previous investigations on the underlying mechanisms of this formation used histological methods. Therefore, this study aimed to apply a minimally invasive method using dynamic contrast-enhanced MRI (DCE-MRI). MATERIALS AND METHODS: Changes in functional parameters related to the microvessel system were assessed in a longitudinal study of a cohort of an animal model applying a reference region model. The development of morphology of the injured physis was investigated with 3D high-resolution MRI. To acquire complementary information for MRI-related findings qRT-PCR and immunohistochemical data were acquired for a second cohort of the animal model. RESULTS: The evaluation of the pharmacokinetic parameters showed a first rise of the transfer coefficient 7 days post-lesion and a maximum 42 days after operation. The analysis of the complementary data showed a connection of the first rise to microvessel proliferation while the maximum value was linked to bone remodeling. CONCLUSION: The pharmacokinetic analysis of DCE-MRI provides information on a proliferation of microvessels during the healing process as a sign for bone bridge formation. Thereby, DCE-MRI could identify details, which up to now required analyses of highly invasive methods.

In vivo visualization using MRI T2 mapping of induced osteochondrosis and osteochondritis dissecans lesions in goats undergoing controlled exercise.

In vivo visualization of subclinical osteochondrosis (OC) lesions, characterized by necrosis of epiphyseal growth cartilage, is necessary to clarify the pathogenesis of this disease. Hence, our objectives were to demonstrate induced necrosis of the epiphyseal cartilage in vivo using MRI and to monitor progression or resolution of resulting lesions. We also aimed to improve the goat model of OC by introducing controlled exercise. Vascular supply to the epiphyseal cartilage was surgically interrupted in four 5-day-old goats to induce ischemic cartilage necrosis in a medial femoral condyle. Starting 3 weeks postoperatively, goats underwent daily controlled exercise until euthanasia at 6, 10, 11 (n = 2) weeks postoperatively. T2 maps of operated and control femora were obtained in vivo at 3 (n = 4), 6 (n = 4), 9 (n = 3), and 11 (n = 2) weeks postoperatively using a 3 T MR scanner. In vivo MRI findings were validated against MRI results obtained ex vivo at 9.4 T in three goats and compared to histological results in all goats. Surgical interruption of the vascular supply caused ischemic cartilage necrosis in three out of four goats. T2 maps obtained in vivo at 3 T identified regions of increased relaxation time consistent with discrete areas of cartilage necrosis 3-11 weeks postoperatively and demonstrated delayed progression of the ossification front at 9 (n = 1) and 11 (n = 2) weeks postoperatively. In vivo MRI findings were confirmed by ex vivo MRI at 9.4 T and by histology. Identification of cartilage necrosis in clinical patients in the early stages of OC using T2 maps may provide valuable insight into the pathogenesis of this condition. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:868-875, 2017.

Differences in the vascular tree of the femoral trochlear growth cartilage at osteochondrosis-susceptible sites in foals revealed by SWI 3T MRI.

Focal ischemic chondronecrosis of epiphyseal growth cartilage (EGC) during endochondral ossification is believed to be a key early event on the pathway to osteochondrosis (OC) in both animals and humans. The lateral ridge of the equine trochlea is a site where severe osteochondritis dissecans lesions frequently arise and is a model for the study of naturally occurring disease. Non-invasive imaging to investigate EGC vascularity may help elucidate why focal ischemia occurs. 3T MRI susceptibility-weighted imaging (SWI) of femoral trochlea of OC predisposed (n = 10) and control (n = 6) day-old foals, with minimal joint loading after birth, was performed. SWI and 3D images revealed the EGC vascular architecture without a contrast agent, and matched histologic observations. No vascular lesions were identified. There was no difference in the vascular density and architecture between control and OC specimens, but a striking difference in vascular pattern was seen at the OC-predilected site in the lateral ridge of the trochlea in all specimens, when compared to the medial ridge of the trochlea, where OC lesions are rarely observed. This site was less ossified with more perichondrial vessels not yet bridging with the subchondral bone. Furthermore, the mean vascular density of all specimens was significantly higher at this site. We speculate that joint morphology and focal internal trauma on this site with a unique vascular architecture may trigger ischemic events at this site. SWI permitted visualization of EGC in young foals with a clinical 3T MRI and paves the way for non-destructive longitudinal studies to improve understanding of OC in all species. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1539-1546, 2016.

Discontinuities in the endothelium of epiphyseal cartilage canals and relevance to joint disease in foals.

Cartilage canals have been shown to contain discontinuous blood vessels that enable circulating bacteria to bind to cartilage matrix, leading to vascular occlusion and associated pathological changes in pigs and chickens. It is also inconsistently reported that cartilage canals are surrounded by a cellular or acellular wall that may influence whether bacterial binding can occur. It is not known whether equine cartilage canals contain discontinuous endothelium or are surrounded by a wall. This study aimed to examine whether there were discontinuities in the endothelium of cartilage canal vessels, and whether canals had a cellular or acellular wall, in the epiphyseal growth cartilage of foals. Epiphyseal growth cartilage from the proximal third of the medial trochlear ridge of the distal femur from six healthy foals that were 1, 24, 35, 47, 118 and 122 days old and of different breeds and sexes was examined by light microscopy (LM), transmission electron microscopy (TEM) and immunohistochemistry. The majority of patent cartilage canals contained blood vessels that were lined by a thin layer of continuous endothelium. Fenestrations were found in two locations in one venule in a patent cartilage canal located deep in the growth cartilage and close to the ossification front in the 118-day-old foal. Chondrifying cartilage canals in all TEM-examined foals contained degenerated endothelial cells that were detached from the basement membrane, resulting in gap formation. Thirty-three percent of all canals were surrounded by a hypercellular rim that was interpreted as contribution of chondrocytes to growth cartilage. On LM, 69% of all cartilage canals were surrounded by a ring of matrix that stained intensely eosinophilic and consisted of collagen fibres on TEM that were confirmed to be collagen type I by immunohistochemistry. In summary, two types of discontinuity were observed in the endothelium of equine epiphyseal cartilage canal vessels: fenestrations were observed in a patent cartilage canal in the 118-day-old foal; and gaps were observed in chondrifying cartilage canals in all TEM-examined foals. Canals were not surrounded by any cellular wall, but a large proportion was surrounded by an acellular wall consisting of collagen type I. Bacterial binding can therefore probably occur in horses by mechanisms that are similar to those previously demonstrated in pigs and chickens.

Novel Application of Magnetic Resonance Imaging Demonstrates Characteristic Differences in Vasculature at Predilection Sites of Osteochondritis Dissecans.

BACKGROUND: Understanding the pathogenesis of osteochondrosis/osteochondritis dissecans and other developmental orthopaedic diseases that are thought to occur secondary to defects in vascular supply to growth/epiphyseal cartilage has been hampered by the inability to image the vasculature in this tissue. This is particularly true in human beings due to limitations of current imaging techniques and the lack of availability of appropriate cadaveric samples for histological studies. HYPOTHESIS: Susceptibility-weighted imaging, an MRI sequence, allows identification of characteristic differences in the vascular architecture in species that are affected by osteochondrosis/osteochondritis dissecans on the femoral condyle (humans and pigs) versus a species that is free of the disease (goat). STUDY DESIGN: Controlled laboratory study. MATERIALS: Distal femora from cadavers of juvenile humans (n = 5), pigs (n = 3), and goats (n = 3) were scanned in a 9.4-T MRI scanner using susceptibility-weighted imaging. Three-dimensional reconstructions were created, and minimum intensity projections were calculated in 3 planes to enhance visualization of the vascular architecture. RESULTS: Susceptibility-weighted imaging allowed clear visualization of the epiphyseal vasculature in all species. Vascular architecture, with vessels primarily arising from the perichondrium, was similar in humans and pigs, which are predisposed to osteochondrosis/osteochondritis dissecans, and was starkly different from that present in goats, a species in which there are no reports of osteochondrosis/osteochondritis dissecans. Furthermore, vessels in the distal femoral predilection site disappeared with age in humans in a pattern similar to that reported previously in pigs. CONCLUSION: Nearly identical vascular architecture at the shared primary predilection site of osteochondrosis/osteochondritis dissecans in the femoral condyles in human beings and pigs suggests that vascular failure, which is known to be central to the pathogenesis of this disease in pigs, may also play a role in humans. CLINICAL RELEVANCE: This assumption of a shared pathogenesis is supported by the pattern of disappearance of vessels with age at the primary predilection site of osteochondritis dissecans in humans, which is essentially identical to that which has been reported in pigs. Susceptibility-weighted imaging will likely help further elucidate this potential relationship in the future.

Reconstruction of the Pediatric Lateral Malleolus and Physis by Free Microvascular Transfer of the Proximal Fibular Physis.

Traumatic injury to the pediatric growth plate can result in growth disturbances, late angular deformity, and limb length inequalities. Complete traumatic loss the entire growth plate complex (physis, epiphysis, and distal metaphysis) of the ankle can lead to severe joint instability and loss of function. In the growing child, physeal preservation is paramount; however, the reconstructive options are limited. We report a case of post-traumatic loss of the distal fibular physis resulting in severe ankle valgus in a pediatric patient after a Gustilo grade 3B open injury. Ankle valgus secondary to post-traumatic necrosis of the lateral ankle physeal complex was successfully managed by microvascular free transfer of the ipsilateral proximal fibula physis. The 24-month follow-up examination demonstrated continued growth of the free vascularized physeal graft and a stable ankle. The donor site had healed without incident. The patient was able to return to age-appropriate play, sports, and social integration.

Surgical induction, histological evaluation, and MRI identification of cartilage necrosis in the distal femur in goats to model early lesions of osteochondrosis.

OBJECTIVE: Identify and interrupt the vascular supply to portions of the distal femoral articular-epiphyseal cartilage complex (AECC) in goat kids to induce cartilage necrosis, characteristic of early lesions of osteochondrosis (OC); then utilize magnetic resonance imaging (MRI) to identify necrotic areas of cartilage. DESIGN: Distal femora were perfused and cleared in goat kids of various ages to visualize the vascular supply to the distal femoral AECC. Vessels located on the axial aspect of the medial femoral condyle (MFC) and on the abaxial side of the lateral trochlear ridge were transected in eight 4- to 5-day-old goats to induce cartilage necrosis. Goats were euthanized 1, 2, 3, 4, 5, 6, 9, and 10 weeks post operatively and operated stifles were harvested. Adiabatic T1ρ relaxation time maps of the harvested distal femora were generated using a 9.4 T MR scanner, after which samples were evaluated histologically. RESULTS: Interruption of the vascular supply to the MFC caused lesions of cartilage necrosis in 6/8 goat kids that were demonstrated histologically. Adiabatic T1ρ relaxation time mapping identified these areas of cartilage necrosis in 5/6 cases. No significant findings were detected after transection of perichondrial vessels supplying the lateral trochlear ridge. CONCLUSIONS: Cartilage necrosis, characteristic of early OC, can be induced by interrupting the vascular supply to the distal femoral AECC in goat kids. The ability of high field MRI to identify these areas of cartilage necrosis in the AECC using the adiabatic T1ρ sequence suggests that this technique may be useful in the future for the early diagnosis of OC.

Comparison of the blood supply to the articular-epiphyseal growth complex in horse vs. pony foals.

REASONS FOR PERFORMING STUDY: To increase understanding of why the prevalence of clinical/radiographic osteochondrosis (OC) dissecans is high in horses and low in ponies. OBJECTIVES: To investigate whether the clinical difference in OC occurrence between horses and ponies could partly be explained by a difference in: 1) number of patent vessels in the epiphyseal growth cartilage; 2) duration of the presence of patent cartilage canals; or 3) growth cartilage thickness at predilection sites for OC. The hypothesis was that pony foals would have fewer cartilage canals, shorter duration of blood supply and thinner growth cartilage than horse foals. STUDY DESIGN: Observational, cross-sectional study. METHODS: Nine Standardbred foals (horse group) 1-49 days old and 11 Norwegian Fjord foals (pony group) 1-62 days old were included. A total of 15 anatomical locations in the tarsocrural and metatarsophalangeal joints were examined by one or more of the following techniques: arterial perfusion; photography of cleared specimens; microcomputed tomography; radiography; and histology. The number of cartilage canals was counted. Cartilage thickness was measured. Duration of blood supply was assessed in histological sections. RESULTS: Of the 3 common predilection sites for OC investigated, there were significantly fewer vessels (P = 0.003) and thinner cartilage (P = 0.002) at the distal lateral trochlear ridge of the talus in the pony group. There was no difference in the duration of presence of cartilage canals between the groups. CONCLUSION: The hypothesis that pony foals would have fewer cartilage canals and thinner growth cartilage than horse foals was confirmed for the lateral trochlear ridge of the talus. The current results may contribute towards an explanation for the low prevalence of OC at the distal lateral trochlear ridge of the talus in pony foals.

PTH receptor signaling in osteoblasts regulates endochondral vascularization in maintenance of postnatal growth plate.

Longitudinal growth of postnatal bone requires precise control of growth plate cartilage chondrocytes and subsequent osteogenesis and bone formation. Little is known about the role of angiogenesis and bone remodeling in maintenance of cartilaginous growth plate. Parathyroid hormone (PTH) stimulates bone remodeling by activating PTH receptor (PTH1R). Mice with conditional deletion of PTH1R in osteoblasts showed disrupted trabecular bone formation. The mice also exhibited postnatal growth retardation with profound defects in growth plate cartilage, ascribable predominantly to a decrease in number of hypertrophic chondrocytes, resulting in premature fusion of the growth plate and shortened long bones. Further characterization of hypertrophic zone and primary spongiosa revealed that endochondral angiogenesis and vascular invasion of the cartilage were impaired, which was associated with aberrant chondrocyte maturation and cartilage development. These studies reveal that PTH1R signaling in osteoblasts regulates cartilaginous growth plate for postnatal growth of bone.

Intra-epiphyseal stress injury of the proximal tibial epiphysis: preliminary experience of magnetic resonance imaging findings.

Stress induced injuries affecting the physeal plate or cortical bone in children and adolescents, especially young athletes, have been well described. However, there are no reports in the current English language literature of stress injury affecting the incompletely ossified epiphyseal cartilage. We present four cases of stress related change to the proximal tibial epiphysis (PTE) along with their respective magnetic resonance imaging (MRI) appearances ranging from subtle oedema signal to a pseudo-tumour like appearance within the epiphyseal cartilage. The site and pattern of intra-epiphyseal injury is determined by the type of tissue that is affected, the maturity of the skeleton and the type of forces that are transmitted through the tissue. We demonstrate how an awareness of the morphological spectrum of MRI appearances in intra-epiphyseal stress injury and the ability to identify concomitant signs of stress in other nearby structures can help reduce misdiagnosis, avoid invasive diagnostic procedures like bone biopsy and reassure patients and their families.

The potential role of VEGF-induced vascularisation in the bony repair of injured growth plate cartilage.

Growth plate injuries often result in undesirable bony repair causing bone growth defects, for which the underlying mechanisms are unclear. Whilst the key importance of pro-angiogenic vascular endothelial growth factor (VEGF) is well-known in bone development and fracture repair, its role during growth plate bony repair remains unexplored. Using a rat tibial growth plate injury repair model with anti-VEGF antibody, Bevacizumab, as a single i.p. injection (2.5 mg/kg) after injury, this study examined the roles of VEGF-driven angiogenesis during growth plate bony repair. Histology analyses observed isolectin-B4-positive endothelial cells and blood vessel-like structures within the injury site on days 6 and 14, with anti-VEGF treatment significantly decreasing blood-vessel-like structures within the injury site (P<0.05). Compared with untreated controls, anti-VEGF treatment resulted in an increase in undifferentiated mesenchymal repair tissue, but decreased bony tissue at the injury site at day 14 (P<0.01). Consistently, microcomputed tomography analysis of the injury site showed significantly decreased bony repair tissue after treatment (P<0.01). RT-PCR analyses revealed a significant decrease in osteocalcin (P<0.01) and a decreasing trend in Runx2 expression at the injury site following treatment. Furthermore, growth plate injury-induced reduced tibial lengthening was more pronounced in anti-VEGF-treated injured rats on day 60, consistent with the observation of a significantly increased height of the hypertrophic zone adjacent to the growth plate injury site (P<0.05). These results indicate that VEGF is important for angiogenesis and formation of bony repair tissue at the growth plate injury site as well as for endochondral bone lengthening function of the uninjured growth plate.

Hindlimb heating increases vascular access of large molecules to murine tibial growth plates measured by in vivo multiphoton imaging.

Advances in understanding the molecular regulation of longitudinal growth have led to development of novel drug therapies for growth plate disorders. Despite progress, a major unmet challenge is delivering therapeutic agents to avascular-cartilage plates. Dense extracellular matrix and lack of penetrating blood vessels create a semipermeable "barrier," which hinders molecular transport at the vascular-cartilage interface. To overcome this obstacle, we used a hindlimb heating model to manipulate bone circulation in 5-wk-old female mice (n = 22). Temperatures represented a physiological range of normal human knee joints. We used in vivo multiphoton microscopy to quantify temperature-enhanced delivery of large molecules into tibial growth plates. We tested the hypothesis that increasing hindlimb temperature from 22°C to 34°C increases vascular access of large systemic molecules, modeled using 10, 40, and 70 kDa dextrans that approximate sizes of physiological regulators. Vascular access was quantified by vessel diameter, velocity, and dextran leakage from subperichondrial plexus vessels and accumulation in growth plate cartilage. Growth plate entry of 10 kDa dextrans increased >150% at 34°C. Entry of 40 and 70 kDa dextrans increased <50%, suggesting a size-dependent temperature enhancement. Total dextran levels in the plexus increased at 34°C, but relative leakage out of vessels was not temperature dependent. Blood velocity and vessel diameter increased 118% and 31%, respectively, at 34°C. These results demonstrate that heat enhances vascular carrying capacity and bioavailability of large molecules around growth plates, suggesting that temperature could be a noninvasive strategy for modulating delivery of therapeutics to impaired growth plates of children.