Generation and characterization of mesenchymal stem cells from the affected femoral heads of dogs with Legg Calvé Perthes disease.

Background: Canine Legg Calvé Perthes disease (LCPD) occurs during the growth period, and the cause of ischemic necrosis of the femoral head during growth remains unclear. If LCPD-affected femoral head-derived mesenchymal stem cells (LCPD-MSCs) can be generated, they can be used as a new tool for the pathophysiological analysis of canine LCPD. Aim: To generate affected femoral head-derived mesenchymal stem cells (MSCs) from dogs with LCPD and investigate the mRNA expression levels of angiogenesis-related factors and osteogenic differentiation potency of LCPD-MSCs. Methods: This study was performed using affected femoral heads from dogs diagnosed with LCPD and underwent femoral head and neck ostectomy. The necrotic tissue was harvested from the LCPD-affected femoral head and cultured statically (LCPD group, n = 6). Canine bone marrow-derived MSCs (BM-MSCs) were used as controls (control group, n = 6). First, the morphology of the cultured cells was observed, and the expression of CD29, CD34, CD44, CD45, CD90, and major histocompatibility complex class II was analyzed using flow cytometry. Additionally, the trilineage differentiation potency of the LCPD-affected head-derived adherent cells was examined. Furthermore, the expression levels of HIF1A, VEGFA, VEGFB, and PDGFB mRNAs and the bone differentiation potency of LCPD-affected head-derived adherent cells were investigated. Results: LCPD-affected femoral head-derived adherent cells showed a fibroblast-like morphology, and the expression of cell surface antigens was similar to that of BM-MSCs. In addition, LCPD-affected femoral head-derived adherent cells showed the same trilineage differentiation potency as BM-MSCs and were consistent with MSC characteristics. Furthermore, the mRNA expression levels of angiogenesis-related factors could be objectively measured in LCPD-MSCs and those MSCs had bone differentiation potency. Conclusion: In the present study, canine LCPD-MSCs were successfully generated, suggesting their usefulness as a tool for pathological analysis of LCPD in dogs.

Intravoxel incoherent motion (IVIM) detects femoral head ischemia in a piglet model of Legg-Calvé-Perthes disease.

There is a clinical need for alternatives to gadolinium contrast-enhanced magnetic resonance imaging (MRI) to facilitate early detection and assessment of femoral head ischemia in pediatric patients with Legg-Calvé-Perthes disease (LCPD), a juvenile form of idiopathic osteonecrosis of the femoral head. The purpose of this study was to determine if intravoxel incoherent motion (IVIM), a noncontrast-enhanced MRI method to simultaneously measure tissue perfusion and diffusion, can detect femoral head ischemia using a piglet model of LCPD. Twelve 6-week-old piglets underwent unilateral hip surgery to induce complete femoral head ischemia. The unoperated, contralateral femoral head served as a perfused control. The bilateral hips of the piglets were imaged in vivo at 3T MRI using IVIM and contrast-enhanced MRI 1 week after surgery. Median apparent diffusion coefficient (ADC) and IVIM parameters (diffusion coefficient: Ds; perfusion coefficient: Df; perfusion fraction: f; and perfusion flux: f*Df) were compared between regions of interest comprising the epiphyseal bone marrow of the ischemic and control femoral heads. Contrast-enhanced MRI confirmed complete femoral head ischemia in 11/12 piglets. IVIM perfusion fraction (f) and flux (f*Df) were significantly decreased in the ischemic versus control femoral heads: on average, f decreased 47 ± 27% (Δf = -0.055 ± 0.034; p = 0.0003) and f*Df decreased 50 ± 27% (Δf*Df = -0.59 ± 0.49 × 10-3 mm2/s; p = 0.0026). In contrast, IVIM diffusion coefficient (Ds) and ADC were significantly increased in the ischemic versus control femoral heads: on average, Ds increased 78 ± 21% (ΔDs = 0.60 ± 0.14 × 10-3 mm2/s; p < 0.0001) and ADC increased 60 ± 36% (ΔADC = 0.50 ± 0.23 × 10-3 mm2/s; p < 0.0001). In conclusion, IVIM is sensitive in detecting bone marrow ischemia in a piglet model of LCPD.

Exosomes from chondrocytes overexpressing miR-214-3p facilitate M2 macrophage polarization and angiogenesis to relieve Legg Calvé-Perthes disease.

OBJECTIVE: Legg-Calvé-Perthes disease (LCPD) is a partial or total necrosis of femoral head bone caused by blood supply disorder and its etiology is not clear. Studies have revealed that microRNA-214-3p (miR-214-3p) plays a vital role in LCPD, however, its exact mechanism is still unclear. In this study, we investigated the potential role of chondrocytes-derived exosomes carrying miR-214-3p (exos-miR-214-3p) in the pathogenesis of LCPD. METHODS: RT-qPCR was performed to evaluate miR-214-3p expression level in femoral head cartilage, serum and chondrocytes of patients with LCPD, as well as dexamethasone (DEX)-exposed TC28 cells. Effects of exos-miR-214-3p on the proliferation and apoptosis were verified via MTT assay, TUNEL staining and caspase3 activity assay. The M2 macrophage markers were assessed by flow cytometry, RT-qPCR and Western blot. Moreover, angiogenic effects of human umbilical vein endothelial cells (HUVECs) were tested using CCK-8 and tube formation assays. Bioinformatics prediction, luciferase assay and ChIP were applied to verify the association between ATF7, RUNX1 and miR-214-3p. RESULTS: miR-214-3p was found to be decreased in patients with LCPD and DEX-treated TC28 cells, of which overexpression promoted cell proliferation and suppressed apoptosis. Mechanistically, exos-miR-214-3p facilitated M2 polarization by ATF7/TLR4 axis and HUVECs angiogenesis via RUNX1/VEGFA axis. CONCLUSION: miR-214-3p alleviates LCPD by promoting M2 polarization of macrophages and angiogenesis.

Inhibition of Toll-Like Receptor 4 Signaling Pathway Accelerates the Repair of Avascular Necrosis of Femoral Epiphysis through Regulating Macrophage Polarization in Perthes Disease.

BACKGROUND: Legg-Calvé-Perthes disease (LCPD) is still a refractory disease in children's orthopedics. With the introduction of the concept of "osteoimmunology", the immune-inflammatory mechanisms between bone and immune system have become a research focus of LCPD. However, few studies have reported on the pathological role of inflammation-related receptors such as toll-like receptors (TLRs) as well as immune cells such as macrophages in LCPD. This study was for investigating the mechanism of TLR4 signaling pathway on the direction of macrophage polarization and the repair of avascular necrosis of femoral epiphysis in LCPD. METHODS: With GSE57614 and GSE74089, differentially expressed genes were screened. Through enrichment analysis and protein-protein interaction network, the functions of TLR4 were explored. Furthermore, immunohistochemistry, enzyme-linked immunosorbent assay (ELISA), hematoxylin & eosin (H&E) staining, micro-CT, tartrate-resistant acid phosphatase (TRAP) dyeing and western blotting were performed for determining the influences of TAK-242 (a TLR4 inhibitor) on the repair of avascular necrosis of femoral epiphysis in rat models. RESULTS: Totally 40 co-expression genes were screened as well as enriched in TLR4 signaling pathway. Immunohistochemistry and ELISA analyses certified that TLR4 facilitated macrophage polarization toward the M1 phenotype and prevented macrophage polarization toward the M2 phenotype. Besides, the results of H&E and TRAP staining, micro-CT, and western blotting showed that TAK-242 can inhibit osteoclastogenesis and promote osteogenesis. CONCLUSION: Inhibition of TLR4 signaling pathway accelerated the repair of avascular necrosis of femoral epiphysis by regulating macrophage polarization in LCPD.

Effects of acute femoral head ischemia on the growth plate and metaphysis in a piglet model of Legg-Calvé-Perthes disease.

OBJECTIVE: To determine the effects of acute (≤7 days) femoral head ischemia on the proximal femoral growth plate and metaphysis in a piglet model of Legg-Calvé-Perthes disease (LCPD). We hypothesized that qualitative and quantitative histological assessment would identify effects of ischemia on endochondral ossification. DESIGN: Unilateral femoral head ischemia was surgically induced in piglets, and femurs were collected for histological assessment at 2 (n = 7) or 7 (n = 5) days post-ischemia. Samples were assessed qualitatively, and histomorphometry of the growth plate zones and primary spongiosa was performed. In a subset of samples at 7 days, hypertrophic chondrocytes were quantitatively assessed and immunohistochemistry for TGFß1 and Indian hedgehog was performed. RESULTS: By 2 days post-ischemia, there was significant thinning of the proliferative and hypertrophic zones, by 63 µm (95% CI -103, -22) and -19 µm (95% CI -33, -5), respectively. This thinning persisted at 7 days post-ischemia. Likewise, at 7 days post-ischemia, the primary spongiosa was thinned to absent by an average of 311 µm (95% CI -542, -82) in all ischemic samples. TGFß1 expression was increased in the hypertrophic zone at 7 days post-ischemia. CONCLUSIONS: Alterations to the growth plate zones and metaphysis occurred by 2 days post-ischemia and persisted at 7 days post-ischemia. Our findings suggest that endochondral ossification may be disrupted at an earlier time point than previously reported and that growth disruption may occur in the piglet model as occurs in some children with LCPD.

Physical disability in Late Antiquity Milan: slipped capital femoral epiphysis with severe secondary joint disease in the Basilica of San Dionigi.

The paper presents the skeletal remains of an adult male of 30-40 years with bone lesions and deformity on the left hip, recovered during the archaeological excavation below the former Basilica of San Dionigi, dated to Late Antiquity (3rd - 5th century AD) Milan. Biological profile and paleopathological analysis were performed following standard references and the bones underwent radiological examination. Differential diagnosis included congenital anomaly, active rickets, infectious diseases, femoral neck trauma, Legg-Carvé-Perthes disease, Slipped Capital Femoral Epiphysis (SFCE), osteogenesis imperfecta and osteoporosis. While the lesions were highly consistent with Legg-Calvé-Perthes disease, they were typical of SFCE, complicated by an avascular necrosis and secondary osteoarthrosis. The alteration of the femoral head led to a 7.8% leg-length discrepancy, causing gait alteration with partial compensation though increased muscular activity on the right leg, reduced mobility of the joint and potentially hip pain, difficulty in walking and running and even limping. This paper explores a case of physical disability from the Roman era found near a Christian place of worship and represents a rare case of SFCE in the paleopathological record.

Quantitative T2 and T1ρ mapping are sensitive to ischemic injury to the epiphyseal cartilage in an in vivo piglet model of Legg-Calvé-Perthes disease.

OBJECTIVE: To determine if the quantitative MRI techniques T2 and T1ρ mapping are sensitive to ischemic injury to epiphyseal cartilage in vivo in a piglet model of Legg-Calvé-Perthes disease using a clinical 3T MRI scanner. We hypothesized that T2 and T1ρ relaxation times would be increased in the epiphyseal cartilage of operated vs contralateral-control femoral heads 1 week following onset of ischemia. DESIGN: Unilateral femoral head ischemia was surgically induced in eight piglets. Piglets were imaged 1 week post-operatively in vivo at 3T MRI using a magnetization-prepared 3D fast spin echo sequence for T2 and T1ρ mapping and a 3D gradient echo sequence for cartilage segmentation. Ischemia was confirmed in all piglets using gadolinium contrast-enhanced MRI. Median T2 and T1ρ relaxation times were measured in the epiphyseal cartilage of the ischemic and control femoral heads and compared using paired t-tests. Histological assessment was performed on a subset of five piglets. RESULTS: T2 and T1ρ relaxation times were significantly increased in the epiphyseal cartilage of the operated vs control femoral heads (ΔT2 = 11.9 ± 3.7 ms, 95% CI = [8.8, 15.0] ms, P < 0.0001; ΔT1ρ = 12.8 ± 4.1 ms, 95% CI = [9.4, 16.2] ms, P < 0.0001). Histological assessment identified chondronecrosis in the hypertrophic and deep proliferative zones within ischemic epiphyseal cartilage. CONCLUSIONS: T2 and T1ρ mapping are sensitive to ischemic injury to the epiphyseal cartilage in vivo at clinical 3T MRI. These techniques may be clinically useful to assess injury and repair to the epiphyseal cartilage to better stage the extent of ischemic damage in Legg-Calvé-Perthes disease.

Legg-Calvé-Perthes disease overview.

BACKGROUND: Legg-Calvé-Perthes Disease (LCPD) is a necrosis of the femoral head which affects the range of motion of the hips. Its incidence is variable, ranging from 0.4/100,000 to 29.0/ 100,000 children. Although LCPD was first described in the beginning of the past century, limited is known about its etiology. Our objective is to describe the main areas of interest in Legg-Calve-Perthes disease. METHODS: A review of the literature regarding LCPD etiology was performed, considering the following inclusion criteria: Studies reporting clinical or preclinical results. The research group carried out a filtered search on the PubMed and Science Direct databases. To maximize the suitability of the search results, we combined the terms ''Perthes disease" OR "LCPD" OR "children avascular femoral head necrosis" with "diagnostic" OR "treatment" OR "etiology" as either key words or MeSH terms. RESULTS: In this article been described some areas of interest in LCPD, we include topics such as: history, incidence, pathogenesis, diagnosis, treatment and possible etiology, since LCPD has an unknown etiology. CONCLUSIONS: This review suggests that LCPD has a multifactorial etiology where environmental, metabolic and genetic agents could be involved.

T1ρ and T2 mapping detect acute ischemic injury in a piglet model of Legg-Calvé-Perthes disease.

This study investigated the sensitivity of T1ρ and T2 relaxation time mapping to detect acute ischemic injury to the secondary ossification center (SOC) and epiphyseal cartilage of the femoral head in a piglet model of Legg-Calvé-Perthes disease. Six piglets underwent surgery to induce global right femoral head ischemia and were euthanized 48 h later. Fresh operated and contralateral-control femoral heads were imaged ex vivo with T1, T2, and T1ρ mapping using a 9.4T magnetic resonance imaging scanner. The specimens were imaged a second time after a freeze/thaw cycle and then processed for histology. T1, T2, and T1ρ measurements in the SOC, epiphyseal cartilage, articular cartilage, and metaphysis were compared between operated and control femoral heads using paired t tests. The effects of freeze/thaw, T1ρ spin-lock frequency, and fat saturation were also investigated. Five piglets with histologically confirmed ischemic injury were quantitatively analyzed. T1ρ was increased in the SOC (101 ± 15 vs. 73 ± 16 ms; p = 0.0026) and epiphyseal cartilage (84.9 ± 9.2 vs. 74.3 ± 3.6 ms; p = 0.031) of the operated versus control femoral heads. T2 was also increased in the SOC (28.7 ± 2.0 vs. 22.7 ± 1.7; p = 0.0037) and epiphyseal cartilage (57.4 ± 4.7 vs. 49.0 ± 2.7; p = 0.0041). No changes in T1 were detected. The sensitivities of T1ρ and T2 mapping in detecting ischemic injury were maintained after a freeze/thaw cycle, and T1ρ sensitivity was maintained after varying spin-lock frequency and applying fat saturation. In conclusion, T1ρ and T2 mapping are sensitive in detecting ischemic injury to the SOC and epiphyseal cartilage of the femoral head as early as 48 h after ischemia induction.

Retroversion of the hemipelvis rather than hypoplastic posterior wall decreases acetabular anteversion in hips affected by Perthes disease.

The acetabular retroversion has a moderate incidence of 31-60% in all patients of the Perthes disease. It might be caused by posterior wall dysplasia based on recent animal researches. However, some studies support that hemipelvic retroversion is the main factor for the acetabular retroversion. The primary pathological factor of increasing retroversion angle is still controversial anatomically. This study aimed to identify whether there is acetabular retroversion in children with Perthes disease,and to find a method to distinguish version types. Forty children with unilateral Perthes disease who were admitted to our hospital from January 1, 2012 to December 31, 2018 were enrolled, and 40 controls were matched based on sex and age. The acetabular anteversion angle (AAA), internal wall anteversion angle (IWAA), anterior wall height of the acetabulum (A), acetabular posterior wall height (P), and acetabular width (W) were assessed on computed tomography (CT) at the level of the femoral head center. The acetabular wall difference index (AWDI; AWDI = P-A)/W*100) was calculated. The mean AAA was significantly lower in Perthes disease hips (10.59 (8.05-12.46)) than in contralateral hips (12.04 (9.02-13.33)) (p = 0.002) but did not differ from control hips (9.68 ± 3.76) (p = 0.465). The mean IWAA was significantly lower in Perthes hips (9.16 ± 3.89) than in contralateral hips (11.31 ± 4.04) (p = 0.000) but did not differ from control hips (9.43 ± 3.82) (p = 0.753). The mean AWDI did not differ between Perthes hips (0.41 ± 4.94) and contralateral hips (- 1.12 (- 4.50, 2.17)) (p = 0.06) or control hips (- 0.49 ± 5.46) (p = 0.437). The mean W was significantly higher in Perthes hips (44.61 ± 5.06) than in contralateral hips (43.36 ± 4.38) (p = 0.000) but did not differ from control hips (45.02 ± 5.01) (p = 0.719). The mean A and P did not differ between Perthes hips and contralateral hips or control hips. Correlation analysis of all hip joints revealed a significant correlation between AAAs and IWAAs (r = 0.772; r = 0.643; r = 0.608; and r = 0.540). Linear regression analysis revealed that AAAs increased with IWAAs. Multiple linear regression showed that IWAAs and AWDIs have good predictive value for AAAs in both Perthes and control hips (R2 = 0.842, R2 = 0.869). In patients with unilateral Perthes disease, the affected acetabulum is more retroverted than that on the contralateral side, which may be caused by hemipelvic retroversion. The measurements in this study could distinguish the form of acetabular retroversion. IWAAs and AWDIs can be used as new observations in future studies of acetabular version.

Hip Morphology in Early-stage LCPD: Is There an Argument for Anatomic-specific Containment?

BACKGROUND: Early containment surgery has become increasingly popular in Legg-Calvé-Perthes Disease (LCPD), especially for older children. These procedures treat the proximal femur, the acetabulum, or both, and most surgeons endorse the same surgical option regardless of an individual patient's anatomy. This "one-surgery-fits-all" approach fails to consider potential variations in baseline anatomy that may make one option more sensible than another. We sought to describe hip morphology in a large series of children with newly diagnosed LCPD, hypothesizing that variation in anatomy may support the concept of anatomic-specific containment. METHODS: A retrospective review of a prospectively collected multicenter database was conducted for patients aged 6 to 11 at diagnosis. To assess anatomy before significant morphologic changes secondary to the disease itself, only patients in Waldenström stages IA/IB were included. Standard hip radiographic measurements including acetabular index, lateral center-edge angle, proximal femoral neck-shaft angle (NSA), articulotrochanteric quartiles, and extrusion index (EI) were made on printed anteroposterior pelvis radiographs. Age-specific percentiles were calculated for these measures using published norms. Significant outliers (≤10th/≥90th percentile) were reported where applicable. RESULTS: A total of 168 patients with mean age at diagnosis of 8.0±1.3 years met inclusion criteria (81.5% male). Mean acetabular index for the entire cohort was 16.8±4.1 degrees; 58 hips (34.5%) were significantly dysplastic compared with normative data. Mean lateral center-edge angle was 15.9±5.2 degrees at diagnosis; 110 (65.5%) were ≤10th percentile indicating dysplasia (by this metric). Mean NSA overall was 136.5±7.0 degrees. Fifty-one (30.4%) and 20 (11.9%) hips were significantly varus (≤10th percentile) or valgus (≥90th percentile), respectively. Thirty-five hips (20.8%) were the third articulo-trochanteric quartiles or higher suggesting a higher-riding trochanter at baseline. Mean EI was 15.5%±9.0%, while 63 patients (37.5%) had an EI ≥20%. CONCLUSIONS: The present study finds significant variation in baseline anatomy in children with early-stage LCPD, including a high prevalence of coexisting acetabular dysplasia as well as high/low NSAs. These variations suggest that the "one-surgery-fits-all" approach may lack specificity for a particular patient; a potentially wiser option may be an anatomic-specific containment operation (eg, acetabular-sided osteotomy for coexisting dysplasia, varus femoral osteotomy for valgus NSA). LEVEL OF EVIDENCE: Level IV.

Increased circulating CD31+/CD42b-EMPs in Perthes disease and inhibit HUVECs angiogenesis via endothelial dysfunction.

AIMS: Endothelial microparticles (EMPs) are extracellular vesicles secreted by endothelial cells. The purpose of this research is to explore that the clinical significance and roles in angiogenesis and endothelial dysfunction of circulating microparticles in Perthes disease. MAIN METHODS: We collected platelet-poor plasma (PPP) from patients and controls, then microparticles (MPs) were extracted. Flow cytometry was performed to calculate the concentrations of CD31+/CD42b-, CD62E+ and CD31+/CD42b+ MPs. ELISA was performed to detect the expression level of biomarkers of endothelial dysfunction and inflammatory factors in plasma. In vitro experiments to evaluate the effect of circulating MPs and EMPs derived from IL-6-stimulated human umbilical vein endothelial cells (HUVECs) on angiogenesis and endothelial dysfunction. KEY FINDINGS: Our results revealed that the CD31+/CD42b- EMPs were significantly higher in Perthes disease group than in the control group. The Perthes-MPs being taken up by HUVECs promoted endothelial cell apoptosis, endothelial dysfunction and inhibited angiogenesis in vitro. Moreover, the level of IL-6 in plasma significantly increased in patients with Perthes, which was tightly correlated with the elevated level of circulating CD31+/CD42b- EMPs. IL-6 promoted HUVECs to secrete CD31+/CD42b- MPs, and EMPs derived from high concentration IL-6-stimulated (100 and 1000 pg/mL) HUVECs promoted endothelial cell apoptosis, endothelial dysfunction and inhibited angiogenesis. SIGNIFICANCE: In summary, our study suggests that circulating EMPs in the phenotypic spectrum revealed unique phenotypes of endothelial dysfunction, showing close correlation with the secretion of IL-6. These circulating EMPs may give rise to endothelial cell apoptosis, endothelial dysfunction and angiogenesis in Perthes disease.

A Comparison of Transphyseal Neck-Head Tunneling and Multiple Epiphyseal Drilling on Femoral Head Healing Following Ischemic Osteonecrosis: An Experimental Investigation in Immature Pigs.

BACKGROUND: Two operative procedures are currently advocated to stimulate the necrotic femoral head healing in children with Legg-Calve-Perthes disease: transphyseal neck-head tunneling (TNHT) and multiple epiphyseal drilling (MED). The purpose of this study was to compare the bone healing and physeal function after treatment with TNHT or MED in a piglet model of ischemic osteonecrosis. METHODS: Eighteen piglets were induced with osteonecrosis by surgically placing a ligature tightly around the right femoral neck. One week later, the piglets were assigned to 1 of 3 treatment groups (n=6/group): (1) local nonweight bearing only (NWB), (2) TNHT plus NWB, or (3) MED plus NWB. The unoperated left femoral heads were used as normal controls. The animals were euthanized at 8 weeks after osteonecrosis induction. Histologic, histomorphometric, radiographic, microcomputed tomography (CT), and calcein-labeling assessments were performed. Statistical analysis included a 1-way ANOVA. RESULTS: Micro-CT analyses showed higher femoral head bone volume in the MED group compared with the TNHT and the NWB groups (P<0.01). The MED group had a higher mean trabecular number (P<0.001) and new bone formation (P=0.001) based on calcein-labeling parameters compared with the TNHT and the NWB groups. In addition, the osteoclast number per bone surface was lower in the MED group compared with the NWB group (P=0.001). Histologic and micro-CT assessments of the proximal femoral physis revealed a larger physeal disruption at the site of physeal drilling in the TNHT group compared with the MED group. However, no significant differences in physeal elongation (P=0.61) and femoral neck length (P=0.31) were observed between the treatment groups. CONCLUSIONS: MED produced a higher bone volume and stimulated greater bone formation than the TNHT or the NWB alone. Both procedures did not produce a significant physeal growth disturbance during the study period. CLINICAL RELEVANCE: This preclinical study provides evidence that MED produces more favorable bone healing than the TNHT in a large animal model of Legg-Calve-Perthes disease.

The role and underlying mechanisms of microRNA­214 in Legg­Calvé­Perthes disease.

Legg­Calvé­Perthes disease (LCPD) is a pediatric form of femoral head osteonecrosis with unknown etiology. MicroRNAs (miRs) have been revealed to serve an important role in LCPD. MiR­214 serves an important role in chondrogenesis. The aim of the present study was to investigate the potential role of miR­214 in LCPD and the underlying mechanisms. The expression levels of miR­214 and B­cell lymphoma 2 (Bcl­2)­associated X protein (Bax) in dexamethasone (DEX)­treated TC28 cells, and the femoral head cartilage tissues, serum and primary chondrocytes of patients with LCPD, and healthy individuals were determined via reverse transcription quantitative polymerase chain reaction and western blot analysis. A luciferase reporter assay was conducted to investigate the association between miR­214 and Bax, while cell viability was determined via an MTT assay, and flow cytometry was performed to investigate cell apoptosis. The results revealed that miR­214 was downregulated and Bax was upregulated in DEX­treated TC28 cells and tissues obtained from patients with LCPD. MiR­214 was demonstrated to directly target Bax and negatively regulate its expression. DEX administration significantly suppressed cell proliferation, promoted apoptosis and decreased the Bcl­2/Bax ratio in TC28 cells; overexpression of miR­214 induced opposing effects, which were reversed by Bax overexpression. In conclusion, the results indicated that miR­214 and Bax may be potential therapeutic targets for the future clinical treatment of LCPD.

The possible origin of Legg-Calvé-Perthes disease / El posible origen de la enfermedad de Legg-Calvé-Perthes

Legg-Calvé-Perthes disease (LCPD) is a childhood orthopedic pathology that affects the development of the hip. It is a rare disease with a huge variation in annual incidence. It occurs approximately five times more often in boys than in girls. The objective of this article was to formulate a hypothesis about the possible origin of LCPD, from the paleopathological findings of this disease reported until 2017, and to highlight the importance of anthropology, history, paleontology and paleopathology to the study of the origins of disease. By using eight web-based search engines, we performed a review of articles focused on the history, genetics and paleopathological findings of LCPD; we evaluated in total 133 articles published between 1910 and 2017. Out of these, 20 articles belonging to the same publication period were included in this analysis. LCPD was described for the first time approximately 100 years ago and without knowing it was a new disease. In the last years, human remains have been found in which LCPD has been identified, providing relevant information about the origin of this pathology. These data and their historical context can be a basis to propose the Asian continent as the site of origin of LCPD; however, new anthropological, genetic and paleopathological studies are needed to reinforce or refute this hypothesis.

La enfermedad de Legg-Calvé-Perthes (ELCP) es una afección ortopédica infantil que repercute en el desarrollo de la cadera. Es una enfermedad rara con incidencia anual variable. Es aproximadamente cinco veces más frecuente en niños que en niñas. El objetivo de este artículo fue formular una hipótesis acerca del posible origen de la ELCP a partir de hallazgos paleopatológicos reportados hasta el año 2017, además de resaltar la importancia que ofrecen la antropología, la historia, la paleontología y la paleopatología para el estudio del origen de las enfermedades. Mediante ocho buscadores se hizo una revisión de artículos referentes a la historia, la genética y los hallazgos paleopatológicos de la ELCP; se evaluaron un total de 133 artículos publicados entre 1910 y 2017. De ellos, fueron incluidos en este análisis 20 artículos que abarcaron el mismo periodo de publicación. La ELCP comenzó a describirse hace poco más de 100 años y sin el conocimiento de que se trataba de una entidad nueva. En los últimos años se han encontrado restos humanos en los que se ha identificado la ELCP, lo cual ha brindado información relevante respecto al origen de este padecimiento. Estos datos y su contexto histórico pueden ser fundamentos para plantear al continente asiático como el sitio de origen de la ELCP; sin embargo, se requiere de nuevos estudios antropológicos, genéticos y paleopatológicos para reforzar o refutar esta hipótesis.

Analysis of Trabecular Microstructure and Vascular Distribution of Capital Femoral Epiphysis Relevant to Legg-Calve-Perthes Disease.

Legg-Calve-Perthes disease is characterized by the capital femoral epiphyseal collapse, which occurs more reliably in the anterior quadrant than the more weight-bearing lateral quadrant. The purpose of this study was to determine whether there is a vascular or microstructural predisposition for anterior femoral epiphyseal collapse in Perthes disease. Thirty-two cadaveric proximal femoral epiphyses from 17 subjects (age 4-14 years old) underwent micro-computed tomography at 10-µm resolution. Each quadrant was analyzed for four markers of trabecular architecture: bone volume fraction (BV/TV), trabecular thickness, trabecular separation (TbSp), and trabecular number (TbN). Vascular channels were then mapped in each quadrant, identified by correlating surface topography with cross-sectional imaging. One-way analysis of variance revealed an overall difference between quadrants (p < 0.001) in BV/TV, TbN, and TbSp. However, post hoc analysis revealed there was no significant difference between the anterior and lateral quadrants for any of the four markers of trabecular architecture. Vascular channel mapping illustrated a predominance of vessels in the posterior half of the epiphysis compared to the anterior half (8.7 ± 4.0 vs. 3.4 ± 3.1 vascular channels, p < 0.001). The lack of microstructural differences between the anterior and lateral quadrants, and the predominance of vascular channels in the posterior half of the epiphysis with posteriorly-based medial femoral circumflex and ligamentum teres vessels suggests that the anterior femoral epiphysis may be a relative vascular watershed region, which predisposes it to collapse after the vascular insult of Perthes disease. Clinical significance: Improved understanding of the pathophysiology of anterior femoral epiphyseal collapse may inform future treatments aimed at revascularization. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:1784-1789, 2019.

Determining Hinge Abduction in Legg-Calvé-Perthes Disease: Can We Reliably Make the Diagnosis?

BACKGROUND: Although hinge abduction is recognized as an important finding in children with Legg-Calvé-Perthes disease, variable diagnostic criteria exist. The purpose of this study was (1) to test the interobserver and intraobserver agreement of the current definition of hinge abduction and (2) to develop consensus regarding key diagnostic features that could be used to improve our diagnostic criteria. METHODS: Four orthopaedic surgeons with subspecialty pediatric hip interest independently assessed 30 randomly ordered cases of Legg-Calvé-Perthes disease. Each case included 2 fluoroscopic images of hip arthrograms (anteroposterior and abduction views). Surgeons graded the cases in a binary manner (hinge/no-hinge) on 2 separate occasions separated by a 4-week interval. Following reliability testing and comprehensive review of the literature, consensus-building sessions were conducted to identify key diagnostic features. Surgeons then regraded a new series of cases. Interobserver and intraobserver agreement between first/second and third/fourth readings were assessed using the Fleiss κ. RESULTS: Interobserver κ for hinge abduction between the first and second surveys was 0.52 (with 0.41 to 0.60 considered moderate agreement), compared with 0.56 for the third and fourth surveys. First and second reading intraobserver agreement ranged from 0.59 to 0.83 compared with 0.75 to 1.00 for third and fourth reading. Consensus sessions identified several key diagnostic factors including: adequate visualization of the labral contour and ability of the lateral epiphysis to slip below the chondrolabral complex in abduction. Medial dye pooling, often due to asphericity of the femoral head, was not found to be a useful diagnostic criterion. CONCLUSIONS: Despite a combined experience of over 70 years among the reviewers, we found just slightly better than 50:50 agreement in what constitutes hinge abduction. Consensus discussions did improve our agreement but these modest changes emphasize how difficult it is to develop reliable diagnostic criteria for hinge abduction. As a result, we caution against using hinge abduction as an inclusion criteria or outcome measure for research purposes, as the diagnostic agreement can be inconsistent. LEVEL OF EVIDENCE: Level III-diagnostic study.

Quantitative MRI Helps to Detect Hip Ischemia: Preclinical Model of Legg-Calvé-Perthes Disease.

Purpose To determine whether quantitative MRI relaxation time mapping techniques can help to detect ischemic injury to the developing femoral head. Materials and Methods For this prospective animal study conducted from November 2015 to February 2018, 10 male 6-week-old piglets underwent an operation to induce complete right femoral head ischemia. Animals were humanely killed at 48 hours (n = 2) or 4 weeks (n = 8) after the operation, and the operated and contralateral-control femoral heads were harvested and frozen. Thawed specimens were imaged at 9.4-T MRI by using T1, T2, T1 in the rotating frame (T1ρ), adiabatic T1ρ, relaxation along a fictitious field (RAFF), and T2* mapping and evaluated with histologic analysis. Paired relaxation time differences between the operated and control femoral heads were measured in the secondary ossification center (SOC), epiphyseal cartilage, articular cartilage, and metaphysis and were analyzed by using a paired t test. Results In the SOC, T1ρ and RAFF had the greatest percent increases in the operated versus control femoral heads at both 48 hours (112% and 72%, respectively) and 4 weeks (74% and 70%, respectively). In the epiphyseal and articular cartilage, T2, T1ρ, and RAFF were similarly increased at both points (range, 24%-49%). At 4 weeks, T2, T1ρ, adiabatic T1ρ, and RAFF were increased in the SOC (P = .004, .018, < .001, and .001, respectively), epiphyseal cartilage (P = .009, .008, .011, and .007, respectively), and articular cartilage (P = .005, .016, .033, and .018, respectively). Histologic assessment identified necrosis in SOC and deep layer of the epiphyseal cartilage at both points. Conclusion T2, T1 in the rotating frame, adiabatic T1 in the rotating frame, and relaxation along a fictitious field maps are sensitive in helping to detect ischemic injury to the developing femoral head. © RSNA, 2018 Online supplemental material is available for this article.

Natural History of Structural Hip Abnormalities and the Potential for Hip Preservation.

Hip osteoarthritis (OA) exerts a significant burden on society, affecting 3% of Americans aged >30 years. Recent advances in the understanding of the pathoanatomy and pathomechanics of the hip have led to treatment options for young adults with hip pain. Femoroacetabular impingement, specifically cam-type femoroacetabular impingement, hip dysplasia, and the sequelae of pediatric hip disease can predispose the hip to early OA. However, many patients with abnormal anatomic findings do not develop early OA, suggesting that there exist other patient characteristics that are protective despite abnormal bony anatomy. Outcome studies show that arthroscopic and open hip procedures improve pain and function in patients with symptomatic hips. However, there is currently limited evidence that these procedures extend the life of the patient's natural hip. Additional studies are needed to determine protective or adaptive factors in patients with abnormal anatomy who do not develop early OA and to determine whether joint preserving hip surgery extends the life of the native hip joint.

The potential role of variations in juvenile hip geometry on the development of Legg-Calvé-Perthes disease: a biomechanical investigation.

Legg-Calvé-Perthes disease (LCP) is one of the most poorly understood diseases in paediatric orthopaedics. One common trait of LCP is the marked morphological difference between healthy and pathological hips, early deviations of which (i.e. prior to disease onset) have been suggested to lead to the overload and collapse of the epiphysis. Here, the impact of common variations in geometry is investigated with a finite element model of a juvenile femur under single leg standing and landing. Here, the impact of typical variations in geometry is investigated with a finite element model of a juvenile femur under single leg standing and landing. The variations appear to have only a limited effect on the stress distribution in the femoral epiphysis even during high impact activities. This suggests that, for this individual at least, they would be unlikely to cause epiphyseal overload and collapse, even in the presence of a skeletally immature epiphysis.