Won Joon Yoo

Mobilization of endothelial progenitor cells in fracture healing and distraction osteogenesis

INTRODUCTION Fracture healing and distraction osteogenesis (DO) are unique postnatal bone formation processes, and neovascularization is critically required for successful bone regeneration. We investigated endothelial progenitor cell (EPC) mobilization during bone regeneration, and the possible contribution of EPCs to increased vascularization and new bone formation, especially in DO. METHODS Mouse tibia fracture and rat tibia DO models were used in this study. The proportion of EPCs among the peripheral and splenic mononuclear cells (MNCs) was determined by examining the endothelial lineage staining characteristics and EPC cell surface markers. Messenger RNA expression of molecules related to EPC mobilization and homing at the fracture site were analyzed by ribonuclease protection assay and reverse-transcription polymerase chain reaction. In the rat tibia DO model, we measured blood flow during DO, and determined the distribution of ex vivo-expanded and intravenously-infused EPCs. RESULTS The proportion of EPCs among the peripheral and splenic MNCs increased after fracture, peaked on post-fracture day 3, and returned to basal levels during the healing period. Messenger RNA expression of EPC mobilizing cytokines such as vascular endothelial growth factor (VEGF), stem cell factor, monocyte chemoattractant protein-1, and stromal cell-derived factor-1, were upregulated at the fracture callus. The plasma VEGF levels peaked prior to the increase in the EPC proportion. Adhesion molecules involved in EPC homing were expressed at the fracture callus. In the DO model, the temporal pattern of the increase in the EPC proportion was similar to that in the fracture healing model, but the EPC proportion increased again during the distraction and consolidation phases. The distraction gap was relatively ischemic during the distraction phase and blood flow increased profusely later in the consolidation phase. The number of EPCs homing to the bone regeneration site in the DO model correlated with the number of transplanted EPCs in a dose-dependent manner. CONCLUSIONS These findings suggest that signals from the bone regeneration site mobilize EPCs from the bone marrow into the peripheral circulation. Increased EPC mobilization and homing may contribute to neovascularization and thus to new bone formation in fracture healing and DO.

Reliability, Validity, and Responsiveness of a Modified International Knee Documentation Committee Subjective Knee Form (Pedi-IKDC) in Children With Knee Disorders

Background: The International Knee Documentation Committee (IKDC) Subjective Knee Form is a knee-specific measure of symptoms, function, and sports activity. A modified IKDC Subjective Knee Form (pedi-IKDC) has been developed for use in children and adolescents. The purpose of this study was to determine the psychometric characteristics of the pedi-IKDC in children and adolescents with knee disorders. Hypothesis: The pedi-IKDC is a reliable, valid, and responsive patient-administered outcome instrument in the pediatric population with knee disorders. Study Design: Cohort study (diagnosis); Level of evidence, 2. Methods: Test-retest reliability, content validity, criterion validity, construct validity, and responsiveness to change were determined for the pedi-IKDC in patients aged 10 to 18 years with a variety of knee disorders. Test-retest reliability was measured in a group of 72 patients with a stable knee disorder. Validity was measured in a group of 589 patients with the Child Health Questionnaire to determine criterion validity. Responsiveness was measured in a group of 98 patients undergoing a variety of knee surgical procedures. Results: The overall pedi-IKDC had acceptable test-retest reliability (intraclass correlation coefficient, .91) and excellent internal consistency (Cronbach alpha, .91). The form also demonstrated acceptable floor (0%) and ceiling (6%) effects. There was acceptable criterion validity with significant (P < .01) correlation between the overall pedi-IKDC and 9 relevant domains of the Child Health Questionnaire. Construct validity was acceptable, with all 11 hypotheses demonstrating significance (P < .0001). Responsiveness to change was acceptable (effect size, 1.39; standardized response mean, 1.35). Conclusion: The pedi-IKDC demonstrated overall acceptable psychometric performance for outcome assessment of children and adolescents with various disorders of the knee.

Growth plate disturbance after transphyseal reconstruction of the anterior cruciate ligament in skeletally immature adolescent patients: an MR imaging study.

Background There are concerns of potential growth disturbance after transphyseal reconstruction of the anterior cruciate ligament in skeletally immature patients. The authors used magnetic resonance (MR) imaging to evaluate growth disturbance and associated physeal abnormalities after index surgery. Methods We retrospectively reviewed the follow-up MR imaging studies of 43 patients who underwent transphyseal reconstruction of the anterior cruciate ligament using a soft-tissue graft at the mean age of 14.8 years (range, 12.4 to 16.5 y). Mean time from surgery to follow-up MR imaging was 16 months (range, 6 to 36 mo). Bone tunnel to growth plate cross-sectional area ratios were calculated as percentages. Focal growth disturbances were assessed in the follow-up MR images in terms of physeal tenting, the presence of a focal bone bridge, an asymmetric growth arrest line of Harris, and metaphyseal extension of physeal cartilage. Physeal angles with respect to the longitudinal axes of the corresponding bones were measured in preoperative MR images and compared with those measured in follow-up images. Premature physeal closure was assessed using the proximal fibular growth plate as an internal control. Clinically, growth disturbances were assessed with physical examinations regarding standing pelvic heights and alignments of the lower extremities. Results The bone tunnel to growth plate ratio was <3% for proximal tibia and distal femur. A focal bone bridge was observed in 5 patients—4 at the tibial physis and 1 at the femoral physis. Physeal angles did not change significantly during follow-up in either the coronal or sagittal plane. Earlier physeal closure than other physes was observed in 2 proximal tibiae. Clinically, there were no perceived growth disturbances. Conclusions MR imaging revealed that focal physeal disruption developed after index procedure in 5 of 43 adolescent patients (11.6%) without a perceived clinical growth disturbance. The results suggest that transphyseal reconstruction of the anterior cruciate ligament may not be a benign procedure that can be applied safely to younger children with substantial growth remaining. Level of Evidence Retrospective Case Series, Therapeutic Level IV.

Novel and recurrent TRPV4 mutations and their association with distinct phenotypes within the TRPV4 dysplasia family

Background Mutations in TRPV4, a gene that encodes a Ca2+ permeable non-selective cation channel, have recently been found in a spectrum of skeletal dysplasias that includes brachyolmia, spondylometaphyseal dysplasia, Kozlowski type (SMDK) and metatropic dysplasia (MD). Only a total of seven missense mutations were detected, however. The full spectrum of TRPV4 mutations and their phenotypes remained unclear. Objectives and methods To examine TRPV4 mutation spectrum and phenotype−genotype association, we searched for TRPV4 mutations by PCR-direct sequencing from genomic DNA in 22 MD and 20 SMDK probands. Results TRPV4 mutations were found in all but one MD subject. In total, 19 different heterozygous mutations were identified in 41 subjects; two were recurrent and 17 were novel. In MD, a recurrent P799L mutation was identified in nine subjects, as well as 10 novel mutations including F471del, the first deletion mutation of TRPV4. In SMDK, a recurrent R594H mutation was identified in 12 subjects and seven novel mutations. An association between the position of mutations and the disease phenotype was also observed. Thus, P799 in exon 15 is a hot codon for MD mutations, as four different amino acid substitutions have been observed at this codon; while R594 in exon 11 is a hotspot for SMDK mutations. Conclusion The TRPV4 mutation spectrum in MD and SMDK, which showed genotype−phenotype correlation and potential functional significance of mutations that are non-randomly distributed over the gene, was presented in this study. The results would help diagnostic laboratories establish efficient screening strategies for genetic diagnosis of the TRPV4 dysplasia family diseases.

Calcaneal lengthening for the planovalgus foot deformity in children with cerebral palsy.

The authors studied the outcomes of calcaneal lengthening for the treatment of planovalgus foot deformity in ambulatory children with cerebral palsy (92 feet in 56 children, mean age 9.2 years), attempting to define the surgical indication in terms of the severity of the foot deformity. Sixty-nine cases (75%) showed satisfactory clinical outcomes at an average follow-up of 5.2 years (range 4.0-17.2 years). Gait parameters such as foot progression angle, ankle motion in sagittal plane, and its power generation improved after operation. Preoperative talocalcaneal angle, talo-first metatarsal angle, and calcaneal pitch on weight-bearing lateral radiographs were predictive of the satisfactory results of the index operation. The authors conclude that calcaneal lengthening is an effective procedure for moderate to severe planovalgus foot deformities in children with cerebral palsy, but there is a limit under which the index operation can be performed safely: less than 35 degrees of talocalcaneal angle, less than 25 degrees of talo-first metatarsal angle, and more than 5 degrees of calcaneal pitch on weight-bearing lateral radiographs.

Interlocking telescopic rod for patients with osteogenesis imperfecta.

BACKGROUND Intramedullary fixation with use of a telescopic rod with a T-piece is one of the standard methods for long-bone stabilization in growing children with osteogenesis imperfecta. However, installation and removal of this device can cause substantial damage to the distal joint, which limits its use, especially in the tibia. We devised a modification of the telescopic rod system--the interlocking telescopic rod--in which the obturator is a simple rod with a hole, instead of a T-piece, at its distal end. METHODS The clinical and radiographic outcomes were evaluated more than two years following treatment of thirty-two limb segments (twenty-three tibiae and nine femora) with this new rod system in fifteen patients with osteogenesis imperfecta. RESULTS All rods were inserted without an arthrotomy of the distal joint, and all telescoped successfully. The interlocking pin used in the first five limb segments backed out between five and thirty-three months postoperatively. A revised fixation technique was used in the remaining twenty-seven limb segments, and the interlocking pin had not backed out at an average 3.1 years postoperatively. Proximal migration of the obturator was observed in four tibiae after 2.5 years. The cumulative survival rate of the rod at four years postoperatively was 88.7%. CONCLUSIONS Both insertion and removal of an interlocking telescopic rod are much less invasive than insertion and removal of a conventional telescopic rod with a T-piece anchor. The interlocking pin at the distal epiphysis provides effective anchorage for telescoping. Our interim results showed survival of the device to be comparable with, or better than, that of the conventional telescopic rod.

Expression and Role of Interleukin-6 in Distraction Osteogenesis

Distraction osteogenesis is a special form of bone healing in which well-controlled distraction stresses and consequent tensile strains within callus tissue induce very efficient new bone formation. Proinflammatory cytokines are involved during the early phase of fracture healing and callus remodeling. Temporal expression patterns of proinflammatory cytokines were assessed in Sprague-Dawley rat tibial models of distraction osteogenesis and acute lengthening, and only interleukin-6 (IL-6) was found to be specifically induced during the distraction phase. IL-6 immunoreactivity was detected not only in hemopoietic cells and osteoblasts but also in the spindle-shaped cells of the fibrous interzone, where most of the tensile strains are concentrated. In vitro study revealed that IL-6 did not affect the proliferation of C3H10T1/2 cells, mouse bone marrow stromal cells (MSCs), or MC3T3-E1 cells; but its blocking antibody reduced the proliferation of C3H10T1/2 cells and MSCs. The mRNA expression of COL1A1 and osteopontin were not changed by IL-6 or its blocking antibody, but the alkaline phosphatase activities of MC3T3-E1 cells were increased by IL-6 and decreased by its blocking antibody. These findings indicate that IL-6 is a proinflammatory cytokine that responds to tensile strain during distraction osteogenesis. IL-6 negatively affects the proliferation of primitive mesenchymal cells, whereas the differentiation of more mature osteoblastic lineage cells is enhanced by IL-6 in vitro. IL-6 appears to be one of the cytokines involved in the complex network of signal cascades evoked during distraction osteogenesis and may differentially affect immature and mature osteoblastic lineage cells.

Overgrowth syndrome associated with a gain-of-function mutation of the natriuretic peptide receptor 2 (NPR2) gene

The signal pathway of the C‐type natriuretic (CNP) and its receptor, natriuretic peptide receptor 2 (NPR2) is involved in the longitudinal growth of long bones. Loss of function mutations at NPR2 cause acromesomelic dysplasia, type Maroteaux, while overproduction of CNP by chromosomal translocation and a gain‐of‐function mutation at NPR2 have been reported to be responsible for an overgrowth syndrome in three cases and one family, respectively. We identified a four‐generation family with an overgrowth syndrome characterized by tall stature, macrodactyly of the great toes, scoliosis, coxa valga and slipped capital femoral epiphysis, similar to those previously reported in association with CNP/NPR2 overactivity. The serum level of amino‐terminal proCNP was normal in the proband. A novel missense mutation of NPR2, c.1462G>C (p.Ala488Pro) was found to co‐segregate with the phenotype in this family. In vitro transfection assay of the mutant NPR2 revealed overactivity of the mutant receptor at baseline as well as with the ligand. This overgrowth syndrome caused by a gain‐of‐function mutation at NPR2 should be differentiated from Marfan or related syndromes, and may be categorized along with the overgrowth syndrome caused by overproduction of CNP due to its phenotypical similarity as overgrowth CNP/NPR2 signalopathy.

Surgical treatment of the severe sequelae of infantile septic arthritis of the hip.

We retrospectively reviewed 45 hips in 43 patients with severe sequelae of infantile septic arthritis of the hip to compare the efficacy of various hip reconstructive and salvage surgeries, and to propose an algorithmic treatment protocol for the different types. Ten hips were classified as Choi Type IIIA, three as Type IIIB, 14 as Type IVA, and 18 as Type IVB sequelae. A total of 78 hip surgeries and 18 limb-length equalizations (three contralateral femoral epiphysiodesis and 15 ipsilateral femoral and/or tibial lengthenings) were done. The first surgical reconstructions were done at an average age of 5.9 years (range, 1.1–14.8 years), with a 9.5-year average followup. Type IIIA hips had better functional results than the other types. In Type III hips, early realignment osteotomy of the proximal femur or bone-grafting of the pseudarthrosis was indicated. In Type IVB hips, satisfactory results were observed in only five of 10 hips treated by trochanteric arthroplasty compared with satisfactory results in all four hips treated by Ilizarov’s hip reconstruction osteotomy. The latter operation seems to be better indicated in older patients with Type IVB hips, or with Type IVA hips in which previous reconstructive surgery was unsuccessful. Level of Evidence: Prognostic study, Level II-1 (retrospective study). See the Guidelines for Authors for a complete description of levels of evidence.

TRPV4-pathy manifesting both skeletal dysplasia and peripheral neuropathy: A report of three patients†‡

Heterozygous missense mutations of transient receptor potential vanilloid 4 channel (TRPV4) cause a spectrum of skeletal disorders, including brachyolmia, spondylometaphyseal dysplasia Kozlowski type, metatropic dysplasia, parastremmatic dysplasia, and spondyloepimetaphyseal dysplasia Maroteaux type. Similarly, heterozygous missense mutations of TRPV4 cause a spectrum of peripheral neuropathy, including hereditary motor and sensory neuropathy type IIC, congenital spinal muscular atrophy, and scapuloperoneal spinal muscular atrophy. There are no apparent differences in the amino acid positions affected or type of change predicted by the TRPV4 mutations responsible for the two disease spectrums; nevertheless, no fundamental phenotypic overlap has been shown between the two spectrums. Here, we report on three patients who had both skeletal dysplasia and peripheral neuropathy caused by heterozygous TRPV4 missense mutations. The skeletal and neurologic phenotypes of these patients covered the wide spectrum of reported TRPV4‐pathies (disease caused by TRPV4 mutations). The molecular data are complementary, proving that “neuropathic” mutations can cause skeletal dysplasia but also the “skeletopathic” mutations can lead to neuropathies. Our findings suggest that pathogenic mechanisms of TRPV4‐pathies in skeletal and nervous systems are not always mutually exclusive and provide further evidence that there is no clear genotype–phenotype correlation for either spectrum. Co‐occurrence of skeletal dysplasia and degenerative neuropathy should be kept in mind in clinical practice including diagnostic testing, surgical evaluation, and genetic counseling.