Can DNA Methylation Profiling Classify Histologic Subtypes and Grades in Soft Tissue Sarcoma?

BACKGROUND: A clear classification of the subtype and grade of soft tissue sarcoma is important for predicting prognosis and establishing treatment strategies. However, the rarity and heterogeneity of these tumors often make diagnosis difficult. In addition, it remains challenging to predict the response to chemotherapy and prognosis. Thus, we need a new method to help diagnose soft tissue sarcomas and determine treatment strategies in conjunction with traditional methods. Genetic alterations can be found in some subtypes of soft tissue sarcoma, but many other types show dysregulated gene expression attributed to epigenetic changes, such as DNA methylation status. However, research on DNA methylation profiles in soft tissue sarcoma is still insufficient to provide information to assist in diagnosis and therapeutic decisions. QUESTIONS/PURPOSES: (1) Do DNA methylation profiles differ between normal tissue and soft tissue sarcoma? (2) Do DNA methylation profiles vary between different histologic subtypes of soft tissue sarcoma? (3) Do DNA methylation profiles differ based on tumor grade? METHODS: Between January 2019 and December 2022, we treated 85 patients for soft tissue sarcomas. We considered patients whose specimens were approved for pilot research by the Human Biobank of St. Vincent's Hospital, The Catholic University of Korea, as potentially eligible. Based on this, 41% (35 patients) were eligible; 1% (one patient) was excluded because of gender mismatch between clinical and genetic data after controlling for data quality. Finally, 39 specimens (34 soft tissue sarcomas and five normal samples) were included from 34 patients who had clinical data. All tissue samples were collected intraoperatively. The five normal tissue samples were from muscle tissues. There were 20 female patients and 14 male patients, with a median age of 58 years (range 19 to 82 years). Genomic DNA was extracted from frozen tissue, and DNA methylation profiles were obtained. Genomic annotation of DNA methylation sites and hierarchical cluster analysis were performed to interpret results from DNA methylation profiling. A t-test was used to analyze different methylation probes. Benjamini-Hochberg-adjusted p value calculations were used to account for bias resulting from evaluating thousands of methylation sites. RESULTS: The most common histologic subtypes were liposarcoma (n = 10) and leiomyosarcoma (n = 9). The tumor grade was Fédération Nationale des Centres de Lutte Contre Le Cancer Grades 1, 2, and 3 in 3, 15, and 16 patients, respectively. DNA methylation profiling demonstrated differences between soft tissue sarcoma and normal tissue as 21,188 cytosine-phosphate-guanine sites. Despite the small number of samples, 72 of these sites showed an adjusted p value of < 0.000001, suggesting a low probability of statistical errors. Among the 72 sites, 70 exhibited a hypermethylation pattern in soft tissue sarcoma, with only two sites showing a hypomethylation pattern. Thirty of 34 soft tissue sarcomas were distinguished from normal samples using hierarchical cluster analysis. There was a different methylation pattern between leiomyosarcoma and liposarcoma at 7445 sites. Using the data, hierarchical clustering analysis showed that liposarcoma was distinguished from leiomyosarcoma. When we used the same approach and included other subtypes with three or more samples, only leiomyosarcoma and myxofibrosarcoma were separated from the other subtypes, while liposarcoma and alveolar soft-part sarcoma were mixed with the others. When comparing DNA methylation profiles between low-grade (Grade 1) and high-grade (Grades 2 and 3) soft tissue sarcomas, a difference in methylation pattern was observed at 144 cytosine-phosphate-guanine sites. Among these, 132 cytosine-phosphate-guanine sites exhibited hypermethylation in the high-grade group compared with the low-grade group. Hierarchical clustering analysis showed a division into two groups, with most high-grade sarcomas (28 of 31) separated from the low-grade group and few (3 out of 31) clustered together with the low-grade group. However, three high-grade soft tissue sarcomas were grouped with the Grade 1 cluster, and all of these sarcomas were Grade 2. When comparing Grades 1 and 2 to Grade 3, Grade 3 tumors were separated from Grades 1 and 2. CONCLUSION: We observed a different DNA methylation pattern between soft tissue sarcomas and normal tissues. Liposarcoma was distinguished from leiomyosarcoma using methylation profiling. High-grade soft tissue sarcoma samples showed a hypermethylation pattern compared with low-grade ones. Our findings indicate the need for research using methylation profiling to better understand the diverse biological characteristics of soft tissue sarcoma. Such research should include studies with sufficient samples and a variety of subtypes, as well as analyses of the expression and function of related genes. Additionally, efforts to link this research with clinical data related to treatment and prognosis are necessary. LEVEL OF EVIDENCE: Level III, diagnostic study.

Mesencephalic astrocyte-derived neurotrophic factor promotes axonal regeneration and the motor function recovery after sciatic nerve injury.

Peripheral nerve injuries have common clinical problems that are often accompanied by sensory and motor dysfunction and failure of axonal regeneration. Although various therapeutic approaches have been attempted, full functional recovery and axonal regeneration are rarely achieved in patients. In this study, we investigated the effects of recombinant adeno-associated virus (AAV) of mesencephalic astrocyte-derived neurotrophic factor (AAV-MANF) or placental growth factor (AAV-PlGF) transduced into mesenchymal stem cells (hMSC-MANF and hMSC-PlGF), which were then transplanted using human decellularized nerves (HDN) into sciatic nerve injury model. Our results showed that both AAV-MANF and AAV-PlGF were expressed in MSCs transplanted into the injury site. Behavioral measurements performed 2, 4, 6, 8, and 12 weeks after injury indicated that MANF facilitated the rapid and improved recovery of sensory and motor functions than PlGF. In addition, immunohistochemical analysis was used to quantitatively analyze the myelination of neurofilaments, Schwann cells, and regrowth axons. Both hMSC-MANF and hMSC-PlGF increased axon numbers and immunoreactive areas of axons and Schwann cells compared with the hMSC-GFP group. However, hMSC-MANF significantly improved the thickness of axons and Schwann cells compared with hMSC-PlGF. G-ratio analysis also showed a marked increase in axon myelination in axons thicker than 2.0 µm treated with MANF than that treated with PlGF. Our study suggests that transplantation of hMSC transduced with AAV-MANF has a potential to provide a novel and efficient strategy for promoting functional recovery and axonal regeneration in peripheral nerve injury.

Diagnosis of Marginal Infiltration in Soft Tissue Sarcoma by Radiomics Approach Using T2-Weighted Dixon Sequence.

BACKGROUND: Determination of preoperative soft tissue sarcoma (STS) margin is crucial for patient prognosis. PURPOSE: To evaluate diagnostic performance of radiomics model using T2-weighted Dixon sequence for infiltration degree of STS margin. STUDY TYPE: Retrospective. POPULATION: Seventy-two STS patients consisted of training (n = 58) and test (n = 14) sets. FIELD STRENGTH/SEQUENCE: A 3.0 T; T2-weighted Dixon images. ASSESSMENT: Pathologic result of marginal infiltration in STS (circumscribed margin; n = 27, group 1, focally infiltrative margin; n = 31, group 2-A, diffusely infiltrative margin; n = 14, group 2-B) was the reference standard. Radiomic volume and shape (VS) and other (T2) features were extracted from entire tumor volume and margin, respectively. Twelve radiomics models were generated using four combinations of classifier algorithms (R, SR, LR, LSR) and three different inputs (VS, T2, VS + T2 [VST2] features) to differentiate the three groups. Three radiologists (reader 1, 2, 3) analyzed the marginal infiltration with 6-scale confidence score. STATISTICAL TESTS: Area under the receiver operating characteristic curve (AUC) and concordance rate. RESULTS: Averaged AUCs of R, SR, LR, LSR models were 0.438, 0.466, 0.438, 0.466 using VS features, 0.596, 0.584, 0.814, 0.815 using T2 features, and 0.581, 0.587, 0.821, 0.821 using VST2 features, respectively. The LR and LSR models constructed with T2 or VST2 features showed higher AUC and concordance rate compared to radiologists' analysis (AUC; 0.730, 0.675, 0.706, concordance rate; 0.46, 0.43, 0.47 in reader 1, 2, 3). DATA CONCLUSION: Radiomics model constructed with features from tumor margin on T2-weighted Dixon sequence is a promising method for differentiating infiltration degree of STS margin. EVIDENCE LEVEL: 4 TECHNICAL EFFICACY: Stage 2.

Development and Validation of a Convolutional Neural Network Model to Predict a Pathologic Fracture in the Proximal Femur Using Abdomen and Pelvis CT Images of Patients With Advanced Cancer.

BACKGROUND: Improvement in survival in patients with advanced cancer is accompanied by an increased probability of bone metastasis and related pathologic fractures (especially in the proximal femur). The few systems proposed and used to diagnose impending fractures owing to metastasis and to ultimately prevent future fractures have practical limitations; thus, novel screening tools are essential. A CT scan of the abdomen and pelvis is a standard modality for staging and follow-up in patients with cancer, and radiologic assessments of the proximal femur are possible with CT-based digitally reconstructed radiographs. Deep-learning models, such as convolutional neural networks (CNNs), may be able to predict pathologic fractures from digitally reconstructed radiographs, but to our knowledge, they have not been tested for this application. QUESTIONS/PURPOSES: (1) How accurate is a CNN model for predicting a pathologic fracture in a proximal femur with metastasis using digitally reconstructed radiographs of the abdomen and pelvis CT images in patients with advanced cancer? (2) Do CNN models perform better than clinicians with varying backgrounds and experience levels in predicting a pathologic fracture on abdomen and pelvis CT images without any knowledge of the patients' histories, except for metastasis in the proximal femur? METHODS: A total of 392 patients received radiation treatment of the proximal femur at three hospitals from January 2011 to December 2021. The patients had 2945 CT scans of the abdomen and pelvis for systemic evaluation and follow-up in relation to their primary cancer. In 33% of the CT scans (974), it was impossible to identify whether a pathologic fracture developed within 3 months after each CT image was acquired, and these were excluded. Finally, 1971 cases with a mean age of 59 ± 12 years were included in this study. Pathologic fractures developed within 3 months after CT in 3% (60 of 1971) of cases. A total of 47% (936 of 1971) were women. Sixty cases had an established pathologic fracture within 3 months after each CT scan, and another group of 1911 cases had no established pathologic fracture within 3 months after CT scan. The mean age of the cases in the former and latter groups was 64 ± 11 years and 59 ± 12 years, respectively, and 32% (19 of 60) and 53% (1016 of 1911) of cases, respectively, were female. Digitally reconstructed radiographs were generated with perspective projections of three-dimensional CT volumes onto two-dimensional planes. Then, 1557 images from one hospital were used for a training set. To verify that the deep-learning models could consistently operate even in hospitals with a different medical environment, 414 images from other hospitals were used for external validation. The number of images in the groups with and without a pathologic fracture within 3 months after each CT scan increased from 1911 to 22,932 and from 60 to 720, respectively, using data augmentation methods that are known to be an effective way to boost the performance of deep-learning models. Three CNNs (VGG16, ResNet50, and DenseNet121) were fine-tuned using digitally reconstructed radiographs. For performance measures, the area under the receiver operating characteristic curve, accuracy, sensitivity, specificity, precision, and F1 score were determined. The area under the receiver operating characteristic curve was used to evaluate three CNN models mainly, and the optimal accuracy, sensitivity, and specificity were calculated using the Youden J statistic. Accuracy refers to the proportion of fractures in the groups with and without a pathologic fracture within 3 months after each CT scan that were accurately predicted by the CNN model. Sensitivity and specificity represent the proportion of accurately predicted fractures among those with and without a pathologic fracture within 3 months after each CT scan, respectively. Precision is a measure of how few false-positives the model produces. The F1 score is a harmonic mean of sensitivity and precision, which have a tradeoff relationship. Gradient-weighted class activation mapping images were created to check whether the CNN model correctly focused on potential pathologic fracture regions. The CNN model with the best performance was compared with the performance of clinicians. RESULTS: DenseNet121 showed the best performance in identifying pathologic fractures; the area under the receiver operating characteristic curve for DenseNet121 was larger than those for VGG16 (0.77 ± 0.07 [95% CI 0.75 to 0.79] versus 0.71 ± 0.08 [95% CI 0.69 to 0.73]; p = 0.001) and ResNet50 (0.77 ± 0.07 [95% CI 0.75 to 0.79] versus 0.72 ± 0.09 [95% CI 0.69 to 0.74]; p = 0.001). Specifically, DenseNet121 scored the highest in sensitivity (0.22 ± 0.07 [95% CI 0.20 to 0.24]), precision (0.72 ± 0.19 [95% CI 0.67 to 0.77]), and F1 score (0.34 ± 0.10 [95% CI 0.31 to 0.37]), and it focused accurately on the region with the expected pathologic fracture. Further, DenseNet121 was less likely than clinicians to mispredict cases in which there was no pathologic fracture than cases in which there was a fracture; the performance of DenseNet121 was better than clinician performance in terms of specificity (0.98 ± 0.01 [95% CI 0.98 to 0.99] versus 0.86 ± 0.09 [95% CI 0.81 to 0.91]; p = 0.01), precision (0.72 ± 0.19 [95% CI 0.67 to 0.77] versus 0.11 ± 0.10 [95% CI 0.05 to 0.17]; p = 0.0001), and F1 score (0.34 ± 0.10 [95% CI 0.31 to 0.37] versus 0.17 ± 0.15 [95% CI 0.08 to 0.26]; p = 0.0001). CONCLUSION: CNN models may be able to accurately predict impending pathologic fractures from digitally reconstructed radiographs of the abdomen and pelvis CT images that clinicians may not anticipate; this can assist medical, radiation, and orthopaedic oncologists clinically. To achieve better performance, ensemble-learning models using knowledge of the patients' histories should be developed and validated. The code for our model is publicly available online at https://github.com/taehoonko/CNN_path_fx_prediction . LEVEL OF EVIDENCE: Level III, diagnostic study.

Evaluation of anatomical and histological characteristics of human peripheral nerves: as an effort to develop an efficient allogeneic nerve graft.

Management of peripheral nerve defects is a complicated problem in clinical contexts. Autologous nerve grafting, a gold standard for surgical treatment, has been well known to have several limitations, such as donor site morbidity, a limited amount of available donor tissue, and size mismatches. Acellular nerve allografts (ANAs) have been developed as an alternative and have been applied clinically with favorable outcomes. However, because of the limited availability of commercialized ANAs due to supplier-related issues and high costs, efforts continue to produce alternative sources for ANAs. The present study evaluated the anatomical and histological characteristics of human peripheral nerves using 25 donated human cadavers. The length, diameter, and branching points of various peripheral nerves (median, ulnar, tibial, lateral femoral cutaneous, saphenous, and sural nerves) in both the upper and lower extremities were evaluated. The cross-sectional area (CSA), ratio of fascicular area, and numbers of fascicles were also evaluated via histologic analysis. CSA, the ratio of fascicular area, and the number of fascicles were analyzed statistically in correlation with demographic data (age, sex, height, weight, BMI). The mean length of all evaluated nerves ranged from 17.1 to 41.4 cm, and the mean diameter of all evaluated nerves ranged from 1.2 to 4.9 mm. Multiple regression analysis revealed correlations between the ratio of fascicular area and sex (p = 0.005) and BMI (p = 0.024) (R2 = 0.051). The results of the present study will be helpful in selecting necessary nerve allograft sources while considering the characteristics of each nerve in the upper and lower extremities during ANAs production.

Why locking plates for the proximal humerus do not fit well.

INTRODUCTION: We compared the angle of the humerus and plate and to assess compatibility of a plate to the proximal humerus using three-dimensional (3D) printed models. MATERIALS AND METHODS: A total of 120 cases were included, who underwent anteroposterior shoulder radiographs. From these, 30 cases with 3D shoulder computed tomography scans were randomly selected to print 3D model. The lateral angle between the lateral cortex of the humeral shaft and lateral border of the greater tuberosity (GT), neck-shaft angle, and height from the most proximal point of the GT to the angular point were measured. When the plates were applied on the 3D models, the gap from the most proximal point of the GT to the proximal rim of the plate was measured. RESULTS: The mean lateral angle in plain radiographs was 12.9 ± 2.2° and height from the most proximal point of the GT to the angular point was 44.4 ± 4.7 mm. The bending angles of the three plates were 8° and 10°. Height from the proximal rim of the plate to the bending point was 42.4, 42.0 and 43.8 mm. In 98% of cases, the lateral angle of the humerus was larger than all three plates. In 43% of cases, height of the GT was smaller than height of plates. When plates were applied to the 3D model, the mean gap from GT to plate was 4.8 ± 2.8 mm. CONCLUSIONS: There was large variation in the lateral angle of the proximal humerus, which was not correlated with the neck-shaft angle. The lateral angle of the humerus was larger than the plates and prone to varus reduction and medial collapse. LEVEL OF EVIDENCE OR CLINICAL RELEVANCE: Basic science study.

Differentiation of soft-tissue lymphoma from undifferentiated sarcoma: apparent diffusion coefficient histogram analysis.

BACKGROUND: Making the preoperative diagnosis of soft-tissue lymphoma is important because the treatments for lymphoma and sarcoma are different. PURPOSE: To determine the reliability and accuracy of single-slice and whole-tumor apparent diffusion coefficient (ADC) histogram analysis when differentiating soft-tissue lymphoma from undifferentiated sarcoma. MATERIAL AND METHODS: Patients with confirmed soft-tissue lymphoma or undifferentiated sarcoma who underwent 3-T magnetic resonance imaging (MRI), including diffusion-weighted imaging, were included. Single-slice and whole-tumor ADC histogram analyses were performed using software. Mean, standard deviation (SD), 5th and 95th percentiles, skewness, and kurtosis were compared between groups, and a receiver operating characteristic curve with area under the curve (AUC) was obtained. RESULTS: Thirteen patients with soft-tissue lymphoma and 12 patients with undifferentiated sarcoma were included. ADC histogram analysis of single-slice and whole-tumor, mean, SD, and 5th and 95th percentiles was significantly lower in lymphoma than in undifferentiated sarcoma. Whole-tumor analysis kurtosis was significantly higher in lymphoma than in undifferentiated sarcoma. All AUCs were high in single-slice and whole-tumor analysis: 0.987 vs. 1.000 in mean; 0.821 vs. 0.782 in SD; 0.949 vs. 0.949 in 5th percentile; and 1.000 vs. 1.000 in 95th percentile without significant difference. AUC of kurtosis in whole-tumor ADC histogram analysis was 0.750. CONCLUSION: Single-slice and whole-tumor ADC histogram analysis seems to be reliable and accurate for differentiating soft-tissue lymphoma from undifferentiated sarcoma.

Comparison of two different decellularization methods for processed nerve allograft.

The use of processed nerve allografts as an alternative to autologous nerve grafts, the gold standard treatment for peripheral nerve defects, is increasing. However, it is not widely used in Korea due to cost and insurance issues. Moreover, the main detergent used in the conventional Hudson method is unavailable. Therefore, a new nerve allograft decellularization process is needed. We aimed to compare the traditional Hudson method with a novel decellularization process that may remove cellular content more efficiently while preserving the extracellular matrix (ECM) structure using low concentration sodium dodecyl sulfate (SDS) and nuclease. After each decellularization process, DNA content was measured in nerve tissue. Masson's trichrome staining and scanning electron microscopy were performed to determine the state of preservation of the ECM. A significantly greater amount of DNA content was removed in the novel method, and the ECM structure was preserved in both methods. For the in vivo study, a 15-mm long sciatic nerve defect was created in two groups of Sprague-Dawley rats, and processed nerve allografts decellularized using the Hudson or novel method were transplanted. Functional and histological recovery results were measured 12 weeks post-transplantation. Ankle contracture angle, maximal isometric tetanic force of the tibialis anterior (TA), and the TA mass were compared between the groups, as well as the percent neural tissue (100 × neural area/intrafascicular area). There was no significant difference in functional and histological nerve recovery between the methods. The novel method is appropriate for developing a processed nerve allograft.

Germline Mutations for Novel Candidate Predisposition Genes in Sporadic Schwannomatosis.

BACKGROUND: Schwannomatosis is a late-onset tumor predisposition syndrome associated with the development of many different types of malignancies. A relevant genetic mechanism can be explained by three mutational events. The first-hit mutation is a germline mutation, and the SMARCB1 mutation on chromosome 22 is the most well-known genetic abnormality in patients with schwannomatosis. LZTR1 is another major predisposing gene in 22q-related schwannomatosis that lacks SMARCB1 variants. Although these two variants account for the occurrence of most familiar schwannomatoses, the genetic causes of sporadic schwannomatosis for the most part remain unknown. Therefore, current molecular diagnostic criteria cannot completely explain the basis of this disease. The common genetic background between schwannomatosis and other related malignant tumors is also unclear. Moreover, it is not easy to explain various clinical manifestations by only two known mutations. QUESTION/PURPOSES: (1) Are there important sequences outside the SMARCB1 or LZTR1 region on chromosome 22 that might carry a first-hit mutational predisposition to sporadic schwannomatosis? Or are there alternative evolutionarily conserved loci that might carry a first-hit mutational predisposition? (2) Is the age of disease onset associated to such genetic variants? METHODS: This study was a retrospective chart review and prospective genetic study on patients with schwannomatosis who were treated surgically. The clinical criteria to diagnose schwannomatosis were as follows: (1) histologically proven nonvestibular schwannomas; (2) no evidence of vestibular schwannomas on 3-mm brain MRI. A total of 21 patients were treated between March 2006 and June 2015. Since nine patients did not visit the outpatient clinic during the recruitment period, we obtained blood samples from 12 patients with schwannomatosis for a genetic analysis. After two patients were excluded because of their family history of schwannomatosis, genetic analyses were finally performed on 10 patients. Then, those with NF2, SMARCB1 or LZTR1 variants were screened by whole exome sequencing. All 10 patients passed our screening strategy. There were eight men and two women, with a median (range) age of 43 years (24 to 66) at the time of diagnosis. To select candidate genes, common ethnic variants and frequent mutations in in-house exome sequencing data were removed to exclude the population-specific polymorphisms not found in other population and to generalize the findings. Frameshift, nonsense, and splice-site variants were deemed pathogenic. Missense variants were classified as potentially pathogenic, variants of uncertain significance, or benign using in silico (via computer simulation) prediction algorithms, Sorting Intolerant From Tolerant (SIFT), Polymorphism Phenotyping v2 (PolyPhen-2), and Combined Annotation Dependent Depletion (CADD). A variant was considered potentially pathogenic if two or more algorithms predicted the variant to be damaging and benign if none considered it damaging. Then, potentially pathogenic variants only in the genes associated with cancer-predisposition or DNA damage repair were classified as the pathogenic candidate variants of sporadic schwannomatosis. The predictions for pathogenic candidate variants were checked again on Clinical Interpretation of Genetic Variants (InterVar) based on the American College of Medical Genetics guidelines and validated against Mendelian clinically applicable pathogenicity scores (M-CAP scores). RESULTS: We detected 26 variants; 13 variants across 10 genes were predicted to be pathogenic and found in seven patients, two each in ARID1A, PTCH2, and NOTCH2 and one each in MSH6, ALPK2, MGMT, NOTCH1, CIC, TSC2, and CDKN2A. One frameshift deletion in PTCH2 met the criteria for pathogenic or likely pathogenic classification, as recommended by the American College of Medical Genetics guidelines. Six missense mutations were classified as possibly pathogenic variants based on M-CAP scores. Four predicted pathogenic missense variants were detected in DNA damage repair (DDR) genes. Three DDR genes were affected: ARID1A, MGMT, and MSH6. Among the nine predicted pathogenic mutations detected in known cancer-predisposing genes, one was a frameshift deletion and the others were missense mutations. Seven tumor suppressor genes were involved: PTCH2, ALPK2, CIC, NOTCH1, NOTCH2, TSC2, and CDKN2A. One patient with multiple pathogenic variants in two DDR genes, ARID1A and MSH6, received a schwannomatosis diagnosis at 33 years old. Each of the other patients who had single variants in the DDR gene received their diagnoses at 41 years of age. The age at diagnosis was 40 years or older in patients with variants in cancer-predisposing genes, except for one patient who had multiple variants in TSC2 and CDKN2A. The carrier of those variants received the diagnosis at 24 years old. CONCLUSIONS: This study identified first-hit candidate mutations predisposing patients to schwannomatosis that were not related to SMARCB1 or LZTR1 variations in a cohort of patients with sporadic schwannomatosis. Patients with sporadic schwannomatosis without SMARCB1 or LZTR1 genetic variation may have developed the disease because of genomic variants related to cancer initiation in areas other than chromosome 22. Seven of 10 patients had predicted pathogenic germline mutations in DDR and cancer predisposition genes. We detected multiple cancer-related mutations in each patient. The age at the time schwannomatosis was diagnosed might be associated with a combination of variants and characteristics of the genes containing the variants; however, we did not have enough patients to confirm this association. CLINICAL RELEVANCE: The germline mutations identified in this study and the ideas related to the age of disease onset may provide potential candidate variants for future research on sporadic schwannomatosis and help to revise the current clinical and molecular diagnostic criteria. Further in vivo and in vitro studies are needed for these variants.

The anatomy of the saphenous and sural nerves as a source of processed nerve allografts.

As an alternative to autologous nerve donors, acellular nerve allografts (ANAs) have been studied in many experiments. There have been numerous studies on processing ANAs and various studies on the clinical applications of ANA, but there have not been many studies on sources of ANAs. The purposes of the present study were to evaluate the course of the saphenous and sural nerves in human cadavers and help harvest auto- or allografts for clinical implications. Eighteen lower extremities of 16 fresh cadavers were dissected. For the saphenous nerve and sural nerve, the distances between each branch and the diameters at the midpoint between each branch were measured. In the saphenous nerve, the mean length between each branch ranged from 7.2 to 28.6 cm, and the midpoint diameter ranged from 1.4 to 3.2 mm. In the sural nerve, the mean length between each branch ranged from 17.4 to 21 cm, and the midpoint diameter ranged from 2.3 to 2.8 mm. The present study demonstrates the length of the saphenous and sural nerve without branches with diameters larger than 1 mm. With regard for the clinical implications of allografts, the harvest of a selective nerve length with a large enough diameter could be possible based on the data presented in the present study.

Soft tissue sarcoma: adding diffusion-weighted imaging improves MR imaging evaluation of tumor margin infiltration.

PURPOSE: To determine the added value of diffusion-weighted imaging (DWI) to conventional magnetic resonance (MR) imaging in assessment of tumor margin infiltration in soft tissue sarcoma (STS) at 3T. MATERIALS AND METHODS: The institutional review board approved this retrospective study. Forty-five patients who underwent 3T MR imaging including DWI and were pathologically confirmed were included in this study. Two readers retrospectively scored conventional MR imaging alone. Then, they assessed a combination of conventional MR imaging and DWI. At pathology, margin infiltration was retrospectively reviewed by one pathologist blinded to MR findings. Areas under the curve (AUCs) of the receiver-operating characteristic curve were obtained for diagnostic performance. Interobserver agreement for the scoring of margin infiltration of STS was assessed with kappa statistics. RESULTS: Among 45 cases of STS, 33 had infiltrative tumor margin at pathology. Sensitivity, specificity, and accuracy of each reader were 100%, 17%, and 78%; 97%, 25%, and 78% on conventional MR imaging alone and 94%, 67%, and 87%; 94%, 42%, and 80% on conventional MR imaging combined with DWI. AUCs of conventional MR imaging combined with DWI were significantly higher than those of conventional MR imaging alone: 0.890 vs 0.678 (p = .0123) and 0.846 vs 0.640 (p = .0305) for each reader. Interobserver agreements of conventional MR imaging alone and conventional MR imaging combined with DWI were moderate to substantial (κ = 0.646, κ = 0.496). CONCLUSION: The addition of DWI to conventional MR imaging may improve specificity for assessing tumor margin infiltration in STS at 3T. KEY POINTS: • DWI has added value for assessment of tumor margin infiltration in soft tissue sarcoma. • Addition of DWI to conventional MRI at 3T may improve specificity. • Addition of DWI to conventional MRI may help orthopedic surgeon determine the extent of the resection margin.

Comparison of the regeneration induced by acellular nerve allografts processed with or without chondroitinase in a rat model.

There have been various studies about the acellular nerve allograft (ANA) as the alternative of autologous nerve graft in the treatment of peripheral nerve defects. As well as the decellularization process methods of ANA, the various enhancement methods of regeneration of the grafted ANA were investigated. The chondroitin sulfate proteoglycans (CSPGs) inhibit the action of laminin which is important for nerve regeneration in the extracellular matrix of nerve. Chondroitinase ABC (ChABC) has been reported that it enhances the nerve regeneration by degradation of CSPGs. The present study compared the regeneration of ANA between the processed without ChABC group and the processed with ChABC group in a rat sciatic nerve 15 mm gap model. At 12 weeks postoperatively, there was not a significant difference in the histomorphometric analysis. In the functional analysis, there were no significant differences in maximum isometric tetanic force, wet muscle weight of tibialis anterior. The processed without ChABC group had better result in ankle contracture angle significantly. In conclusion, there were no significant differences in the regeneration of ANA between the processed without ChABC group and the processed with ChABC group.

Differentiation of focal indeterminate marrow abnormalities with multiparametric MRI.

PURPOSE: To explore magnetic resonance imaging (MRI) parameters from intravoxel incoherent motion diffusion-weighted imaging (IVIM-DWI), multiecho Dixon imaging (ME-Dixon), and dynamic contrast-enhanced imaging (DCE) for differentiating focal indeterminate marrow abnormalities MATERIALS AND METHODS: Forty-two patients with 14 benign and 28 malignant focal marrow abnormalities were included. The following were independently analyzed by two readers: signal intensity (SI), contour, and margin on conventional MR images; SI on b-800 images (SIb-800 ), apparent diffusion coefficient (ADC), IVIM parameters (Dslow, Dfast , and f), fat fraction (Ff), and DCE parameters (time-to-signal intensity curve pattern, iAUC, Ktrans , kep , and ve ). The MR characteristics and parameters from benign and malignant lesions were compared with a chi-squared test and the Mann-Whitney U-test, respectively. The area under receiver operating characteristic (ROC) curves (AUC) of each sequence were also compared. Interobserver agreements were assessed with Cohen's κ, and intraclass correlation coefficient (ICC). RESULTS: ADC, Dslow , and Ff demonstrated a significant difference between benign and malignant marrow abnormalities for both readers (P < 0.001). SIb-800 and perfusion-related parameters from IVIM-DWI and DCE were not significantly different between the two groups (P = 0.145, 0.439, and 0.337 for reader 1, P = 0.378, 0.368, and 0.343 for reader 2, respectively). The AUCs of ADC, Dslow , and Ff were significantly higher for differentiating indeterminate marrow abnormalities in both readers (P < 0.001). Interobserver agreements were substantial in SIb-800 , and ICCs were almost perfect for ADC, Dslow , f, and Ff, and substantial for iAUC, kep , Ktrans , ve , and Dfast . CONCLUSION: ADC, Dslow , and Ff may provide information for differentiating focal indeterminate abnormalities. LEVEL OF EVIDENCE: 3 Technical Efficacy: Stage 2 J. MAGN. RESON. IMAGING 2017;46:49-60.

The Effect of Autologous Platelet-Rich Plasma on Bone Regeneration by Autologous Mesenchymal Stem Cells Loaded onto Allogeneic Cancellous Bone Granules.

To develop a clinically effective bone regeneration strategy, we compared bone regeneration using allogeneic cancellous bone granule scaffolds loaded with autologous bone marrow-derived mesenchymal stem cells (BM-MSC) with or without autologous platelet-rich plasma (PRP). Critical-sized segmental bone defects were made at the mid-shaft of both radiuses in 41 New Zealand White rabbits. Small-sized allogeneic cancellous bone granules (300-700 µm in diameter) loaded with BM-MSC were implanted on one side, and PRP was added. On the other side, autologous BM-MSC loaded onto allogeneic cancellous granules were grafted as a control. Bone regeneration was assessed by radiographic evaluation at 4, 8, and 16 weeks postimplantation and by micro-computed tomography (micro-CT) and histological evaluation of the retrieved specimens at 8 and 16 weeks. The experimental group did not show significantly higher bone quantity indices than the control group at any time point. Micro-CT analysis revealed that both groups had similar mean total volumes, surface areas, and other parameters at 8 and 16 weeks. Histological evaluation of 8- and 16-week specimens also showed a similar progression of new bone formation and maturation. In this experiment using a contralateral control group in the same individual, an initial single addition of PRP in allogeneic cancellous bone granules loaded with BM-MSC for critical-sized bone defects in the weight-bearing area did not induce a consequent difference in bone healing. Further research into the optimal preparation and application of PRP is necessary. Furthermore, studies involving a greater number of subjects and larger experimental animals could determine the clinical relevance of PRP treatment.

Differentiation of malignant from benign soft tissue tumours: use of additive qualitative and quantitative diffusion-weighted MR imaging to standard MR imaging at 3.0 T.

OBJECTIVES: To determine the added value of diffusion-weighted imaging (DWI) to standard magnetic resonance imaging (MRI) to differentiate malignant from benign soft tissue tumours at 3.0 T. METHODS: 3.0 T MR images including DWI in 63 patients who underwent surgery for soft tissue tumours were retrospectively analyzed. Two readers independently interpreted MRI for the presence of malignancy in two steps: standard MRI alone, standard MRI and DWI with qualitative and quantitative analysis combined. RESULTS: There were 34 malignant and 29 non-malignant soft tissue tumours. In qualitative analysis, hyperintensity relative to skeletal muscle was more frequent in malignant than benign tumours on DWI (P=0.003). In quantitative analysis, ADCs of malignant tumours were significantly lower than those of non-malignant tumours (P≤0.002): 759±385 vs. 1188±423 µm(2)/sec minimum ADC value, 941±440 vs. 1310±440 µm(2)/sec average ADC value. The mean sensitivity, specificity and accuracy of both readers were 96%, 72%, and 85% on standard MRI alone and 97%, 90%, and 94% on standard MRI with DWI. CONCLUSIONS: The addition of DWI to standard MRI improves the diagnostic accuracy for differentiation of malignant from benign soft tissue tumours at 3.0 T. KEY POINTS: DWI has added value for differentiating malignant from benign soft tissue tumours. Addition of DWI to standard MRI at 3.0 T improves the diagnostic accuracy. Measurements of both ADC min within solid portion and ADC av are helpful.

Osteosarcoma in Asian Populations Over the Age of 40 Years: A Multicenter Study.

BACKGROUND: Elderly patients with osteosarcoma (OSA) are no longer uncommon; however, many questions remain regarding this population. We investigated the clinicopathological characteristics and prognostic factors of OSA in an Asian population over the age of 40 years. METHODS: This was a multi-national, multi-institutional study by the Eastern Asian Musculoskeletal Oncology Group (EAMOG). RESULTS: A total of 232 patients were enrolled (116 males and 116 females), with a median age of 50 years at diagnosis; 25 (10.8 %) patients exhibited initial metastasis. Median follow-up was 52 months for survivors. We observed 102 osteolytic and mixed radiographic findings for 173 lesions. Histological subtypes other than osteoblastic type were frequent. Radiation-associated OSA was seen in seven patients, with a 5-year overall survival (OS) of 16.7 %. No Paget's OSA was observed. High-grade spinopelvic OSA was seen in 29 (12.5 %) patients. The 5-year OS was 59.4 % in patients without initial metastasis and 45.2 % in patients with spinopelvic OSA. While surgery and initial metastasis were common prognostic factors for OS, chemotherapy was not. Histologic response to neoadjuvant chemotherapy was poor in 61 of 83 patients. CONCLUSION: This study revealed distinct clinicopathological features of OSA patients over 40 years of age compared with younger patients, such as the high incidence of axial tumors, common osteolytic and mixed radiographic findings, the high frequency of unusual histologic subtypes, and poor prognosis. Contrary to Western elderly patients with OSA, there was no Paget's OSA in this study, which may result in a lower incidence of secondary OSA. Prognostic factor analyses demonstrated chemotherapy did not influence OS.

Pseudomyxoma peritonei extending to the lower extremity: a case report.

Pseudomyxoma peritonei is characterized by mucinous ascites originating from a mucin-producing neoplasm; however, even the definition is still under debate. Tumor deposits extend and ultimately engulf the entire cavity, causing death from cachexia due to limited intestinal movement. Here, we report a unique case of an 80-year-old woman with pseudomyxoma peritonei, which extended to the lower extremity mimicking infectious condition. The patient survived for a long time without bowel obstruction despite having the histologic subtype that has an unfavorable prognosis. The extremity lesion was treated with limited extensive surgery. The origin of the disease and the mechanism of extension to the extremity could not be clarified. Clinicians should be aware of the original disease entity and this unusual presentation and determine its mechanism and the best management strategy.

Giant cell tumor of soft tissue: a case report with emphasis on MR imaging.

Giant cell tumor of soft tissue is a rare neoplasm, histologically resembling giant cell tumor of bone. In this report, we describe a deep and solid giant cell tumor of soft tissue interpreted as a benign soft tissue tumor based on magnetic resonance (MR) findings with hypointense to intermediate signals on T2-weighted images and impeded diffusivity (water movement) on diffusion-weighted imaging (DWI), which could suggest a giant-cell-containing benign soft tissue tumor, despite the malignancy suggested by 18F-fluorodeoxyglucose positron emission tomography-computed tomography in a 35-year-old male. To our knowledge, this report introduces the first deep, solid giant cell tumor of soft tissue with MR features of a giant-cell-containing benign soft tissue tumor, despite the malignancy-mimicking findings on 18F-FDG PET-CT.

Bone regeneration potential of allogeneic or autogeneic mesenchymal stem cells loaded onto cancellous bone granules in a rabbit radial defect model.

For developing a clinically effective bone regeneration strategy, we compare the bone regeneration potential of cultured allogeneic bone marrow-derived mesenchymal stem cells (BM-MSCs) and of autologous BM-MSCs loaded onto allogeneic cancellous bone granule scaffolds. A critical-sized segmental bone defect was made at the mid-shaft of both radiuses in 19 New Zealand White rabbits (NWRs). In the experimental group, allogeneic BM-MSCs loaded onto small-sized allogeneic cancellous bone granules (300~700 um in diameter) were implanted in one side of a bone defect. In the control group, autologous BM-MSCs loaded onto allogeneic cancellous granules were grafted in the other side. Bone regeneration was assessed by radiographic evaluation at 4, 8, 12 and 16 weeks post-implantation and by micro-computed tomography (micro-CT) and histological evaluation at 8 and 16 weeks. The experimental groups showed lower bone quantity indices (BQIs) than the control groups at 12 and 16 weeks (p < 0.05), although no significant difference was observed at 4 and 8 weeks (p > 0.05). Micro-CT analysis revealed that both groups had similar mean total bone volume and other parameters including trabecular thickness, number and separation at either 8 or 16 weeks. Only bone surface area revealed less area in the experimental group at 16 weeks. Histological evaluation of 8-week and 16-week specimens showed similar biologic processes of new bone formation and maturation. There was no inflammatory reaction indicating an adverse immune response in both allogeneic and autologous MSC groups. In conclusion, allogeneic BM-MSCs loaded onto allogeneic cancellous bone granules had comparable bone regeneration potential to autologous BM-MSCs in a rabbit radial defect model.

Does size difference in allogeneic cancellous bone granules loaded with differentiated autologous cultured osteoblasts affect osteogenic potential?

We study the efficacy of bone regeneration by using two differently sized allogeneic cancellous bone granules loaded with autologous cultured osteoblasts in a rabbit model. Critical-sized bone defects of the radial shaft were made in 40 New Zealand White rabbits. Small allogeneic bone granules (150-300 µm in diameter) loaded with cultured differentiated autologous osteoblasts were implanted into one forearm (SBG group) and large bone granules (500-710 µm) loaded with osteoblasts were implanted into the forearm of the other side (LBG group). Radiographic evaluations were performed at 3, 6, 9 and 12 weeks and histology and micro-CT image analysis were carried out at 6 and 12 weeks post-implantation. On radiographic evaluation, the LBG group showed a higher bone quantity index at 3 and 6 weeks post-implantation (P < 0.05) but statistical significance was lost at 9 and 12 weeks. The progression of biological processes of the SBG group was faster than that of the LBG group. On micro-CT image analysis, the LBG group revealed a higher total bone volume and surface area than the SBG group at 6 weeks (P < 0.05) but the difference decreased at 12 weeks and was without statistical significance. Histological evaluation also revealed faster progression of new bone formation and maturation in the SBG group. Thus, the two differently sized allogeneic bone granules loaded with co-cultured autologous osteoblasts show no differences in the amount of bone regeneration, although the SBG group exhibits faster progression of bone regeneration and remodeling. This method might therefore provide benefits, such as a short healing time and easy application in an injectable form, in a clinical setting.