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Summary Brown tumors are osteoclastic, benign lesions characterized by fibrotic stroma, intense vascularization and multinucleated giant cells. They are the terminal expression of the bone remodelling process occurring in advanced hyperparathyroidism. Nowadays, due to earlier diagnosis, primary hyperparathyroidism keeps few of the classical manifestations and brown tumors are definitely unexpected. Thus, it may happen that they are misdiagnosed as primary or metastatic bone cancer. Besides bone imaging, endocrine evaluation including measurement of serum parathyroid hormone and calcium (Ca) levels supports the pathologist to address the diagnosis. Herein, a case of multiple large brown tumors misdiagnosed as a non-treatable osteosarcoma is described, with special regards to diagnostic work-up. After selective parathyroidectomy, treatment with denosumab was initiated and a regular follow-up was established. The central role of multidisciplinary approach involving pathologist, endocrinologist and oncologist in the diagnostic and therapeutic work-up is reported. In our opinion, the discussion of this case would be functional especially for clinicians and pathologists not used to the differential diagnosis in uncommon bone disorders. Learning points: Brown tumors develop during the remodelling process of bone in advanced and long-lasting primary or secondary hyperparathyroidism. Although rare, they should be considered during the challenging diagnostic work-up of giant cell lesions. Coexistence of high parathyroid hormone levels and hypercalcemia in primary hyperparathyroidism is crucial for the diagnosis. A detailed imaging study includes bone X-ray, bone scintiscan and total body CT; to rule out bone malignancy, evaluation of bone lesion biopsy should include immunostaining for neoplastic markers as H3G34W and Ki67 index. If primary hyperparathyroidism is confirmed, selective parathyroidectomy is the first-line treatment. In advanced bone disease, treatment with denosumab should be considered, ensuring a strict control of Ca levels.

BACKGROUND Phosphaturic mesenchymal tumors are rare neoplasms, frequently presenting with osteomalacia. These neoplasms usually grow at a slow rate and are associated with unspecific symptoms. CASE In this study, we present the case of a 70-year-old woman who had been suffering from musculoskeletal pain, hypophosphatemia, and spontaneous fractures. Positron emission tomography with Gallium showed increase uptake in a subpleural lesion. CONCLUSION The patient underwent surgical excision of the subpleural lesion with a non-intubated uniportal video-assisted thoracoscopic surgery approach.

To diagnose a bone tumor, an analytical approach is required, as well as the use of clinical, histological, and radiological means.

Although remarkable progress has been made in the histological typing and the molecular genotyping of bone tumors, their classification remains extremely complex; in some instances, the integration of histomorphological, phenotypic, and molecular features with the clinical appearance does not provide sufficient prognostic information to assess the clinical course and plan treatment.

Conventional central chondrosarcoma (CCC) is a malignant bone tumor that is characterized by the production of chondroid tissue. Since radiation therapy and chemotherapy have limited effects on CCC, treatment of most patients depends on surgical resection. This study aimed to identify the expression profiles of microRNAs (miRNAs) and isomiRs in CCC tissues to highlight their possible participation to the regulation of pathways critical for the formation and growth of this type of tumor. Our study analyzed miRNAs and isomiRs from Grade I (GI), Grade II (GII), and Grade III (GIII) histologically validated CCC tissue samples. While the different histological grades shared a similar expression profile for the top abundant miRNAs, we found several microRNAs and isomiRs showing a strong different modulation in GII + GIII vs GI grade samples and their involvement in tumor biology could be consistently hypothesized. We then in silico validated these differently expressed miRNAs in a larger chondrosarcoma public dataset and confirmed the expression trend for 17 out of 34 miRNAs. Our results clearly suggests that the contribution of miRNA deregulation, and their targeted pathways, to the progression of CCC could be relevant and strongly indicates that when studying miRNA deregulation in tumors, not only the canonical miRNAs, but the whole set of corresponding isomiRs should be taken in account. Improving understanding of the precise roles of miRNAs and isomiRs over the course of central chondrosarcoma progression could help identifying possible targets for precision medicine therapeutic intervention.

Selection of cancer patients for treatment with immune checkpoint inhibitors remains a challenge due to tumour heterogeneity and variable biomarker detection. PD-L1 expression in 24 surgical chordoma specimen was determined immunohistochemically with antibodies 28-8 and E1L3N. The ability of patient-derived organoids to detect treatment effects of nivolumab was explored by quantitative and qualitative immunofluorescence and FACS analysis. The more sensitive antibody, E1L3N (ROC = 0.896, p = 0.001), was associated with greater tumour diameters (p = 0.014) and detected both tumour cells and infiltrating lymphocytes in 54% of patients, but only 1–15% of their cells. Organoids generated from PD-L1-positive patients contained both tumour cells and PD-1/CD8-positive lymphocytes and responded to nivolumab treatment with marked dose-dependent diameter reductions of up to 50% and increased cell death in both PD-L1-positive and negative organoids. Patient-derived organoids may be valuable to predict individual responses to immunotherapy even in patients with low or no immunohistochemical PD-L1 expression.

Autologous fat tissue grafting (AFTG) has been successfully used in the treatment of different sclerotic conditions, including localized scleroderma. Patients with advanced systemic sclerosis (SSc)-related perioral thickening and mouth opening limitation are candidates for this therapeutic approach. AFTG of the lips was performed to improve mouth opening in patients with SSc. We enrolled in the study 20 female patients with diffuse SSc (median age 35 ± 15 years and 11 ± 10 years of disease duration). Two-milliliter fractions of autologous fat drawn from trochanteric or periumbilical areas were injected in eight different sites around the mouth. Baseline and after-treatment mouth opening changes were assessed by measuring interincisal distance and oral perimeter, while skin hardness was tested by digital durometer. Pre- and posttreatment modifications of microvascular architecture were assessed by counting capillaries in the inferior lip videocapillaroscopy (VC) images and by scoring the microvascular density (MVD) in anti-CD34/CD31 immunohistochemical (IH) stained perioral skin biopsy sections. Similarly, histological sections were examined to evaluate dermoepidermic junction (DEJ) modifications. Three months after treatment, both the interincisal distance and oral perimeter significantly increased ( p < 0.001). At the same time, a significant skin neovascularization became evident, both considering the VC images ( p< 0.001) and MVD scores in IH sections ( p< 0.0001). Finally, some skin histological aspects also improved, as shown by the significant changes in DEJ flattening scores ( p < 0.0001). The present study suggests that, in patients with SSc, AFTG can improve mouth opening and function, induce a neovascularization, and partially restore the skin structure.

Background: Soft tissue giant cell tumor (GCT-ST) of low malignant potential is an uncommon neoplasm, considered the soft tissue counterpart of giant cell tumor of bone. GCT-ST mainly affects young to middle-age adults and presents as a painless growing mass mainly located in the lower extremities and trunk. Histologically, this tumor is characterized by a mixture of uniformly scattered osteoclast-like multinucleated giant cells intimately admixed with short fascicles of spindled cells. Complete excision with negative surgical margins is associated with a benign clinical course in most cases. Methods: The authors report the clinicopathological and immunohistochemical features of an unusual GCT-ST of 46 years duration previously histologically misdiagnosed as Kaposi's sarcoma. Results: Histologically, the tumor was characterized by a multinodular growth pattern with osteoclast-like multinucleated giant cells admixed with spindle cells partially arranged in a storiform pattern, fibrosis and foci of haemorrhage and mature bone. Immunohistochemistry revealed CD68 reactivity of the multinucleated giant cells. Conclusion: GCT-ST is a rare neoplasm characterized by benign clinical course if excised adequately, as shown by our case of exceptionally long duration. Emphasis is placed on the importance of differential diagnosis with other giant cell-rich soft tissue neoplasms because clinical behaviour, prognosis and treatment significantly differ.

Background: Achondroplasia (ACH) represents the major cause of dwarfism and is due to mutations in the fibroblast growth factor receptor 3 (FGFR3) gene. The cellular mechanisms involved in the reduced growth have been mainly described for in vitro or in vivo models, but few data have been obtained for humans. Methods: Thirteen children with ACH were enrolled in the study; the presence of FGFR3 mutations was determined by restriction fragment length polymorphism analysis and sequencing, whereas protein expression in cartilage biopsy was assessed by immunohistochemistry. Results: Chondrocytes in cartilage biopsies of ACH children were characterized by the presence of growth arrest mediated by STAT activation (both STAT1 and STAT5) and increased expression of p21 and cyclin D1, whereas no expression of either p53 or cyclin D3 could be detected. This mechanism was present in ACH children carrying the G380R mutation but also in a patient in whom no mutation could be detected in the entire coding region of the FGFR3 gene. Conclusions: These data thus demonstrate the presence of a common final mechanism involving p21 and possibly leading to a block in chondrocyte proliferation. © 2009 The Japanese Orthopaedic Association.

Iron deficiency is a manifestation of celiac disease (CD) usually attributed to a decreased absorptive surface, although no data on the regulation of iron transport under these conditions are currently available. Our aim was to evaluate divalent metal transporter 1 (DMT1), duodenal cytochrome b (Dcytb), ferroportin 1 (FP1), hephaestin, and transferrin receptor 1 (TfR1) expression, as well as iron regulatory protein (IRP) activity in duodenal biopsies from control, anemic, and CD patients. We studied 10 subjects with dyspepsia, 6 with iron-deficiency anemia, and 25 with CD. mRNA levels were determined by real-time PCR, protein expression by Western blotting or immunohistochemistry, and IRP activity by gel shift assay. Our results showed that DMT1, FP1, hephaestin, and TfR1 mRNA levels were significantly increased in CD patients with reduced body iron stores compared with controls, similar to what was observed in anemic patients. Protein expression paralleled the mRNAs changes. DMT1 protein expression was localized in differentiated enterocytes at the villi tips in controls, whereas with iron deficiency it was observed throughout the villi. FP1 expression was localized on the basolateral membrane of enterocytes and increased with low iron stores. TfR1 was localized in the crypts in controls but also in the villi with iron deficiency. These changes were paralleled by IRP activity, which increased in all iron-deficient subjects. We conclude that duodenal DMT1, FP1, hephaestin, and TfR1 expression and IRP activity, thus the iron absorption capacity, are upregulated in CD patients as a consequence of iron deficiency, whereas the increased enterocyte proliferation observed in CD has no effect on iron uptake regulation.

We examined 28 cases of primary bone lymphomas (PBL; stage IE) and 26 cases of systemic lymphomas involving the bone (SBL; stage IIE to IV). Two histologic types were prevalent: Diffuse large B-cell lymphomas (DLBCL; 26 PBL and 21 SBL) and CD30+ anaplastic large cell lymphomas (ALCL; 1 PBL and 4 SBL). A mature B phenotype (CD45+, CD20+, CD79a+, CDw75+/-, CD10-/+) was established in the DLBCL group. Bcl-2 immunoreactivity was demonstrated in 13/37 cases (35%), and bcl-6 immunostaining was observed in 22/32 cases (69%). ALCL showed null/T phenotype (CD3-/+; CD43+/-; CD30+), with ALKI-1 expression in 3/3 cases. With use of a FR3A primer, a monoclonal pattern was demonstrated by PCR analysis in 22/41 lymphomas (54%). Bcl-2 translocation was identified in 2/41 cases (5%). This study details the clinical and pathological characteristics of bone lymphomas. Our immunohistochemical and molecular data suggest that most of them are “de novo” DLBCL and support their follicle center origin.

Background & Aims: Stimulator of Fe Transport (SFT) and transferrin receptor (TfR) are proteins involved in iron transport. This study evaluated iron metabolism protein expression in duodenal biopsy specimens from controls and patients with abnormal iron metabolism. Methods: Twelve controls, 8 patients with iron deficiency anemia, 7 with HFE-related hemochromatosis, and 6 with non–HFE-related iron overload were studied. Immunohistochemistry was performed on duodenal biopsy specimens with anti-TfR and anti-SFT antibodies which recognize a putative stimulator of Fe transport of ~80 kilodaltons. Results: In controls, the putative stimulator of Fe transport was expressed in the middle and distal part of the villi in the subapical cytoplasmatic region. Its expression increased in anemics and, to a lesser degree, in HFE-related hemochromatotics, whereas it was reduced in patients with non–HFE-related iron overload. TfR expression showed a crypt-to-tip gradient in controls, but not in anemics, in whom it was uniformly overexpressed. TfR expression was intermediate in HFE-related hemochromatotics and similar to controls in non–HFE-related iron overload. Conclusions: Expression of the putative stimulator of Fe transport and TfR increases in iron deficiency. Increased expression of both proteins is present only in HFE-related hemochromatotics suggesting that other factors may be involved in determining non–HFE-related iron overload phenotype. GASTROENTEROLOGY 2001;120:1404-1411

The molecular mechanism responsible for Ewing's Sarcoma (ES) remains largely unknown. MicroRNAs (miRNAs), a class of small non-coding RNAs able to regulate gene expression, are deregulated in tumors and may serve as a tool for diagnosis and prediction. However, the status of miRNAs in ES has not yet been thoroughly investigated. This study compared global miRNAs expression in paraffin-embedded tumor tissue samples from 20 ES patients, affected by primary untreated tumors, with miRNAs expressed in normal human mesenchymal stromal cells (MSCs) by microarray analysis. A miRTarBase database was used to identify the predicted target genes for differentially expressed miRNAs. The miRNAs microarray analysis revealed distinct patterns of miRNAs expression between ES samples and normal MSCs. 58 of the 954 analyzed miRNAs were significantly differentially expressed in ES samples compared to MSCs. Moreover, the qRT-PCR analysis carried out on three selected miRNAs showed that miR-181b, miR-1915 and miR-1275 were significantly aberrantly regulated, confirming the microarray results. Bio-database analysis identified BCL-2 as a bona fide target gene of the miR-21, miR-181a, miR-181b, miR-29a, miR-29b, miR-497, miR-195, miR-let-7a, miR-34a and miR-1915. Using paraffin-embedded tissues from ES patients, this study has identified several potential target miRNAs and one gene that might be considered a novel critical biomarker for ES pathogenesis.

Permanent synovial sarcoma cell lines are invaluable tools for understanding of the biology of this tumor. The present study reports the establishment of a new human cell line, PDSS-26, derived from a surgical specimen of a poorly differentiated synovial sarcoma. PDSS-26 has a doubling time of a 72 hours and grows as a monolayer of spindle cells that retain immunoreactivity for bcl-2 and vimentin. Karyotypic analysis revealed a rearrangement involving chromosomes 17 and 18, at the breakpoints q11.2 and q11.2, respectively, as the only structural aberrations. Analysis by reverse transcriptase polymerase chain reaction showed the presence of the SYT-SSX1 fusion transcript in both the primary tumor and the cell line. Cytoplasmic PTEN staining was detected by immunohistochemistry in both the PDSS-26 cell line and in original tumor, whereas no mutation was identified by automatic sequencing. Thus, PDSS-26 cells could be useful for future functional studies. (C) 2003 Elsevier Inc. All rights reserved.

Somatic point mutations of the 5' noncoding region of the BCL-6 gene have been described as genetic alterations in non-Hodgkin lymphomas (NHL). They are more frequent in diffuse large B cell (DLBCL) and follicular lymphomas (FL). This study aims to analyse the presence and distribution of BCL-6 gene mutations in a large series of primary bone lymphomas (PBL), a rare extranodal presentation of NHL frequently associated with diffuse large cell morphology. Fifty-three cases of PBL were examined. Mutations were detected with non-radioisotopic polymerase chain reaction-single strand conformation polymorphism and visualized with fluorescent cycle sequencing. Among stage I(E) PBL, there were 30 cases of DLBCL and one each of follicular, anaplastic large cell and peripheral T-cell lymphoma. The stage II(E) PBL included six DLBCL and one lymphoplasmacytic lymphoma, whereas within stage IV PBL there were 12 DLBCL and one Burkitt lymphoma. Fifteen patients (28%) displayed mutational events. In nine cases there were more than one BCL-6 mutation. Only DLBCL displayed mutations (31%). Mutations included single base-pair substitutions (16 transitions and ten transversions) and a single point insertion (ins A 427-28). The frequency of mutations resulted lower in DLBCL of the PBL category than in the majority of other extranodal large cell lymphomas. The prevalence of mutations was higher in stage I(E) PBL than in more advanced stages of the disease (II(E) + IV) ( p=0.02). Our results reinforce the observation of heterogeneity of the DLBCL included in the clinical category of PBL.

Achondroplasia, the most common skeletal dysplasia, is an autosomaldominant disorder caused by a gain-of-function mutation in the gene encoding the type 3 receptor for fibroblast growth factor (FGFR3); in more than 95% of cases, the mutation is G380R. The disease is characterized by a disturbance of endochondral bone formation and affects the cartilaginous growth plate in the growing skeleton, leading to a variety of manifestations and complications.

Background Autologous fat grafts (AFG) for restoring soft tissue defects have been used for many decades and has many clinical applications. Recent studies have shown the efficacy of the AFG in the treatment of localized scleroderma. Besides mechanical lipofilling, AFG seems to produce trophic effects on both vascular and stromal cells. However, the exact mechanisms underlying the clinical results are far to be completely understood. Objectives A prospective pilot study has been carried out aimed at evaluating whether the perioral AFG in SSc patients may improve the mouth functional impairment related to the sclerotic changes. In addition, we tried to clarify whether the perioral fat filling might induce local trophic changes in the stromal and vascular structures. Methods Twenty-five patients with diffuse SSc (dcSSc) (mean age + SD 36.5+16 yrs, disease duration of 12±9 yrs) and perioral skin fibrotic changes with limited mouth opening were treated by topical perioral administration of AFG. Lipofilling was performed according to the Coleman tecnique. Follow-up was performed at 1 and 3 months postoperatively by evaluating measure of inter-incisal distance and mouth perimeter; skin hardness was measured by a digital durometer. A baseline and 3 months post-treatment evaluation of microcirculatory abnormalities was made using labial video capillaroscopy and skin biopsy. Results: Results. All patients made good postoperative recovery without complications showing an immediate subjective improvement of their skin stiffness. One and 3 months after lipofilling, the median measure of inter-incisal distance was significantly increased in comparison with the score before treatment (p=0.02); the same results, with overtime significant improvement from baseline to 1 and 3 months, were confirmed by measuring mouth perimeter (p=0.004 and 0.003). Durometer evaluations showed that skin hardness resulted significantly lower after treatment (p=0.014 and 0.019). Labial capillaroscopy of treated patients showed significant improvement in microvascular patterns after AFG in term of increase in capillary density (p=0.001) with more regular organization of microcirculation, increased length and reduced diameter shapes. Pathological studies confirmed the improvement in capillary density in the areas of treated skin with a significant upregulation of CD31 and CD34 expression (p Conclusions Our study suggest that AFG of SSc fibrotic perioral changes can provide immediate and long-lasting benefits in terms of both aesthetic and functional improvement. Itis a safe, relatively non-invasive, and rapid procedure. In addition we provide evidences that AFG has the potential to favour neovascularization in SSc and revert SSc fibrotic features. The overtime increase of treatment benefits suggests that other mechanisms different from the simple fat filling may be involved. The cellular and/or tissue mechanisms underlying these changes need further investigation. Disclosure of Interest None Declared