Systematic Characterization of the Clinical and Pathological Features of Schwannomas Harboring SH3PXD2A::HTRA1 Fusion.

The understanding of schwannoma tumorigenesis has been reshaped by the recent identification of SH3PXD2A::HTRA1 fusion in 10% of intracranial/spinal schwannomas. Nonetheless, pathologic features of schwannomas harboring this fusion, as well as its prevalence outside intracranial/spinal locations, have not been characterized. We screened 215 consecutive schwannomas for their clinicopathologic characteristics and fusion status using reverse-transcriptase polymerase chain reaction (RT-PCR). Among 29 (13.5%) fusion-positive schwannomas, the most prevalent location was peripheral somatic tissue (30.7%, 19/62), followed by spinal/paraspinal (18.4%, 7/38), body cavity/deep structures (10%, 2/20), intracranial (1.3%, 1/75), and viscera (0/13). All 8 cellular, 4 microcystic/reticular, and 3 epithelioid schwannomas were fusion-negative, as were 41/42 nonschwannomatous peripheral nerve sheath tumors. Remarkably, a distinct 'serpentine' palisading pattern, comprising ovoid/plump cells shorter than usual schwannian cells in a hyalinized stroma, was identified in most fusion-positive cases and the schwannomatous component of the only fusion-positive malignant peripheral nerve sheath tumor. To validate this finding, 60 additional cases were collected, including 36 with (≥10% arbitrarily) and 24 without appreciable serpentine histology, of which 29 (80.6%) and 2 (8.3%) harbored the fusion, respectively. With percentages of 'serpentine' areas scored, 10% was determined as the optimal practical cut-off to predict the fusion status (sensitivity, 0.950; specificity, 0.943). Fusion positivity was significantly associated with serpentine histology, smaller tumors, younger patients, and peripheral somatic tissue, while multivariate logistic linear regression analysis only identified serpentine histology and location as independent fusion-predicting factors. RNA in situ hybridization successfully detected the fusion junction, highly concordant with RT-PCR results. Gene expression profiling on 18 schwannomas demonstrated segregation largely consistent with fusion status. Fusion-positive cases expressed significantly higher HTRA1 mRNA abundance, perhaps exploitable as a biomarker. In summary, we systematically characterize a series of 60 SH3PXD2A::HTRA1 fusion-positive schwannomas, showing their distinctive morphology and location-specific prevalence for the first time.

Identification of COL1A1/2 Mutations and Fusions With Noncoding RNA Genes in Bizarre Parosteal Osteochondromatous Proliferation (Nora Lesion).

Bizarre parosteal osteochondromatous proliferation (BPOP) (Nora lesion) is a benign bone surface lesion, which most commonly occurs in the digits of young patients and has a high rate of recurrence. Histologically, it is composed of a mixture of disorganized bone, cartilage, and spindle cells in variable proportions and characterized by amorphous "blue bone" mineralization. Recurrent chromosomal abnormalities, including t(1;17)(q32-42;q21-23) and inv(7)(q21.1-22q31.3-32), have been reported in BPOP. However, the exact genes involved in the rearrangements remain unknown. In this study, we analyzed 8 BPOP cases affecting the fingers, toe, ulna, radius, and fibula of 5 female and 3 male patients, aged 5 to 68 years. RNA sequencing of 5 cases identified genetic fusions between COL1A2 and LINC-PINT in 3 cases and COL1A1::MIR29B2CHG fusion in 1, both validated using fluorescence in situ hybridization and reverse transcription (RT)-PCR. The remaining fusion-negative case harbored 3 COL1A1 mutations as revealed by whole-exome sequencing and confirmed using Sanger sequencing. All these genetic alterations were predicted to cause frameshift and/or truncation of COL1A1/2. The chromosomal locations of COL1A2 (7q21.3), LINC-PINT (7q32.3), COL1A1 (17q21.33), and MIR29B2CHG (1q32.2) were consistent with the breakpoints identified in the previous cytogenetic studies. Subsequent screening of 3 BPOPs using fluorescence in situ hybridization identified 1 additional case each with COL1A1 or COL1A2 rearrangement. Our findings are consistent with reported chromosomal abnormalities and implicate the disruption of type I collagen, and perhaps of either noncoding RNA gene as a tumor suppressor, in the tumorigenesis of BPOP. The prevalence and tumorigenic mechanisms of these COL1A1/2 alterations in BPOP require further investigation.

Extraskeletal Myxoid Chondrosarcomas: The Uncommon Clinicopathologic Manifestations and Significance of TAF15::NR4A3 Fusion.

Extraskeletal myxoid chondrosarcoma (EMC) is an ultrarare sarcoma typically exhibiting myxoid/reticular histology and NR4A3 translocation. However, morphologic variants and the relevance of non-EWSR1::NR4A3 fusions remain underexplored. Three challenging pan-Trk-expressing cases, featuring cellular to solid histology, were subjected to RNA exome sequencing (RES), unveiling different NR4A3-associated fusions. Alongside RES-analyzed cases, fluorescence in situ hybridization was performed to confirm 58 EMCs, with 48 available for pan-Trk immunostaining and KIT sequencing. Except for 1 (2%) NR4A3-rearranged EMC without identifiable partners, 46 (79%), 9 (16%), and 2 (3%) cases harbored EWSR1::NR4A3, TAF15::NR4A3, and TCF12::NR4A3 fusions, respectively. Five EWSR1::NR4A3-positive EMCs occurred in the subcutis (3) and bone (2). Besides 43 classical cases, there were 8 cellular, 4 rhabdoid/anaplastic, 2 solid, and 1 mixed tumor-like variants. Tumor cells were oval/spindle to pleomorphic and formed loose myxoid/reticular to compact sheet-like or fascicular patterns, imparting broad diagnostic considerations. RES showed upregulation of NTRK2/3, KIT, and INSM1. Moderate-to-strong immunoreactivities of pan-Trk, CD117, and INSM1 were present in 35.4%, 52.6%, and 54.6% of EMCs, respectively. KIT p. E554K mutation was detected in 2/48 cases. TAF15::NR4A3 was significantly associated with size >10 cm (78%, P = .025). Size >10 cm, moderate-to-severe nuclear pleomorphism, metastasis at presentation, TAF15::NR4A3 fusion, and the administration of chemotherapy portended shorter univariate disease-specific survival, whereas only size >10 cm (P = .004) and metastasis at presentation (P = .032) remained prognostically independent. Conclusively, EMC may manifest superficial or osseous lesions harboring EWSR1::NR4A3, underrecognized solid or anaplastic histology, and pan-Trk expression, posing tremendous challenges. Most TAF15::NR4A3-positive cases were >10 cm in size, ie, a crucial independent prognosticator, whereas pathogenic KIT mutation rarely occurred.

Dunaliella salina Alga Protects against Myocardial Ischemia/Reperfusion Injury by Attenuating TLR4 Signaling.

Myocardial ischemia/reperfusion (I/R) injury is marked by rapid increase in inflammation and not only results in myocardial apoptosis but also compromises the myocardial function. Dunaliella salina (D. salina), a halophilic unicellular microalga, has been used as a provitamin A carotenoid supplement and color additive. Several studies have reported that D. salina extract could attenuate lipopolysaccharides-induced inflammatory effects and regulate the virus-induced inflammatory response in macrophages. However, the effects of D. salina on myocardial I/R injury remain unknown. Therefore, we aimed to investigate the cardioprotection of D. salina extract in rats subjected to myocardial I/R injury that was induced by occlusion of the left anterior descending coronary artery for 1 h followed by 3 h of reperfusion. Compared with the vehicle group, the myocardial infarct size significantly decreased in rats that were pre-treated with D. salina. D. salina significantly attenuated the expressions of TLR4, COX-2 and the activity of STAT1, JAK2, IκB, NF-κB. Furthermore, D. salina significantly inhibited the activation of caspase-3 and the levels of Beclin-1, p62, LC3-I/II. This study is the first to report that the cardioprotective effects of D. salina may mediate anti-inflammatory and anti-apoptotic activities and decrease autophagy through the TLR4-mediated signaling pathway to antagonize myocardial I/R injury.

Adult NTRK-rearranged spindle cell neoplasms of the viscera: with an emphasis on rare locations and heterologous elements.

NTRK-rearranged mesenchymal neoplasms mostly affect the soft tissues of pediatric patients. Given the responsiveness to selective NTRK inhibitors, it remains critical to identify those ultra-rare cases occurring in the viscera of adults. In five females and two males aged 18-53 years, we characterized visceral mesenchymal tumors harboring TPM3-NTRK1 [uterine cervix (N = 2), pleura, prostate], LMNA-NTRK1 (lung), SQSTM1-NTRK3 (heart), and NTRK3 rearrangement with unknown fusion partner (colon/mesocolon) with RNA sequencing, FISH, RT-PCR, and immunohistochemistry. The tumors exhibited spindled to ovoid/epithelioid or pleomorphic cells, often arranged in fascicles, and were low-to-intermediate-grade and high-grade in three and four cases, respectively. Keloid-like stromal collagen and perivascular hyalinization was noted in five. Adenosarcoma-like appearances were observed in two, manifesting frond-like protrusions in one cervical tumor and phyllodes-like architecture in the prostatic tumor. Abrupt high-grade transformation into pleomorphic liposarcoma was found in another cervical tumor, while the pleural tumor contained intermixed rhabdomyoblasts. Pan-TRK immunostaining was positive in all cases. All cases expressed CD34, while five were S100-positive. CDKN2A homozygous deletion with concomitant p16 loss occurred in 4/7. Whole-exome sequencing identified TP53 mutation (c.672+2T>C, involving a splice site, with concomitant protein loss) in a cervical sarcoma, limited to its heterologous liposarcomatous component. At least moderate pan-TRK immunoreactivity was present in varying proportions of potential pathologic mimics, with BCOR-positive sarcoma (56%, 5/9), undifferentiated uterine sarcoma (50%, 3/6), and spindle cell/sclerosing rhabdomyosarcoma (33%, 2/6) being among the most frequent. This underscored the unsatisfactory specificity of pan-TRK immunohistochemistry and warranted molecular confirmation in the diagnosis of adult NTRK-rearranged visceral mesenchymal neoplasms. The current report highlights the ever-expanding clinicopathologic and genetic spectrum of this entity by describing the unprecedented cardiac and pleural locations and heterologous differentiation, as well as the second NTRK-rearranged "prostatic stromal sarcoma," while substantiating CDKN2A deletion as a frequent occurrence.

Clinicopathological and molecular characterisation of USP6-rearranged soft tissue neoplasms: the evidence of genetic relatedness indicates an expanding family with variable bone-forming capacity.

AIMS: USP6 rearrangement underpins self-limiting fibroblastic/myofibroblastic neoplasms, including nodular fasciitis (NF), myositis ossificans (MO), aneurysmal bone cyst (ABC), and related variants. The aim of this study was to characterise UPS6 and fusion partners in order to delineate the clinicopathological, genetic and bone-forming features in such lesions of soft tissue (ST). METHODS AND RESULTS: Break-apart fluorescence in-situ hybridisation (FISH) validated USP6 rearrangement in 31 of 35 NF [comprising three of three fasciitis ossificans (FO) cases, seven of eight cellular variant of fibroma of tendon sheath (C-FTS), four of six MO, three of three ST-ABC, and two of two fibro-osseous pseudotumours of digits (FOPD)]. As determined with FISH and reverse transcription polymerase chain reaction, MYH9-USP6 was the commonest fusion in four C-FTS and 20 NF, including one intravascular case and two infantile (one retroperitoneal) cases. The presence of MYH9-USP6 confirmed the diagnosis of two NFs> 50 mm with prominent ischaemic necrosis. COL1A1-USP6 was predominant in ossifying lesions, including all FO, MO, ST-ABC and FOPD with identified partner genes, and was also present in non-ossifying head and neck NF (HN-NF) and C-FTS in two cases each. A cervical NF of a 14-month-old girl harboured the novel COL1A2-USP6. Ossifying lesions showed considerable genetic and morphological overlaps. Sharing COL1A1-USP6, FO and FOPD showed similar central or haphazard bone matrix deposition. Besides zonation of outward bone maturation, four COL1A1-USP6-positive MO had incipient to sieve-like pseudocysts reminiscent of ST-ABC. CONCLUSION: MYH9-USP6 is present in some C-FTS and most NF, including rare variants, but is unrelated to bone formation. All bone-forming USP6-rearranged lesions adopt COL1A1 as the 5' partner, indicating close genetic kinships. However, COL1A1/COL1A2 also contributes to the pathogenesis of minor subsets of non-ossifying USP6-rearranged HN-NF and C-FTS.

Prolonged preoperative fasting induces postoperative insulin resistance by ER-stress mediated Glut4 down-regulation in skeletal muscles.

Preoperative fasting aims to prevent pulmonary aspiration and improve bowel preparation, but it may induce profound systemic catabolic responses that lead to protein breakdown and insulin-resistant hyperglycemia after operation. However, the molecular mechanisms of catabolic reaction induced by prolonged preoperative fasting and surgical stress are undetermined. In this study, anesthetized rats were randomly assigned to receive a sham operation or laparotomy cecectomy. Fasting groups were restricted from food and water for 12 h before operation, while the feeding group had free access to food throughout the study period. Twenty-four hours after operation, the animals were sacrificed to collect blood samples and soleus muscles for analysis. Postoperative blood glucose level was significantly increased in the fasting group with elevated serum insulin and C-peptide. Continuous feeding reduced serum myoglobin and lactate dehydrogenase concentrations. Preoperative fasting activated inositol-requiring transmembrane kinase/endoribonuclease (IRE)-1α and c-Jun N-terminal kinase (JNK) mediated endoplasmic reticulum (ER)-stress, and reduced glucose transporter type 4 (Glut4) expression in the soleus muscle. Phospholamban phosphorylation was reduced and intracellular calcium levels were increased in the isolated skeletal muscle cells. Similar results were found in ER stress-induced C1C12 myoblasts. The expression of Glut4 was suppressed in the stressed C1C12, but was potentiated following inhibition of ER stress and chelation of intracellular free calcium. This study provides evidence demonstrating that prolonged preoperative fasting induces ER stress and generates insulin resistance in the skeletal muscle through suppression of Glut4 and inactivation of Ca2+-ATPase, leading to intracellular calcium homeostasis disruption and peripheral insulin resistance.

Immunohistochemistry Helps to Distinguish Noninvasive Follicular Thyroid Neoplasm with Papillary-like Nuclear Features/Noninvasive Encapsulated Follicular Variant of Papillary Thyroid Carcinoma with Other Follicular Thyroid Lesions.

Background and Objectives: We aimed to assess the diagnostic value of various immunohistochemical (IHC) markers and panels for differentiation among benign follicular nodules (BFNs), noninvasive follicular thyroid neoplasms with papillary-like nuclear features (NIFTPs), noninvasive encapsulated follicular variants of papillary thyroid carcinoma (NEFVPTCs), and infiltrative FVPTC (IFVPTC). Materials and Methods: Sixty-three cases were classified as BFNs, NIFTPs, NEFVPTCs, or IFVPTCs and were evaluated using the following markers: CK19, CD56, galectin-3, CITED1, HBME-1, VE1, and TROP-2. Results: The IHC results for NIFTP and NEFVPTC exhibited no statistically significant differences. In differentiating IFVPTCs from BFNs and NIFTPs/NEFVPTCs, galectin-3 and TROP-2 were the markers with the highest sensitivity plus high specificity, respectively. In various combinations, panel co-expression of two markers, including galectin-3 and/or HBME-1 and/or TROP-2, and the combination of galectin-3 and TROP-2 co-expression could achieve 100% in all aspects. In terms of discrimination of BFNs from NIFTP/NEFVPTC, CK19 was the single most sensitive marker (81.3%), while CD56 was the most specific (100%). The panel consisting of CK19 and/or HBME-1 exhibited the greatest sensitivity (96.9%), but the panel with CD56 and/or HBME-1 exhibited the greatest specificity (90.5%). Conclusions: Our results broaden the use of IHC markers for differential diagnoses among the four groups of follicular-based lesions. In addition, the similar IHC profiles of NIFTP and NEFVPTC also suggest the original criterion of <1% papillae within tumors, providing a reliable NIFTP diagnosis. Their close relationship may represent a spectrum of progressing neoplasia.

Frequent overexpression of klotho in fusion-negative phosphaturic mesenchymal tumors with tumorigenic implications.

Phosphaturic mesenchymal tumors (PMT) are tumors that cause hypophosphatemia/osteomalacia chiefly by secreting FGF23. We have identified FN1-FGFR1/FGF1 fusion genes in nearly half of PMT, suggesting a central role of FGFR1 pathways in the pathogenesis of PMT. Tumorigenic drivers are unknown for tumors where previous study detected neither fusion, including many in bone, where FISH failed because of tissue decalcification. To identify alternative fusions in PMT without known fusions, as well as to validate the positive FISH results and characterize the fusion junctions, 34 PMT were studied, including 12 with known FN1-FGFR1 fusion by FISH (Group A), 2 with FN1-FGF1 (B), 12 with neither fusion (C), and 8 with previous acid-based decalcification and hence unknown fusion status (D). In total, 23 archival samples were subjected to anchored multiplex PCR-based RNA-sequencing (AMP-seq) with primers targeting FN1, genes encoding the FGF/FGFR families, and KL (α-Klotho); five Group C cases were also studied with whole-transcriptomic and exome-captured RNA sequencing, respectively. The AMP-seq results were consistent with previous FISH and/or transcriptomic sequencing data, except in one old Group A sample. One case had a novel FGFR1 exon 9 breakpoint, confirmed by genomic DNA sequencing. One Group D bone tumor was found to harbor FN1-FGF1. All 3 RNA-sequencing platforms failed to identify convincing fusion genes in Group C (N = 10), which instead expressed significantly higher levels of either KL or KLB. This result was further confirmed with KL and KLB RNA CISH semi-quantification (RNAscope). Our results demonstrated the utility of AMP-seq, which was compromised by decalcification and prolonged archiving. Of potential importance, fusion-negative PMT frequently overexpressed α-Klotho (or instead ß-Klotho less commonly), whose role as an obligatory co-receptor for FGF23-FGFR1 binding suggests its aberrant expression in osteocytes/osteoblasts might result in an FGF23-FGFR1 autocrine loop that in turn drives the overexpression of FGF23 and tumorigenesis through activated FGFR pathways.

The expanding morphological and genetic spectrum of MYOD1-mutant spindle cell/sclerosing rhabdomyosarcomas: a clinicopathological and molecular comparison of mutated and non-mutated cases.

AIMS: Spindle cell/sclerosing rhabdomyosarcomas (SC/SRMS) feature spindled and/or rounded rhabdomyosarcomatous cells within variably hyalinised stroma. Only 30-67% of SC/SRMSs harbour neomorphic MYOD1 p.L122R mutations, indicating heterogeneity in this RMS type. We compared MYOD1-mutant and non-mutant cases to characterise the histological and genetic spectrum of mutated SC/SRMS. METHODS AND RESULTS: Seventeen RMSs with spindled, sclerosing or hybrid histology were sequenced to identify MYOD1 and PIK3CA mutations and reappraised to assess histological features and myogenic immunophenotypes. Twelve SC/SRMSs harboured MYOD1 mutations, including homozygous p.L122R (n = 8), heterozygous p.L122R (n = 3) and heterozygous p.E118K (n = 1). MYOD1-mutant tumours affected nine females and three males aged 8-64 years (median = 22.5), had a median size of 4.2 cm (range = 2-22) and involved the head and neck (n = 7), extremities (n = 4) and mediastinum (n = 1). Fascicular/spindle histology was predominant in four cases, including one with heterologous lipoblasts in focally myxoid stroma. Four sclerosing cases mainly comprised rounded cells, including one with multinucleated tumour cells. Four cases were histologically hybrid. The only PIK3CA (p.H1047R) mutation was detected in a predominantly spindled MYOD1-p.L122R-mutated case, but not in its laser-microdissected lipoblast-containing area. All MYOD1-mutant cases exhibited diffuse MYOD1 expression but patchy myogenin reactivity. At final follow-up (median = 13.5 months), recurrences (n = 4), metastases (n = 2) or both (n = 1) occurred in seven MYOD1-mutant cases; one had died of disease. Five non-mutated cases were reclassified as spindle embryonal (n = 3), dense embryonal (n = 1) and unclassifiable (n = 1) RMSs. CONCLUSION: MYOD1-mutant RMSs are uncommonly mutated with PIK3CA and behave aggressively with an expanded morphological and genetic spectrum, including lipoblastic differentiation, multinucleated cells and the alternative p.E118K mutation.

Clinicopathological and genetic heterogeneity of the head and neck solitary fibrous tumours: a comparative histological, immunohistochemical and molecular study of 36 cases.

AIMS: Solitary fibrous tumour (SFT) harbours recurrent inv12(q13q13)-derived NAB2-STAT6 fusions, resulting in STAT6 nuclear expression. SFTs affecting the head and neck are rare, for which we reported their clinicopathological, immunohistochemical, and genetic features. METHODS AND RESULTS: With 19 cases assessable for NAB2-STAT6 fusions, 36 head and neck SFTs (18 males; 18 females) diagnosed between 13 and 79 years (median, 47 years) of age were analysed for clinicopathological features and STAT6 immunoexpression. These SFTs, ranging from 5 to 80 mm (median, 25 mm), affected the oral cavity/pharynx (12), orbit (11), sinonasal structures (seven), and somatic soft tissues or skull (six). Histologically, 20 SFTs were conventional, six were giant-cell angiofibroma-like, one was fat-forming, four were cellular/atypical, and five were malignant (two developing metastases). STAT6 distinctively decorated the tumoral nuclei in 35 (97.2%) SFTs, but not in 29 site-relevant histological mimics categorized into 12 entities. Sixteen (84.2%) SFTs showed NAB2-STAT6 fusions with highly heterogeneous exon compositions, including NAB2ex6-STAT6ex17 in four cases, NAB2ex4-STAT6ex2 in three cases, NAB2ex2-STAT6ex2, NAB2ex4-STAT6ex4, NAB2ex6-STAT6ex16 and NAB2ex6-STAT6ex18 in two cases each, and NAB2ex3-STAT6ex19 in one case. CONCLUSIONS: Nuclear STAT6 immunoexpression is sensitive and specific for distinguishing SFT from mimics. However, considerable heterogeneity exists in the head and neck SFTs regarding the locations, histological patterns, and NAB2-STAT6 fusion variants.

NAB2-STAT6 gene fusion and STAT6 immunoexpression in extrathoracic solitary fibrous tumors: the association between fusion variants and locations.

Solitary fibrous tumor (SFT) is a rare mesenchymal neoplasm harboring NAB2-STAT6 fusion, which drives STAT6 nuclear relocation. For extrathoracic SFTs, the clinical relevance of this molecular hallmark remains obscure. We assessed STAT6 immunoexpression for 61 extrathoracic SFTs exclusive of the meninges and head and neck, and 25 had analyzable RNAs to distinguish fusion variants by RT-PCR. The immunohistochemical and molecular findings were correlated with clincopathological features and disease-free survival (DFS). Twenty-eight males and 33 females had SFTs in the body cavities (n = 31), extremities (n = 17), and trunk (n = 13), categorized into 53 non-malignant and 8 malignant tumors. The vast majority (n = 57, 93%) exhibited distinctive STAT6 nuclear expression, including malignant ones. The common fusion variants were NAB2ex6-STAT6ex16/17 in 13 SFTs and NAB2ex4-STAT6ex2 in 8, while miscellaneous variants were detected only in 4 SFTs in the limbs and trunk but not in any body cavity-based cases (P = 0.026). The worse DFS was univariately associated with malignant histology (P = 0.04) but unrelated to tumor size, location, or fusion variant. Conclusively, extrathoracic SFTs mostly harbor NAB2ex6-STAT6ex16/17, followed by NAB2ex4-STAT6ex2. Miscellaneous variants are significantly rare in SFTs within the body cavities. The clinical aggressiveness of extrathoraic SFTs is associated with malignant histology but unrelated to the NAB2-STAT6 fusion variants.

NAB2-STAT6 fusion types account for clinicopathological variations in solitary fibrous tumors.

Solitary fibrous tumor (SFT) is characterized by the inv12(q13q13)-derived NAB2-STAT6 fusion, which exhibits variable breakpoints and drives STAT6 nuclear expression. The implications of NAB2-STAT6 fusion variants in pathological features and clinical behavior remain to be characterized in a large cohort of SFTs. We investigated the clinicopathological correlates of this genetic hallmark and analyzed STAT6 immunoexpression in 28 intrathoracic, 37 extrathoracic, and 23 meningeal SFTs. These 88 tumors were designated as histologically nonmalignant in 75 cases and malignant in 13, including 1 dedifferentiated SFT. Eighty cases had formalin-fixed and/or fresh samples to extract assessable RNAs for RT-PCR assay, which revealed NAB2-STAT6 fusion variants comprising 12 types of junction breakpoints in 73 fusion-positive cases, with 65 (89%) falling into 3 major types. The predominant NAB2ex4-STAT6ex2 (n=33) showed constant breakpoints at the ends of involved exons, whereas the NAB2ex6-STAT6ex16 (n=16) and NAB2ex6-STAT6ex17 (n=16) might exhibit variable breakpoints and incorporate NAB2 or STAT6 intronic sequence. Including 73 fusion-positive and 7 CD34-negative SFTs, STAT6 distinctively labeled 87 (99%) SFTs in nuclei, exhibited diffuse reactivity in 73, but did not decorate 98 mimics tested. In seven fusion-negative cases, 6 were STAT6-positive, suggesting rare fusion variants not covered by RT-PCR assay. Regardless of histological subtypes, intrathoracic SFTs affected older patients (P=0.035) and tended to be larger in size (P=0.073). Compared with other variants, NAB2ex4-STAT6ex2/4 fusions were significantly predominant in the SFTs characterised by intrathoracic location (P<0.001), older age (P=0.005), decreased mitoses (P=0.0028), and multifocal or diffuse STAT6 staining (P=0.013), but not found to correlate with disease-free survival. Conclusively, STAT6 nuclear expression was distinctive in the vast majority of SFTs, including all fusion-positive tumors, and exploitable as a robust diagnostics of CD34-negative cases. Despite the associations of NAB2-STAT6 fusion variants with several clincopathological factors, their prognostic relevance should be further validated in large-scale prospective studies of SFTs.

Targeting IL-11R/EZH2 signaling axis as a therapeutic strategy for osteosarcoma lung metastases.

Lung metastases are the primary cause of death for osteosarcoma (OS) patients. We recently validated interleukin-11 receptor α (IL-11Rα) as a molecular target for the inhibition of OS lung metastases. Since there is no clinically approved antibody against this receptor, we sought to identify downstream targets that mediate the effects of IL-11Rα signaling. We used shRNA to deplete IL-11Rα from OS cells; as a complementary approach, we added IL-11 exogenously to OS cells. The resulting changes in gene expression identified EZH2 as a downstream candidate. This was confirmed by knockdown of IL-11Rα in OS cells, which led to increased expression of genes repressed by histone methyltransferase EZH2, including members of the WNT pathway, a known target pathway of EZH2. Exogenous IL-11 increased the global levels of histone H3 lysine 27 trimethylation, evidence of EZH2 activation. Treatment with the EZH2 inhibitor GSK126 significantly reduced in vitro proliferation and increased cell-cycle arrest and apoptosis, which were partially mediated through the WNT pathway. In vivo, treatment of an orthotopic nude mouse model of OS with GSK126 inhibited lung metastatic growth and prolonged survival. In addition, significantly shorter recurrence-free survival was seen in OS patients with high levels of EZH2 in their primary tumors (P < .05). This suggests that IL-11Rα promotes OS lung metastasis via activation of EZH2. Thus, blocking EZH2 activity may be an effective strategy for inhibiting OS lung metastasis and improving prognosis.

Cordyceps sinensis protects against renal ischemia/reperfusion injury in rats.

Cordyceps sinensis (CS) is an entomogenous fungus used as a tonic food and Chinese medicine to replenish health. This study investigated the protective effects of CS in rats post-renal ischemia-reperfusion (I/R) sequence by analyzing the influence on stromal cell-derived factor-1α (SDF-1α and chemokine (C-X-C motif) receptor 4 (CXCR4) expressions and senescence during recovery. Chemokine SDF-1 [now called chemokine C-X-C motif ligand 12 (CXCL12)] and its receptor CXCR4 are crucial in kidney repair after ischemic acute renal failure. CS treatment significantly alleviated I/R-induced renal damage assessed by creatinine levels (p < 0.05) and abated renal tubular damages assessed by periodic acid-Schiff with diastase (PASD) staining. CS induced early SDF-1α expression and increased CXCR4 expression 1-6 h post-reperfusion. Histology studies have revealed that CS induced SDF-1α in squamous cells of Bowman's capsule, mesangial cells, distal convoluted tubules (DCT), and proximal convoluted tubules (PCT). CS also improved renal repair in I/R-induced injury by increasing Ki-67 staining. I/R induced renal senescence after 3 and 6 h of reperfusion. However, CS alleviated I/R-induced senescence at early stage (1 and 3 h). We conclude that CS protects against I/R injury via the SDF-1/CXCR4-signaling axis and alleviates senescence.

Recurrent amplification at 7q21.2 Targets CDK6 gene in primary myxofibrosarcomas and identifies CDK6 overexpression as an independent adverse prognosticator.

BACKGROUND: Myxofibrosarcoma is genetically complex and remains obscure in molecular determinants of clinical aggressiveness. Our prior study revealed recurrent gains of 7q in myxofibrosarcomas where MET and CDK6 genes displayed increased DNA copies. Previously, we demonstrated the implication of MET overexpression, prompting us to further elucidate the roles of CDK6 in myxofibrosarcomas. MATERIALS: On tissue microarrays, CDK6 immunoexpression was assessable in 77 primary tumors, 55 of which were successfully quantified for CDK6 and MET genes by real-time PCR using genomic DNA extracted from laser-microdissected tumor cells. Gene status and protein expression of CDK6 were correlated with each other, clinicopathological variables, metastasis-free survival (MFS), and disease-specific survival (DSS). RESULTS: Protein overexpression and gene amplification of CDK6, which were detected in 21 of 77 (27.2 %) and 13 of 55 cases (23.6 %), respectively, were highly related to each other (p < .001) and associated with higher grades (overexpression, p = .004; amplification, p = .014). There was a strong correlation between CDK6 and MET gene copies (p < .001, r = 0.0714). Importantly, CDK6 protein overexpression (MFS, p = .0002; DSS, p = .0015) and gene amplification (MFS, p = .0001; DSS, p = .0083) were both univariately associated with worse outcomes. Together with nonextremity location and AJCC stage III disease, CDK6 overexpression independently portended inferior MFS (p = .0015, risk ratio [RR] = 7.411). This aberration, along with nonextremity location, was also an independent adverse prognosticator of DSS (p = .0069, RR = 6.006). CONCLUSIONS: In approximately a quarter of primary myxofibrosarcomas, CDK6 overexpression is mostly driven by gene amplification on 7q, associated with adverse prognosticators, and independently predictive of worse outcomes, highlighting its possible causative role in tumor aggressiveness.

Molecular Characterization of Dermatofibrosarcoma Protuberans: The Clinicopathologic Significance of Uncommon Fusion Gene Rearrangements and Their Diagnostic Importance in the Exclusively Subcutaneous and Circumscribed Lesions.

The clinicopathologic relevance of various gene rearrangements underlying dermatofibrosarcoma protuberans (DFSP) remains insufficiently characterized. In 188 DFSPs, we determined PDGFB, COL1A1, PDGFD, COL6A3, and EMILIN2 rearrangements by fluorescence in situ hybridization (FISH). The clinicopathologic significance of rearrangement types and factors related to recurrence and metastasis were statistically analyzed. In all, classic PDGFB rearrangement, cryptic COL1A1-PDGFB fusion, and PDGFD rearrangement were identified in 172 (91.4%), 8 (4.3%), and 8 (4.3%: 4 COL6A3-PDFGD, 4 EMILIN2-PDGFD) cases, respectively. In an index DFSP harboring the cryptic fusion, the COL1A1-PDGFB transcript was confirmed by both RNA sequencing and reverse transcription-polymerase chain reaction. In comparison with cases harboring classic PDGFB rearrangement, cryptic PDGFB-rearranged DFSPs usually exhibited higher 5'-COL1A1 copy numbers. In a combined reappraisal of published and current cases, COL6A3-PDGFD-positive DFSPs (n=16) predominated in females (n=14, 88%) and torso (n=14, 88%), especially the breast (n=7, 44%); EMILIN2-PDGFD-positive DFSPs (n=6) preferentially demonstrated near exclusively subcutaneous growth (n=5, 83%) and fibrosarcomatous transformation (n=5, 83%). In our cohort, local recurrence was related to fibrosarcomatous variant (P=0.029, odds ratio=3.478) and head and neck location (P=0.046, odds ratio=3.508). Distant metastasis only occurred in the fibrosarcomatous variant (9/73, 12.3%) but not in other cases. In conclusion, 8.6% of DFSPs are negative for PDGFB break-apart FISH, which, especially those with challenging subcutaneous and circumscribed manifestation, require complementary diagnosis by FISH assays targeting COL1A1 and PDGFD. The types of fusion gene rearrangements, head and neck location, and fibrosarcomatous transformation may account for clinicopathologic and prognostic variations in DFSPs and warrant future independent validation.

Micromachined optical fiber enclosed 4-electrode IPMC actuator with multidirectional control ability for biomedical application.

The present paper examined a novel micromachined column structured Ion Polymer Metal Composite (IPMC) actuator with multidirectional control capability. The developed 4-electrode transducer enclosed a section of optical fiber, thereby allowing electronic directional control of conducted laser light. The fabricated device with IPMC actuator dimensions of 5 mm x 2 mm x 1 mm reached a maximum displacement of 400 µm when a square wave of 9 V was applied to the top-bottom electrode pair. Displacements in different directions and moving angles were characterized with side-side and top-right electrode pairs connected to the actuating signals. Furthermore, the generating moment per volt per second by the transducer was analyzed. The maximum value of approximately 200 µN*m/V/s was displayed when the device actuated with the side-side electrode pair. Controlling the developed IPMC actuator moved the laser beam in multiple directions. This device could be promising for biomedical applications such as microendoscopic ocular surgery.

Morphologic Spectrum of Lymphadenopathy in Adult-onset Immunodeficiency (Anti-interferon-γ Autoantibodies).

Adult-onset immunodeficiency syndrome (AOIS) caused by anti-interferon-γ autoantibodies is an emerging disease. Affected patients present typically with systemic lymphadenopathy, fatigue, and fever. We studied 36 biopsy specimens, 31 lymph nodes, and 5 extranodal sites, of AOIS confirmed by serum autoantibody or QuantiFERON-TB Gold In-Tube assay. We describe the morphologic features and the results of ancillary studies, including special stains, immunohistochemistry, and molecular testing. The overall median age of these patients was 60.5 years (range, 41 to 83 y) with a male-to-female ratio of 20:16. All biopsy specimens showed nontuberculous mycobacterial infection, and most cases showed the following histologic features: capsular thickening with intranodal sclerosing fibrosis, irregularly distributed ill-formed granulomas or histiocytic aggregates with neutrophilic infiltration, interfollicular expansion by a polymorphic infiltrate with some Hodgkin-like cells that commonly effaces most of the nodal architecture and proliferation of high endothelial venules. In situ hybridization analysis for Epstein-Barr virus-encoded RNA showed scattered (<1%) to relatively more common (4% to 5%) positive cells in 29 of 30 (97%) tested specimens, reflecting immune dysregulation due to an interferon-γ defect. In the 31 lymph node specimens, 23 (74%) cases showed increased immunoglobulin G4-positive plasma cells (4 to 145/HPF; mean, 49.7/HPF) with focal areas of sclerosis reminiscent of immunoglobulin G4-related lymphadenopathy, 4 (13%) cases resembled, in part, nodular sclerosis Hodgkin lymphoma, and 9 (29%) cases mimicked T-cell lymphoma. Among 33 patients with available clinical follow-up, 20 (61%) showed persistent or refractory disease despite antimycobacterial therapy, and 1 patient died of the disease. We conclude that the presence of ill-defined granulomas, clusters of neutrophils adjacent to the histiocytic aggregates, and some Epstein-Barr virus-positive cells are features highly suggestive of AOIS. A high index of clinical suspicion and awareness of the morphologic features and differential diagnosis of AOIS are helpful for establishing the diagnosis.