10× single-cell sequencing revealed cellular composition heterogeneity in cardiac myxoma with malignant glandular properties.

Cardiac myxoma is the most common primary cardiac tumor in adults. The histogenesis and cellular composition of myxoma are still unclear. This study aims to reveal the role of myxoma cell components and their gene expression in tumor development. We obtained single living cells by enzymatic digestion of tissues from 4 cases of surgically resected cardiac myxoma. Of course, there was 1 case of glandular myxoma and 3 cases of nonglandular myxoma. Then, 10× single-cell sequencing was performed. We identified 12 types and 11 types of cell populations in glandular myxoma and nonglandular myxoma, respectively. Heterogeneous epithelial cells are the main components of glandular myxoma. The similarities and differences in T cells in both glandular and nonglandular myxoma were analyzed by KEGG and GO. The most important finding was that there was active communication between T cells and epithelial cells. These results clarify the possible tissue occurrence and heterogeneity of cardiac myxoma and provide a theoretical basis and guidance for clinical diagnosis and treatment.

Autocrine Regulation of Interleukin-6 via the Activation of STAT3 and Akt in Cardiac Myxoma Cells.

Plasma concentrations of a pleiotropic cytokine, interleukin (IL)-6, are increased in patients with cardiac myxoma. We investigated the regulation of IL-6 in cardiac myxoma. Immunohistochemical staining and reverse transcription-polymerase chain reaction (RT-PCR) revealed that IL-6 and its receptors, IL-6 receptor (IL-6R) and gp130, co-existed in the myxoma cells. Myxoma cells were cultured, and an antibody array assay showed that a conditioned medium derived from the cultured myxoma cells contained increased amounts of IL-6. Signal transducer and activator of transcription (STAT) 3 and Akt were constitutively phosphorylated in the myxoma cells. An enzyme-linked immunosorbent assay (ELISA) showed that the myxoma cells spontaneously secreted IL-6 into the culture medium. Real-time PCR revealed that stimulation with IL-6 + soluble IL-6R (sIL6R) significantly increased IL-6 mRNA in the myxoma cells. Pharmacological inhibitors of STAT3 and Akt inhibited the IL-6 + sIL-6R-induced gene expression of IL-6 and the spontaneous secretion of IL-6. In addition, IL-6 + sIL-6R-induced translocation of phosphorylated STAT3 to the nucleus was also blocked by STAT3 inhibitors. This study has demonstrated that IL-6 increases its own production via STAT3 and Akt pathways in cardiac myxoma cells. Autocrine regulation of IL-6 may play an important role in the pathophysiology of patients with cardiac myxoma.

The benign nature and rare occurrence of cardiac myxoma as a possible consequence of the limited cardiac proliferative/ regenerative potential: a systematic review.

BACKGROUND: Cardiac Myxoma is a primary tumor of heart. Its origins, rarity of the occurrence of primary cardiac tumors and how it may be related to limited cardiac regenerative potential, are not yet entirely known. This study investigates the key cardiac genes/ transcription factors (TFs) and signaling pathways to understand these important questions. METHODS: Databases including PubMed, MEDLINE, and Google Scholar were searched for published articles without any date restrictions, involving cardiac myxoma, cardiac genes/TFs/signaling pathways and their roles in cardiogenesis, proliferation, differentiation, key interactions and tumorigenesis, with focus on cardiomyocytes. RESULTS: The cardiac genetic landscape is governed by a very tight control between proliferation and differentiation-related genes/TFs/pathways. Cardiac myxoma originates possibly as a consequence of dysregulations in the gene expression of differentiation regulators including Tbx5, GATA4, HAND1/2, MYOCD, HOPX, BMPs. Such dysregulations switch the expression of cardiomyocytes into progenitor-like state in cardiac myxoma development by dysregulating Isl1, Baf60 complex, Wnt, FGF, Notch, Mef2c and others. The Nkx2-5 and MSX2 contribute predominantly to both proliferation and differentiation of Cardiac Progenitor Cells (CPCs), may possibly serve roles based on the microenvironment and the direction of cell circuitry in cardiac tumorigenesis. The Nkx2-5 in cardiac myxoma may serve to limit progression of tumorigenesis as it has massive control over the proliferation of CPCs. The cardiac cell type-specific genetic programming plays governing role in controlling the tumorigenesis and regenerative potential. CONCLUSION: The cardiomyocytes have very limited proliferative and regenerative potential. They survive for long periods of time and tightly maintain the gene expression of differentiation genes such as Tbx5, GATA4 that interact with tumor suppressors (TS) and exert TS like effect. The total effect such gene expression exerts is responsible for the rare occurrence and benign nature of primary cardiac tumors. This prevents the progression of tumorigenesis. But this also limits the regenerative and proliferative potential of cardiomyocytes. Cardiac Myxoma develops as a consequence of dysregulations in these key genes which revert the cells towards progenitor-like state, hallmark of CM. The CM development in carney complex also signifies the role of TS in cardiac cells.

Cerebral embolization associated with parenchymal seeding of the left atrial myxoma: Potential role of interleukin-6 and matrix metalloproteinases.

Systemic embolization has been reported in up to 40% of patients with left atrial myxoma, half of them with cerebral involvement. However, development of intracerebral embolization associated with parenchymal seeding of the myxoma emboli is an extremely rare complication, with only 36 histologically diagnosed cases reported in the published literature. We describe a 69-year-old woman who arrived at the emergency service with hemiparesis associated with drug-resistant epilepsy and a medical history of resection of a left atrial myxoma 10 months previously. Cranial computed tomography revealed multiple large lesions of heterogeneous density and cystic components in the occipital lobes and posterior fossa parenchyma. Histopathological analyses after stereotactic biopsy of the occipital lesion revealed infiltrative myxoma cells with benign histological findings and uniform expression of calretinin similar to that of the primary cardiac myxoma. Additional immunohistochemical studies confirmed brain parenchymal seeding of the myxoma cells with strong expression of interleukin-6 (IL-6) and focal expression of matrix metalloproteinases-2 (MMP-2). Here, we discuss the clinicopathological features of intracerebral embolization of left atrial myxomas associated with progressive parenchymal seeding of the tumor emboli and the potential pathogenic role of IL-6 and MMPs.

Cardiac Myxoma: Review and Update of Contemporary Immunohistochemical Markers and Molecular Pathology.

Cardiac myxoma is an uncommon benign mesenchymal neoplasm of the heart. It usually arises in the left atrium, near the valve of the fossa ovalis, and most frequently affects adults in the third through the sixth decades of life. It is hypothesized to arise from subendothelial vasoformative reserve cells or primitive cells that differentiate along the lines of the endothelium, but this remains speculative. Microscopically, the neoplastic cells are arranged individually, and nests, and are oriented in single or multiple layers around vascular channels. The neoplastic cells are immunoreactive for vimentin, calretinin, S100, nonspecific enolase, factor VIII, CD31, and CD34. The tumor can have diverse clinical presentations depending on its location and extent of disease and is predisposed to embolization. The current treatment is prompt surgical excision.

Next generation sequencing for GNAS uncovers CD34 as a sensitive marker for intramuscular myxoma.

BACKGROUND: Intramuscular myxoma is a soft tissue myxoid tumor with a broad morphological differential diagnosis and recent developments have led to the identification of markers that can exclude some, but not all, differential diagnostic entities. However, a sensitive confirmatory marker for intramuscular myxoma has not been clearly identified. Since there is some evidence that mutations in the GNAS gene could be such a marker, we evaluated our results of next-generation sequencing testing for GNAS mutations performed in recent years on our series of intramuscular myxoma. MATERIALS AND METHODS: Next-generation sequencing was performed on 10 cases of intramuscular myxoma diagnosed between 2015 and 2019, using either the TruSight Tumor 26 panel or an in-house developed 97 cancer gene panel. Additionally, immunohistochemistry for CD34 was performed on all cases. RESULTS: All intramuscular myxomas showed a diffuse and strong expression of CD34 and a GNAS mutation was found in 88% of cases, making this a very sensitive positive test for the diagnosis of intramuscular myxoma. CONCLUSIONS: Under the condition that contemporary next-generation sequencing is applied as testing method, searching for GNAS mutations is a very sensitive confirmatory test for the diagnosis of intramuscular myxoma, obviating the necessity to perform tests that exclude other entities by the virtue of their negative result. The molecular tests results also identified strong and diffuse CD34 expression as a sensitive, albeit non-specific, marker for intramuscular myxoma.

Notch Signaling Affects Oral Neoplasm Cell Differentiation and Acquisition of Tumor-Specific Characteristics.

Histopathological findings of oral neoplasm cell differentiation and metaplasia suggest that tumor cells induce their own dedifferentiation and re-differentiation and may lead to the formation of tumor-specific histological features. Notch signaling is involved in the maintenance of tissue stem cell nature and regulation of differentiation and is responsible for the cytological regulation of cell fate, morphogenesis, and/or development. In our previous study, immunohistochemistry was used to examine Notch expression using cases of odontogenic tumors and pleomorphic adenoma as oral neoplasms. According to our results, Notch signaling was specifically associated with tumor cell differentiation and metaplastic cells of developmental tissues. Notch signaling was involved in the differentiation of the ductal epithelial cells of salivary gland tumors and ameloblast-like cells of odontogenic tumors. However, Notch signaling was also involved in squamous metaplasia, irrespective of the type of developmental tissue. In odontogenic tumors, Notch signaling was involved in epithelial-mesenchymal interactions and may be related to tumor development and tumorigenesis. This signaling may also be associated with the malignant transformation of ameloblastomas. Overall, Notch signaling appears to play a major role in the formation of the characteristic cellular composition and histological features of oral neoplasms, and this involvement has been reviewed here.

Evaluation of the osteoclastogenic process associated with RANK / RANK-L / OPG in odontogenic myxomas.

BACKGROUND: Odontogenic myxoma (OM) is a benign intraosseous neoplasm that exhibits local aggressiveness and high recurrence rates. Osteoclastogenesis is an important phenomenon in the tumor growth of maxillary neoplasms. RANK (Receptor Activator of Nuclear Factor κappa B) is the signaling receptor of RANK-L (Receptor activator of nuclear factor kappa-Β ligand) that activates the osteoclasts. OPG (osteoprotegerin) is a decoy receptor for RANK-L that inhibits pro-osteoclastogenesis. The RANK / RANKL / OPG system participates in the regulation of osteolytic activity under normal conditions, and its alteration has been associated with greater bone destruction, and also with tumor growth. OBJECTIVES: To analyze the immunohistochemical expression of OPG, RANK and RANK-L proteins in odontogenic myxomas (OMs) and their relationship with the tumor size. MATERIAL AND METHODS: Eighteen OMs, 4 small (<3 cm) and 14 large (> 3cm) and 18 dental follicles (DF) that were included as control were studied by means of standard immunohistochemical procedure with RANK, RANKL and OPG antibodies. For the evaluation, 5 fields (40x) of representative areas of OM and DF were selected where the expression of each antibody was determined. Descriptive and comparative statistical analyses were performed with the obtained data. RESULTS: There are significant differences in the expression of RANK in OM samples as compared to DF (p = 0.022) and among the OMSs and OMLs (p = 0.032). Also a strong association is recognized in the expression of RANK-L and OPG in OM samples. CONCLUSIONS: Activation of the RANK / RANK-L / OPG triad seems to be involved in the mechanisms of bone balance and destruction, as well as associated with tumor growth in odontogenic myxomas.

PPARγ suppresses the proliferation of cardiac myxoma cells through downregulation of MEF2D in a miR-122-dependent manner.

Peroxisome proliferator-activated receptor gamma (PPARγ), a multiple functional transcription factor, has been reported to have anti-tumor effects through inhibition of cells proliferation. However, its effects on cardiac myxoma (CM) cells and the underlying signaling mechanism is unclear. In the present study, we demonstrated that the level of PPARγ is inversely correlated with that of myocyte enhancer factor 2D (MEF2D), a biomarker of CM. We found that activation of PPARγ inhibit MEF2D expression via upregulation of miR-122, which can target the 3'-UTR of MEF2D and inhibit MEF2D expression, by directly binding to the PPRE in the miR-122 promoter region. Functional experiments further showed that miR-122-dependent downregulation of MEF2D by PPARγ suppress the proliferation of CM cells. These results suggest that PPARγ may exert its antiproliferative effects by negatively regulating the MEF2D in CM cells, which through upregulation of miR-122, and PPARγ/miR-122/MEF2D signaling pathway may be a novel target for treatment of CM.

Ancient cardiac myxomas - another point of view in the light of tetraspanins.

Myxomas are the most common non-invasive but life-threatening cardiac neoplasms due to obstruction of heart chambers and risk of embolism in a manner resembling thromboembolism as well. They can occasionally disseminate via their detached fragments into the bloodstream to seed and grow as secondary still benign tumors. In this study we evaluated morphological and clinical aspects of 14 ancient, degenerated left or right-sided cardiac atrial myxomas with expression of CD9 and CD63, which are found to contribute to platelet activation, aggregation and, as a result, intratumoral thrombosis or fragmentation. The appearance of tumors varied from sessile to polypoid revealing that a higher rate of endocardial thrombosis was associated with sessile compared to polypoid myxomas and left-sided tumors compared to right-sided ones in our study. In the general aspect of ancient calcifications, amorphous calcification with intra-tumor thrombosis was noted more frequently in sessile tumors, while well-formed osseous metaplasia was usually a feature of polypoid tumors. In our material osseous metaplasia did not coexist with massive thrombosis and was found in polypoid, pedunculated myxomas. Most importantly, CD9 overexpression was recorded in every studied myxoma and CD63 gave a weak reaction in myxoma cells.

Molecular basis of cardiac myxomas.

Cardiac tumors are rare, and of these, primary cardiac tumors are even rarer. Metastatic cardiac tumors are about 100 times more common than the primary tumors. About 90% of primary cardiac tumors are benign, and of these the most common are cardiac myxomas. Approximately 12% of primary cardiac tumors are completely asymptomatic while others present with one or more signs and symptoms of the classical triad of hemodynamic changes due to intracardiac obstruction, embolism and nonspecific constitutional symptoms. Echocardiography is highly sensitive and specific in detecting cardiac tumors. Other helpful investigations are chest X-rays, magnetic resonance imaging and computerized tomography scan. Surgical excision is the treatment of choice for primary cardiac tumors and is usually associated with a good prognosis. This review article will focus on the general features of benign cardiac tumors with an emphasis on cardiac myxomas and their molecular basis.

Modulation of transforming growth factor-ß signaling and extracellular matrix production in myxomatous mitral valves by angiotensin II receptor blockers.

BACKGROUND: Little is known about the pathophysiology of myxomatous degeneration of the mitral valve, the pathological hallmark of mitral valve prolapse, associated with symptomatic mitral regurgitation, heart failure, and death. Excess transforming growth factor (TGF)-ß signaling is known to cause mitral valve degeneration and regurgitation in a mouse model of Marfan syndrome. We examined if TGF-ß signaling is dysregulated in clinical specimens of sporadic mitral valve prolapse compared with explanted nondiseased mitral valves and we tested the effects of angiotensin II receptor blockers on TGF-ß signaling in cultured human mitral valve cells. METHODS AND RESULTS: Operative specimens, cultured valve tissues, and cultured valvular interstitial cells were obtained from patients with mitral valve prolapse undergoing mitral valve repair or from organ donors without mitral valve disease. Increased extracellular matrix in diseased valve tissue correlated with an upregulation of TGF-ß expression and signaling as evidenced by SMAD2/3 phosphorylation. Both TGF-ß ligand and signaling mediators colocalized primarily to valvular interstitial cells suggesting autocrine/paracrine activation. In cultured valve tissue, exogenous TGF-ß increased basal extracellular matrix production, whereas serological neutralization of TGF-ß inhibited disease-driven extracellular matrix overproduction. TGF-ß-induced extracellular matrix production in cultured valvular interstitial cells was dependent on SMAD2/3 and p38 signaling and was inhibited by angiotensin II receptor blockers. CONCLUSIONS: TGF-ß has a profibrotic role in the pathogenesis of sporadic mitral valve prolapse. Attenuation of TGF-ß signaling by angiotensin II receptor blockers may represent a mechanistically based strategy to modulate the pathological progression of mitral valve prolapse in patients.

Benign cutaneous neural tumors.

Benign cutaneous neural neoplasms are one of the most frequent benign mesenchymal tumors in the skin. Because peripheral sheath nerve is composed of different cells, the tumors raised in these structures are varied and usually contain many of these cells. Most of these tumors are easy to diagnose, as usually present characteristic features well-recognized and express -specific immunohistochemical proteins. However, there are so many infrequent variants that many times require distinction from others spindle-cell tumors including melanoma. The tumors differ from one another by displaying a different proportion and arrangement of the various constituents of a peripheral nerve. In this article, we present the most characteristic clinical and histopathological features of many of these frequent benign cutaneous neural tumors including their uncommon variants.

Intramuscular myxoma of the buttock mimicking low-grade fibromyxoid sarcoma: diagnostic usefulness of MUC4 expression.

Intramuscular myxoma (IM) is a benign intramuscular neoplasm composed of fibroblasts and abundant myxoid stroma. Some malignant soft tissue tumors can undergo myxomatous degeneration, which makes it difficult to distinguish them from IM. We describe a case of IM of the buttock region mimicking low-grade fibromyxoid sarcoma. The tumor appeared as a well-defined ovoid mass with a cystic lesion on MRI images, and mild uptake on PET images was seen. This was originally misdiagnosed as low-grade fibromyxoid sarcoma (LGFMS) after core-needle biopsy. The mass was excised en bloc and sent for histology. The surgical specimen showed the features of LGFMS with the same characteristics as those mentioned in the previous biopsy report. After surgery, MUC4 expression, a highly sensitive and specific immunohistochemical marker for LGFMS, and FUS gene rearrangement by FISH was not detected upon re-examination; therefore, a conclusive diagnosis of IM was made. The patient had no local recurrence at the 3-year follow-up. Our case suggests that IM with mild FDG uptake is frequently confused with other low-grade malignant myxoid tumors. In addition, absence of MUC4 expression is the definitive key to distinguish IM from LGFMS.

Crosstalk between KIF1C and PRKAR1A in left atrial myxoma.

Cardiac myxoma (CM) is the most common benign cardiac tumor, and most CMs are left atrial myxomas (LAMs). Six variations of KIF1C, c.899 A > T, c.772 T > G, c.352 A > T, c.2895 C > T, c.3049 G > A, and c.*442_*443dup in left atrial myxoma tissues are identified by whole-exome sequencing (WES) and Sanger sequencing. RNA-seq and function experiments show the reduction of the expression of KIF1C and PRKAR1A caused by rare variations of KIF1C. KIF1C is observed to be located in the nucleus, bind to the promoter region of PRKAR1A, and regulate its transcription. Reduction of KIF1C decreases PRKAR1A expression and activates the PKA, which causes an increase in ERK1/2 phosphorylation and SRC-mediated STAT3 activation, a reduction of CDH1, TP53, CDKN1A, and BAX, and eventually promotes tumor formation both in vitro and in vivo. The results suggest that inhibition of KIF1C promotes the pathogenesis of LAM through positive feedback formed by the crosstalk between KIF1C and PRKAR1A.

Somatic Mutation of PRKAR1A in Four Cases of Sporadic Cardiac Myxoma.

BACKGROUND: Inactivating mutations of the protein kinase A regulatory subunit 1 alpha (PRKAR1A) gene have been reported in familial cardiac myxoma. However, the role of PRKAR1A mutation in sporadic cardiac myxoma remains unknown. METHODS: Targeted next-generation sequencing (NGS) was performed to identify mutations with the PRKAR1A gene in seven cases of sporadic cardiac myxomas. Sanger sequencing of DNA from cardiac myxoma specimens and matched peripheral blood samples was performed to verify the identified mutations. RESULTS: Targeted NGS of myxoma DNA revealed 232 single nucleotide variants in 141 genes and 38 insertion-deletion mutations in 13 genes. Six PRKAR1A mutations were identified in four of the seven cardiac myxoma cases, and thus, the PRKAR1A inactivating mutation rate was 57.2% (4/7, 95% CI=0.44-0.58, P<0.05). The PRKAR1A variants identified by Sanger sequencing analysis were consistent with those from the NGS analysis for the four myxoma specimens. All of the pathogenic PRKAR1A mutations led to premature termination of PRKAR1A, except for one synonymous mutation. Moreover, none of the nonsense and missense mutations found in the myxoma specimens were found in the matched peripheral blood samples. CONCLUSION: Pathogenic mutations of the PRKAR1A gene were identified in tumor specimens from four cases of sporadic cardiac myxoma, and the absence of these mutations in peripheral blood samples demonstrated that they were somatic mutations.

Superficial Angiomyxoma of the Wrist: Case Report and Literature Review.

BACKGROUND/AIM: Superficial angiomyxoma (SAM) is a rare benign soft-tissue tumor that usually occurs in the trunk, head and neck, and lower extremity of middle-aged adults. Herein, we describe an unusual case of SAM of the wrist, which was initially diagnosed as a ganglion cyst on imaging. CASE REPORT: The patient was a 71-year-old man with no history of trauma who presented with a 2-year history of a palpable mass in the left wrist. Physical examination revealed a 2.5-cm, elastic hard, mobile, nontender mass. Magnetic resonance imaging revealed a well-defined mass with iso-signal intensity relative to skeletal muscle on T1-weighted sequences and very high signal intensity on T2-weighted fat-suppressed sequences. Subtle internal enhancement was seen following gadolinium administration. Complete excision was performed under general anesthesia with tourniquet control. Histologically, the lesion was composed of bland spindle to stellate-shaped cells in an abundant myxoid stroma. Immunohistochemically, the lesional cells were positive for CD34 but negative for S-100 protein, smooth-muscle actin, desmin, epithelial membrane antigen and pancytokeratin. These findings were consistent with a diagnosis of SAM. There was no clinical evidence of recurrence during a follow-up period of 3 months. CONCLUSION: Although extremely rare, SAM should be considered in the differential diagnosis of a cyst-like solid lesion near small joints.

Ectomesenchymal chondromyxoid tumor: histopathologic and immunohistochemical study of two cases without a chondroid component.

Ectomesenchymal chondromyxoid tumor (ECT) is a rare benign neoplasm usually affecting the anterior dorsum of the tongue. Histopathologically, it is formed by spindle, round and/or polygonal cells embedded in a chondromyxoid matrix. Immunohistochemical positivity for vimentin, S-100 protein, glial fibrillary acid protein and neuron-specific enolase are helpful to confirm the diagnosis. There are 42 cases of ECT of the tongue reported in the English language literature, three of them showing no chondroid matrix. We describe two additional cases of ECT lacking the chondroid component, exhibiting areas of reticulated myxoid and cellular pattern. Considering the microscopical features, ECT can be classified in classic and 'chondroid-free' variants, the latter including the reticulated myxoid and cellular patterns. It is important to consider that the cellular ECT usually exhibits predominance of an infiltrative atypical cellular component that may mimic a malignant tumor.

Myxoid variant of anaplastic large cell lymphoma involving the skin: a case report.

We report the case of a 62-year-old white male who presented with a 2.6-cm ulcerating mass on the skin of the left buttock and ipsilateral inguinal lymphadenopathy. Microscopic sections of the skin lesion showed a nodular and plaque-like growth pattern of a mixed cellular infiltrate throughout the dermis and subcutaneous tissue with prominent myxoid change. There was a dominant population of medium-sized mitotically active atypical cells that expressed CD30, CD4 and EMA. These atypical cells were mixed with eosinophils, neutrophils, mature lymphocytes and histiocytes. Tissue from the inguinal lymphadenopathy showed similar pathologic features, although no residual lymph node tissue was present. A diagnosis of secondary anaplastic large cell lymphoma, myxoid variant, with skin and lymph node/perinodal soft tissue involvement was rendered at the time of complete excision of the buttock mass. The patient received five cycles of CHOP (cyclophosphamide, doxorubicin, vincristine and prednisone) chemotherapy with complete resolution of lymphadenopathy and no residual cutaneous disease. He was disease-free by PET/CT scan and physical examination at 16 months after chemotherapy. We present this case to highlight the histopathologic and immunophenotypic features of this entity with a discussion of the differential diagnosis and a review of the literature.