GNAQ Q209R Mutations Are Highly Specific for Circumscribed Choroidal Hemangioma.

Several tumors, including uveal melanoma, show somatic mutations of GNAQ/GNA11. Circumscribed choroidal hemangioma is a benign tumor that becomes symptomatic in adulthood. In some patients, morphologic examination of biopsies is required for differential diagnosis between amelanotic choroidal melanoma and circumscribed choroidal hemangioma. Here, we report the results of GNAQ/GNA11 mutation analysis in samples from circumscribed choroidal hemangioma. Deep amplicon sequencing (Illumina MiSeq, San Diego, CA, USA) of positions R183 and Q209 of GNAQ and GNA11 in tissue samples from 33 patients with histologically diagnosed circumscribed choroidal hemangioma. All patients underwent biopsy or enucleation at our clinic between 2008 and 2018. To enable detection of variant alleles at low fractions, read depth exceeded 15,000-fold. DNA for genetic analysis was prepared from either snap-frozen (n = 22) or FFPE (n = 11) tissue samples. Samples from 28/33 patients (85%) showed a somatic missense mutation of GNAQ (c.626 A > G) predicted to result in p.Q209R. Variant allele fraction was variable (range 2.3% to 28%). Variants of GNAQ resulting in p.Q209 are characteristic for circumscribed choroidal hemangiomas. It appears that the GNAQ mutation spectrum in this tumor is narrow, possibly restricted to p.Q209R. Moreover, the spectrum is distinct from that of uveal melanoma, in which alterations resulting in p.Q209R are very rare.

Chromosome 3 is a valid marker for prognostic testing of biopsy material from uveal melanoma later treated by brachytherapy.

PURPOSE: Monosomy 3 (M3) in uveal melanoma (UM) obtained after enucleation is significantly associated with metastatic death. With improved biopsy techniques, samples from patients treated with eye-preserving methods have become available. As the choice of treatment depends on tumour size, patients treated with eye-preserving brachytherapy tend to have smaller tumours. It has to be determined if M3 is a valid marker for prognosis of these patients. METHODS: Follow-up and clinical data were collected from a total of 451 UM patients: 291 patients were treated by brachytherapy. Tumour tissue was sampled by transretinal biopsy using the 23-gauge Essen biopsy forceps prior to therapy in 114 of them. Chromosome 3 status was determined by microsatellite analysis. Data were compared to those from 160 patients treated by enucleation. RESULTS: Chromosome 3 status correlates significantly with disease-related survival in both patient groups. The proportion of tumours with M3 is lower in the brachytherapy group compared to patients treated with enucleation (25/77 32% and 102/144 71%, respectively). CONCLUSIONS: M3 is a valid marker for poor prognosis in uveal melanoma later treated by brachytherapy. The higher proportion of D3 tumours might explain, at least in part, the more favourable prognosis of patients treated by brachytherapy.

Immunohistochemical evidence of BMP-2, -4 and -7 activity in otospongiosis.

CONCLUSION: This study is the first to show that bone morphogenetic proteins (BMP)-2, -4 and -7 play a role in active phase otosclerotic bone remodelling (otospongiosis). OBJECTIVES: The role of BMPs in various tissue growth and repair mechanisms is an ongoing topic in the literature. BMP-2, -4 and -7 are known to be of major importance in bone formation and repair. Their role in otosclerotic bone transformation has not been analysed previously. The main goal of this study was to perform an immunohistological analysis of BMP-2, -4 and -7 in otoclerosis. MATERIALS AND METHODS: Parts of the stapedial footplates, collected during partial stapedectomies in 30 patients with clinical otosclerosis, were analysed for histological otosclerotic lesions after staining haematoxylin and eosin. Immunohistochemical analysis was performed using polyclonal IgG antibodies for BMP-2, -4 and -7, as well as biotinylated secondary antibodies, avidin-biotin-peroxidase complex reaction and alkaline phosphatase staining. RESULTS: In all, 14 specimens contained otosclerosis; 3 of these were otospongiotic, 8 fibrotic, 2 sclerotic and 1 had both sclerotic and fibrotic lesions. Thus in total 14/30 specimens (47%) showed histological otosclerosis. Only the multiple osteoblasts and osteoclasts in those specimens exhibiting an otospongiotic phase showed distinct immunochemical staining for BMP-2, -4 and -7.

Signaling by way of type IB and II bone morphogenetic protein receptors regulates bone formation in otospongiosis.

HYPOTHESIS: The main goal of this study was to perform an immunohistologic analysis of bone morphogenetic protein receptors (BMPR) in otospongiosis. BACKGROUND: BMP-2, -4, and -7 play an essential role in bone formation and repair. They do so as well in otosclerosis. It has been shown that these BMPs are traceable in osteocytes and osteoclasts in the active phase of otosclerosis (otospongiosis). The role of the different BMP receptors in otosclerotic bone transformation has not been previously analyzed. METHODS: The posterior parts of the stapes footplates, collected during partial stapedectomies in 35 patients with clinical otosclerosis, were analyzed for histologic otosclerotic lesions after hematoxylin staining. Immunohistochemical analysis was performed using polyclonal immunoglobulin G antibodies for BMPR-IA, -IB, and -II, as well as biotinylated secondary antibodies, avidin-biotin-peroxidase complex reaction, and alkaline phosphatase staining with nitroblue-tetrazolium-chloride. RESULTS: Seventeen of 35 (49%) specimens contained otosclerosis, but only 5 of these exhibited an otospongiotic phase. The abundant osteoblasts and osteoclasts in these cases showed distinct immunochemical staining for BMP-2, -4, and -7. In two cases, there could also be found an immense positive staining for BMPR-IB and modest staining for BMPR-II, whereas BMPR-1A always remained negative. CONCLUSION: It was demonstrated for the first time that in otospongiosis, the actions of the BMPs are mediated through BMPR-IB and BMPR-II. To determine this role in detail, further investigations, especially for the phosphorylated Smad proteins within the BMP dependent mediator cascade, will be necessary.

Mixed chimerism of cardiomyocytes and vessels after allogeneic bone marrow and stem-cell transplantation in comparison with cardiac allografts.

Controversy persists about mixed chimerism (mCh) occurring in the hearts of patients after orthotopic cardiac transplantation in comparison with allogeneic bone marrow (BM) and peripheral blood stem-cell (PBSC) transplants. Cadaver hearts were examined after sex-mismatched transplantation by immunophenotyping combined with dual color fluorescence in situ hybridization (X and Y chromosome-specific probes). A striking disparity in the extent of mCh depending on the different transplantation procedures was recognizable. After allografting with PBSCs, 1.7% chimeric cardiomyocytes were detectable contrasting 5.4% of donor cells after full BM transplantation. In cardiac transplants, host-type endothelial cells (16.2%) and myocytes (14.3%) of the vessel walls were more often encountered than after BM and PBSC allografting. A sprouting of vascular structures into the donor heart after orthotopic cardiac transplantation has to be assumed, as does a pivotal role of the mesenchymal stem cells of the BM in the development of mCh.