A Tumor-Specific Neo-Antigen Caused by a Frameshift Mutation in BAP1 Is a Potential Personalized Biomarker in Malignant Peritoneal Mesothelioma.

Malignant peritoneal mesothelioma (MPM) is an aggressive rare malignancy associated with asbestos exposure. A better understanding of the molecular pathogenesis of MPM will help develop a targeted therapy strategy. Oncogene targeted depth sequencing was performed on a tumor sample and paired peripheral blood DNA from a patient with malignant mesothelioma of the peritoneum. Four somatic base-substitutions in NOTCH2, NSD1, PDE4DIP, and ATP10B and 1 insert frameshift mutation in BAP1 were validated by the Sanger method at the transcriptional level. A 13-amino acids neo-peptide of the truncated Bap1 protein, which was produced as a result of this novel frameshift mutation, was predicted to be presented by this patient's HLA-B protein. The polyclonal antibody of the synthesized 13-mer neo-peptide was produced in rabbits. Western blotting results showed a good antibody-neoantigen specificity, and Immunohistochemistry (IHC) staining with the antibody of the neo-peptide clearly differentiated neoplastic cells from normal cells. A search of the Catalogue of Somatic Mutations in Cancer (COSMIC) database also revealed that 53.2% of mutations in BAP1 were frameshift indels with neo-peptide formation. An identified tumor-specific neo-antigen could be the potential molecular biomarker for personalized diagnosis to precisely subtype rare malignancies such as MPM.

[The effects of equol on enzymes related to testosterone synthesis and vimentin in the testis of perinatal mice by organ culture in vitro].

OBJECTIVE: To investigate the effects of Equol on genes and protein expression of testosterone synthesis related enzymes and Vimentin in testis of perinatal mice in vitro. METHODS: Testes were isolated and cultured in infiltrating type rotating device for 72 h. The testes were randomly divided into five groups and treated with Equol (DMSO control, 0.01, 0.10, 1.00, 10.00 µmol/L Equol). Morphological changes were observed by HE staining under optical microscope. Expressions of 3ß-hydroxysteroid dehydrogenase (3ß-HSD),P450 side-chain cleavage enzyme (P450scc), Vimentin were detected by real-time PCR and immunohistochemistry. RESULTS: No apparent morphological change in testes was observed compared to control group. The mRNA expression of 3ß HSD, P450Scc, Vimentin show no statistical significance(P> 0. 05) in all Equol group, while the protein expressions of 3ß-HSD, Vimentin, P450scc increased in 0. 10 µmol/L and decreased in 10.00 µmol/L Equol group. CONCLUSION: Equol exposure can affect 3ß-HSD, P450Scc, Vimentin expression in testes in vitro, means Equol may have potential adverse effects on testosterone production and spermatogenesis.

[The effects of di-(2-ethylhexyl) phthalate (DEHP) in testosterone synthesis and its molecular mechanisms in the fetal testis of male mouse by organ culture in vitro].

OBJECTIVE: To investigate the effects of DEHP in testosterone synthesis and the related genes expression in the fetal testis of male mouse by organ culture in vitro. METHODS: The testis tissues were cultured in infiltrating type rotating device for 72 hours. The culture and gas were changed every 24 hours. The testis tissues were divided into the DMSO control group and four DEHP groups (the terminal concentration were 0.1, 1.0, 10.0, 100.0 micromol/L). The testosterone levels in the cultured medium were measured by radioimmunoassay, the INHBbeta levels were measured by Elisa, the gene expressions related with testosterone synthesis were detected by real-time PCR, the morphological changes of cultured testis were observed by HE staining under optical microscope, the expressions of related proteins were measured by immunohistochemistry. RESULTS: Compared to the control group, the levels of testosterone synthesis in 0.1, 1.0, 10.0 micromol/L groups were increased, but decreased in 100.0 micromol/ L group which were exposed in DEHP for 48 hours and 72 hours. There was an increase of INHBbeta synthesis in 0.1 and 1.0 micromol/L groups, but a decrease in 10.0 and 100.0 micromol/L groups. The gene expressions of 3beta-HSD, P450c17, P450Scc, vimentin were significantly decreased compared to that of control group especially in 1.0 and 10.0 micromol/L groups (P < 0.05), but the expressions of INHBbeta had no significant changes. There were no apparent morphological changes in testis tissue by HE staining. A significant increase of the three proteins (3beta-HSD, P450c17, P450Scc) and a significant decrease of vimentin were observed in 10.0 and 100.0 micromol/L groups (P < 0.05). CONCLUSION: DEHP exposure can affect the testosterone synthesis in the fetal testis of male mouse. The regulation of gene expression involving in testosterone synthesis might be the mechanism.

MRI and the pathology of breast invasive micropapillary carcinoma.

Diagnosis of breast invasive micropapillary carcinoma (IMPC) before surgery is of great value for determining the optimal treatment strategy. The aim of the present study was to investigate the magnetic resonance imaging (MRI) and pathological features of IMPC. MRI features of IMPC were characterized in relation to the patients' clinicopathological features. Clinical manifestations, mammography results and/or MRI findings of patients with IMPC were retrospectively analyzed. Parameters included morphology, plain T2-weighted imaging (T2WI) signal intensity, the apparent diffusion coefficient (ADC), the internal enhancement mode, early enhancement rates and time-intensity curve (TIC) types during dynamic enhanced scanning. A total of 10 lesions were detected by MRI in eight patients, with one case having three lesions with the mean diameter of 34.44 mm. In plain T2WI scanning, the lesions appeared inhomogeneous with a moderate or high signal intensity. When the b value was 800 sec/mm2, the average ADC value was 0.823±0.12×10-3 mm2/sec. A total of four cases exhibited mass-like enhancement, including an oval rim in one case (three lesions), irregular inhomogeneous enhancement in two cases and irregular uniform enhancement in one case. The margins were clear in one case (three lesions), irregular in two cases and spiculate in one case. Among the four cases with non-mass enhancement, the distribution was focal in two cases, linear in one case and regional in one case, and the internal enhancement mode was cluster-like in one case, heterogeneous in one case and uniform in two cases. The average early enhancement rate was 116.96±45.26%. TICs of type III were observed in all cases. In conclusion, MRI of IMPC demonstrated typical features of malignant tumors and lymphatic vessel infiltration, suggesting that MRI may exhibit guiding significance for the diagnosis and treatment planning of IMPC.

[Expressions of MMP-2 and COX-2 mRNA in bladder transitional cell carcinoma and their correlation].

OBJECTIVE: To determine the levels of MMP-2 and COX-2 mRNA in bladder transitional cell carcinoma tissues and explore their relationship. METHODS: We enrolled in this study 42 patients with bladder transitional cell carcinoma, including Ta-T1 (n = 18), T2-T4 (n = 24), G1 (n = 12), G2 (n = 19), G3 (n = 11), metastasis (n =26) and non-metastasis (n = 16). Another 5 cases of normal bladder tissues were taken as controls, and the levels of MMP-2 and COX-2 mRNA were detected by RT-PCR. RESULTS: The relative expressions of COX-2 mRNA were 1.038 +/- 0. 484 in Ta-T1, 1.489 +/- 0.584 in T2-T4, 0.920 +/- 0.442 in G1, 1.338 +/- 0.584 in G2 and 1.632 +/- 0.515 in G3, all significantly higher than that of the controls (0.460 +/- 0.224, P < 0.05). And the corresponding relative levels of MMP-2 mRNA were 1.107 +/- 0.384, 1.604 +/- 0.425, 0.971 +/- 0.370, 1.445 +/- 0.378 and 1.755 +/- 0.387, also significantly higher than that of the latter group (0.423 +/- 0.227, P < 0.05). The COX-2 and MMP-2 mRNA levels in the tumor tissues with and without metastasis were 1.591 +/- 0.455 vs 0.815 +/- 0.430 and 1.676 +/- 0.339 vs 0.927 +/- 0.228, (P < 0.01), respectively, with a positive correlation between the mRNA level of COX-2 and that of MMP-2 (r = 0. 703, P < 0.01). CONCLUSION: MMP-2 and COX-2 mRNA are highly expressed in bladder transitional cell carcinoma tissues and their expressions are positively correlated with the degree of malignancy. MMP-2 and COX-2 might play a synergetic role in the pathogenesis and progression of bladder transitional cell carcinoma.

[Establishment of a rotary aerobic culture system for in vitro culture of mouse testis].

OBJECTIVE: To establish an in vitro model of cultured mouse testis using rotary aerobic culture. METHODS: Rotary aerobic incubation with optimized culture conditions was used for in vitro culture of mouse testis, and the morphology of the cultured testicular tissues was compared with that cultured in Transwell chambers. The changes in the testicular tissue structure were examined using HE staining, and the cell proliferation was assessed with BrdU staining. Testosterone concentrations in the culture medium were tested with radioimmunoassay and the expression of the functionally related proteins in the testis was detected using immunohistochemistry. RESULTS: The testicular tissue cultured by optimized rotary aerobic culture presented with more intact histological structure with the size of the testis ranged from 0.3 to 0.8 mm(3). In the two culture systems, the prolifeation index of the spermatogonia increased and that of Sertoli cells decreased with time, and such changes in spermatogonia and Sertoli cell proliferation indices became statistically significant at 3 days (P<0.05) and 5 days (P<0.05) of culture, respectively, as compared with those at 1 day. The concentration of testoerone in the culture media decreased significantly with incubation time (P<0.05). At 3 days of culture, the protein expression of 3ß-hydroxysteroid dehydrogenase, cytochrome P450 17α-hydroxylase and cholesterol side-chain cleavage enzyme was detected in Leydig cell cytoplasm and vimentin expression in Sertoli cell cytoplasm. CONCLUSION: An in vitro model of cultured mouse testis has been successfully established using rotary aerobic incubation.

STR DNA genotyping of hydatidiform moles in South China.

OBJECTIVE: To evacuate whether short-tandem-repeat (STR) DNA genotyping is effective for diagnostic measure to precisely classify hydatidiform moles. METHODS: 150 cases were selected based on histologic features that were previously diagnosed or suspected molar pregnancy. All sections were stained with hematoxylin as a quality control method, and guided the microscopic dissection. DNA was extracted from dissected chorionic villi and paired maternal endometrial FFPE tissue sections. Then, STR DNA genotyping was performed by AmpFlSTR(®) Sinofiler(TM) PCR Amplification system (Applied Biosystems, Inc). Data collection and analysis were carried out using GeneMapper(®) ID-X version 1.2 (Applied Biosystems, Inc). RESULTS: DNA genotyping was informative in all cases, leading to identification of 129 cases with abnormal genotype, including 95 complete and 34 partial moles, except 4 cases failed in PCR. Among 95 complete moles, 92 cases were monospermic and three were dispermic. Among 34 partial moles, 32 were dispermic and 2 were monospermic. The remaining 17 cases were balanced biallelic gestations. CONCLUSION: STR DNA genotyping is effective for diagnostic measure to precisely classify hydatidiform moles. And in the absence of laser capture microdissection (LCM), hematoxylin staining plus manual dissection under microscopic guided is a more economic and practical method.