[Expression of VHL, VEGF and HIF-1α in endolymphatic sac tumors].

Objective: To investigate the expression of von Hippel-Lindau (VHL), vascular endothelial growth factor (VEGF) and hypoxia inducible factor-1α (HIF-1α) in endolymphatic sac tumor (ELST) and its clinical significance, and to analyze its association with VHL gene mutation. Methods: Twenty-four cases of ELST, which were surgically resected and diagnosed by pathological examination in Beijing Tongren Hospital Affiliated to Capital Medical University, Beijing, China during 2012-2020, were recruited as the ELST group, and 24 cases of otitis media diagnosed in the same hospital were selected as the control group. The expression of VHL, VEGF, and HIF-1α was assessed using EnVision immunohistochemical staining and compared between the ELST and control groups. Sanger sequencing was performed to detect the VHL mutation status in 24 ELSTs. The correlations among VHL, VEGF and HIF-1α expression were analyzed. The associations of VHL, VEGF and HIF-1α expression with age of onset, gender, tumor size, bone invasion and clinical stage in ELST were also analyzed. Results: The expression rate of VHL in the ELST group was significantly lower than that in the control group (P<0.05), but the expression rates of VEGF and HIF-1α in the ELST group were significantly higher than those in the control group (P<0.05). VHL expression was inversely correlated with VEGF and HIF-1α expression. The expression of VEGF and HIF-1α was associated with bone invasion and clinical stage (P<0.05), but the expression of VHL, VEGF and HIF-1α had no significant associations with the age of onset, gender, or tumor size of ELST (P>0.05). Conclusions: The expression of VHL is decreased while that of VEGF and HIF-1α increased in ELST. Expression of VHL is inversely correlated with that of VEGF and HIF-1α. The expression of VEGF and HIF-1α is correlated with bone invasion and clinical stage. Thus, VEGF and HIF-1α may be therapeutic targets of ELST.

MicroRNA-370 suppresses SOX12 transcription and acts as a tumor suppressor in bladder cancer.

OBJECTIVE: Dysregulation of microRNA-370 (miR-370) is involved in a variety of cancers, but its roles in bladder cancer (BC) remain largely unexplored. Therefore, we designed this study to explore the role of miR-370 in BC. PATIENTS AND METHODS: We took advantage of biochemical assays, including RT-qPCR, Western blot, CCK-8, flow cytometry, transwell, xenograft tumor formation, and immunohistochemistry (IHC) for research. RESULTS: The expression of miR-370 was found to be downregulated during the development of BC, highly correlating with the malignant transformation of tumors. The overexpression of miR-370 led to enhanced apoptosis in BC cells, while inhibiting cell proliferation, migration, and invasion, effectively blocking cancer metastasis. Additionally, we identified SOX12, a known human oncogene, as a direct target of miR-370, showing that upregulation of SOX12 attenuated miR-370-mediated tumor suppression, promoted tumor growth, and epithelial-mesenchymal transition (EMT) in BC. CONCLUSIONS: Taken together, these findings help to elucidate the roles of miR-370 as a tumor suppressor in BC, providing a potential target for diagnosis and treatment of BC.

Chemical composition, energy, and amino acid digestibility in 7 cottonseed co-products fed to growing pigs.

The objective of this study was to determine the chemical composition, DE and ME contents, and apparent ileal digestibility (AID) and standardized ileal digestibility (SID) of AA in 7 cottonseed co-products fed to growing pigs. The 7 cottonseed co-products were: cottonseed meals (solvent extracted) with CP level of 46%, 50%, and 55% (46CSM, 50CSM, and 55CSM), cottonseed protein with CP level of 50% and 55% (50CSP and 55CSP), fermented cottonseed meal (CSMF), and expelled cottonseed meal (CSME). The DE and ME contents of the 7 test ingredients were measured using 48 crossbreed barrows (BW: 44.1 ± 4.2 kg) and 8 diets (including a corn-soybean meal basal diet) in a completely randomized design. The AID and SID of AA of the 7 test ingredients were determined using 7 crossbreed barrows (initial BW: 67.4 ± 8.5 kg) in a 6 × 8 Youden square design with 6 periods and 8 diets (including one N-free diet). The DE and ME values of the 7 cottonseed co-products varied from 12.72 to 15.63 MJ/kg and 12.24 to 14.83 MJ/kg, respectively. Among the 7 cottonseed co-products, 55CSP and CSME had the greatest (P < 0.05) ME and DE values compared to the other cottonseed co-products. In addition, 55CSP had the greatest (P < 0.05) AID and SID of all the AA tested except for Gly and Pro. In contrast, 46CSM or 55CSM had the lowest (P < 0.05) AID and SID of all the AA tested except for Gly and Pro. The 55CSP also had the greatest (P < 0.05) concentrations of standardized ileal digestible CP and all the AA tested except for Gly and Pro. In conclusion, the chemical composition, energy, and AA digestibility of cottonseed co-products with different processing techniques varied widely. Based on the energy and AA digestibility, cottonseed protein with CP level of 55% is a better dietary ingredient for growing pigs compared with the other cottonseed co-products.

[Role of ash2 (absent, small, or homeotic)-like and Jumonji domain-containing protein 3 on histone methylation of interferon-gamma gene and their associations with vascular damage of Kawasaki disease].

Objective: To investigate the impacts of ash2 (absent, small, or homeotic)-like (Ash2L) and Jumonji domain-containing protein 3 (Jmjd3) on histone methylation of interferon-gamma(IFN-γ) gene and association with vascular damage of Kawasaki disease (KD) in acute phase. Methods: This study was performed among 36 children with KD in acute phase (KD group) and 28 age-matched health children (control group), who were treated or underwent physical examination in our hospital between February 2015 and June 2016. Patients were further divided into KD groups with or without coronary artery lesions (KD-CAL(+) , 16 cases; KD-CAL(-), 20 cases). All KD patients were treated with intravenous immunoglobulin. The proportion of type 1 helper T(Th1) cells and protein levels of IFN-γ, T-box expressed in T cells(T-bet), phosphorylated signal transducer and activator of transcription 1(pSTAT1) and phosphorylated signal transducer and activator of transcription 4(pSTAT4) were analyzed by flow cytometry.Chromatin immunoprecipitation was performed to determine histone methylation (histone H3 tri-methyl K4(H3K4me3), histone H3 tri-methyl K27(H3K27me3)) and binding levels of Ash2L, Jmjd3 and Ezh2 associated with IFN-γ in CD4(+) T cells. Quantitative real-time PCR was used to determine mRNA levels of IFN-γ, interferon γ receptor 1(IFN-γR1), interferon γ receptor 2(IFN-γR2), interleukin 12 receptor subunit beta 1(IL-12Rß1), interleukin 12 receptor subunit beta 2(IL-12Rß2), interleukin 18 receptor subunit beta α(IL-18Rα), interleukin 18 receptor subunit beta ß(IL-18Rß), tumor necrosis factor receptor 1(TNFR1), toll-like receptor 4(TLR4), receptor interacting serine/threonine kinase 1(RIP-1) and myeloid differentiation primary response gene 88(MyD88) in CD4(+) T cells. Plasma concentrations of IFN-γ, interleukin 12(IL-12), interleukin 18(IL-18) and tumor necrosis factor α(TNF-α) were measured by enzyme-linked Immunosorbent assay. Results: (1)The proportion of Th1 and its protein level of IFN-γ were significantly higher in KD group than those in control group and higher in KD-CAL(+) group than in KD-CAL(-) group (all P<0.05), and lower after treatment than before treatment (all P<0.05). (2)Compared with control group, mRNA level of IFN-γ and IFN-γ-associating H3K4me3 was increased, while level of IFN-γ associating H3K27me3 in CD4(+) T cells was reduced in KD group (all P<0.05), which resulted in a higher rate of H3K4me3/H3K27me3 (P<0.05) in KD group, which was positively correlated with IFN-γ mRNA in KD group(r=0.55, P<0.05). Similar results were found between KD-CAL(+) group and KD-CAL(-) group (all P<0.05). Level of IFN-γ associating H3K27me3 was increased, and mRNA level of IFN-γ and IFN-γ associating H3K4me3 was decreased after treatment than before treatment (all P<0.05). (3)Expression of T-bet protein and binding levels of Ash2L and Jmjd3 with IFN-γ gene were significantly higher in KD group than those in control group(all P<0.05), higher in KD-CAL(+) group than those in KD-CAL(-) group (all P<0.05). These parameters were significantly lower after treatment than before treatment (all P<0.05). Binding level of Ezh2 with IFN-γ gene was similar among various groups (all P>0.05). (4)In comparison with control or after treatment, surface receptors(IFN-γR1/2, IL-12Rß1/2, IL-18Rα/ß, TNFR1 and TLR4) and its downstream molecules(pSTAT1, pSTAT4, RIP(1) and MyD88) in CD4(+) T cells, and plasma concentrations of inflammatory cytokines(IFN-γ, IL-12, IL-18 and TNF-α) were found to be higher in KD group(all P<0.05). These parameters were also higher in KD-CAL(+) group than in KD-CAL(-) group (all P<0.05). Conclusion: Aberrant histone methylation of IFN-γ associating H3K4me3 and H3K27me3 caused by over-binding of Ash2L and Jmjd3 might be involved in immune dysfunction and vascular damage in KD in the acute phase.

Dynamic relationship between SIPA1 gene and protein expression and the development of gastric cancer.

Association of signal-induced proliferation-associated 1 (SIPA) gene and protein expression with gastric cancer development was examined. SIPA1 mRNA and protein levels were determined by real-time quantitative polymerase chain reaction and western blot, respectively, in 40 gastric tumor and tumor-adjacent normal tissues. SIPA1, VEGF-A, and FVIII levels in 60 gastric tumor and 40 tumor-adjacent normal tissues were examined by immunohistochemical staining. Correlations between SIPA1, VEGF-A, and microvessel density (MVD) were analyzed. SIPA1 mRNA levels were significantly lower in tumor tissues than in tumor-adjacent normal tissues (P < 0.05). Similarly, protein levels were significantly lower in tumor tissues (0.3043 ± 0.1062) than in tumor-adjacent normal tissues (0.5423 ± 0.0682, P < 0.05). Positive staining rates of SIPA1 (48.3%) and VEGF-A (36.7%) were lower and higher, respectively, in tumor tissues than in tumor-adjacent normal tissues (65.0 and 2.5%, P < 0.05). Positive protein staining rates in tumor tissues correlated with the degree of differentiation, lymph node metastases, and clinical grading (P < 0.05) and not with sex, age, or tumor size (P > 0.05). Significantly higher MVD (57.4 ± 9.3) was observed in tumor tissues displaying positive SIPA1 staining than in tumor-adjacent normal tissues (41.2 ± 5.7, P < 0.05). SIPA1 and VEGF-A expression in tumor tissues were negatively correlated (r = -0.736, P < 0.05). SIPA1 and its protein may play important roles in gastric cancer invasion, metastasis, and biological behavior. Low SIPA1 levels in gastric cancer may accelerate tumor development and progression by promoting VEGF-A expression to increase vascular density.

[Clinicopathological features of acantholytic mammary Paget's disease: a report of 28 cases].

Objective: To investigate the clinicopathological features of acantholytic mammary Paget's disease (AMPD). Methods: From January, 2010 to October, 2016, a total of 28 patients were diagnosed as AMPD in the Department of Dermatology, Peking Union Medical College Hospital. The clinical and histopathological data of these patients were analyzed retrospectively. Results: The patients were all female. The mean age of onset was (51±15)years (range, 24 to 78 years). The median course of disease was 10.5 months (range, 3 months to 2 years). All the cases were unilaterally involved. The ratio of left breast involvement to the right breast involvement was 1.55∶1. In histopathological examination, all the cases showed acantholytic pattern. In addition to that, the prototypical pattern accounted for 50.0% (14/28) of all the AMPD cases. The frequencies of other different accompanied histopathological patterns were: 3 (10.7%) upper nest, 2 (7.1%) budding, 1 (3.6%) tall nest, and 1 (3.6%) sheet-like. Hailey-Hailey-disease-like acantholysis was observed in 18 patients (64.3%), whereas pemphigus-vulgaris-like acantholysis was noted in 7 patients (25.0%) and Darier's-disease-like acantholysis in 3 patients (10.7%). About 75.0% (21/28) of the AMPD cases were found to be accompanied with underlying breast cancers. There was no recurrence in 18 of 20 patients who completed treatment and were followed up for over 1 year. Conclusions: AMPD might involve the left breast more frequently. It is mostly associated with the prototypical pattern of mammary Paget's disease. Hailey-Hailey-disease-like acantholysis may be the most frequent subtype of AMPD. Most AMPD cases are associated with underlying breast cancers, but with a low recurrence rate after treatment.

[Clinicopathological study of 56 cases of extramammary Paget's disease with or without acantholysis].

Objective: To identify the clinicopathological features of extramammary Paget's disease(EMPD) and investigate the clinical and histopathological significance of acantholysis in EMPD. Methods: From June, 2010 to October, 2016, a total of 56 patients were diagnosed as EMPD in the Department of Dermatology, Peking Union Medical College Hospital. Clinical and histopathological data were retrieved from these patients' medical records and analyzed respectively. The cases were divided into two subgroups according to the histopathological pattern (with or without acantholysis): the acantholytic EMPD (AEMPD) group and the non-acantholytic EMPD (N-AEMPD) group. The clinicopathological data were compared and statistically analyzed between the two groups. Results: The AEMPD group included 22 cases (39.3%), while the N-AEMPD group included 34 cases (60.7%). The male: female ratio, the age of onset, and the median duration of the disease were 10∶1 vs 10.3∶1, (64±8)vs (64±10)years, and 42(4-240)months vs 48(3-120) months in the AEMPD and N-AEMPD groups, respectively, with no significant difference (all P>0.05). There was no significant difference in the severe dermal inflammation (27.3% vs 17.6%, P=0.600 ) and cytologic anaplasia(13.6% vs17.6%, P=0.979)between the AEMPD and N-AEMPD groups.Adnexal involvement or dermal invasion (72.7%) was significantly more common in cases with AEMPD, compared to cases with N-AEMPD (23.5%, P<0.001). And 17 cases in the AEMPD group (77.3%) were Hailey-Hailey-disease-like subtype. The recurrence rate after surgical management (29 cases) showed no significant difference between the two groups (1/12 vs 4/17, P=0.370). Conclusions: Acantholysis is common in EMPD. It is not associated with sex, the age of onset, and the duration of the disease. Acantholysis may indicate invasive growth of Paget's cells. Its occurrence has no association with severe dermal inflammation or cytologic anaplasia. Hailey-Hailey-disease-like subtype is the most common subtype in AEMPD, requiring careful consideration to avoid misdiagnosis. Postoperative recurrence is not associated with acantholysis in EMPD.

Genomewide Association Study Identifies Novel Genetic Loci That Modify Antiplatelet Effects and Pharmacokinetics of Clopidogrel.

Genetic variants in the pharmacokinetic (PK) mechanism are the main underlying factors affecting the antiplatelet response to clopidogrel. Using a genomewide association study (GWAS) to identify new genetic loci that modify antiplatelet effects in Chinese patients with coronary heart disease, we identified novel variants in two transporter genes (SLC14A2 rs12456693, ATP-binding cassette [ABC]A1 rs2487032) and in N6AMT1 (rs2254638) associated with P2Y12 reaction unit (PRU) and plasma active metabolite (H4) concentration. These new variants dramatically improved the predictability of PRU variability to 37.7%. The associations between these loci and PK parameters of clopidogrel and H4 were observed in additional patients, and its function on the activation of clopidogrel was validated in liver S9 fractions (P < 0.05). Rs2254638 was further identified to exert a marginal risk effect for major adverse cardiac events in an independent cohort. In conclusion, new genetic variants were systematically identified as risk factors for the reduced efficacy of clopidogrel treatment.