Effects of benzo(a)pyrene on expressions of insulin-degrading enzyme and neprilysin in neuroglia cells / 北京大学学报(医学版)

OBJECTIVE@#To investigate effects of benzo(a)pyrene (BaP) on expressions of insulin-degrading enzyme (IDE) and neprilysin (NEP) which have the ability to degrade β-amyloid (Aβ) in neuroglia cells.@*METHODS@#Primary mix-neuroglia cells were cultured from newborn SD rats. After exposure to BaP, Aβ1-42 oligomer or Aβ1-42 fiber individually or jointly for 24 h, the cell survival rate was measured by cell counting kit-8 (CCK-8). Afterwards, the primary mix-neuroglia cells were divided randomly into six groups: Control group, BaP group (2.00 μmol/L), Aβ1-42 oligomer group (20.00 mg/L), BaP plus Aβ1-42 oligomer group, Aβ1-42 fiber group (20.00 mg/L) and BaP plus Aβ1-42 fiber group, of which BaP was pretreated for 12 h followed by cotreatment with different aggregated Aβ1-42. The expressions of IDE and NEP were measured by quantitative real-time polymerase chain reaction (qRT-PCR) for mRNA level and Western blotting for protein level.@*RESULTS@#The cell survival rate showed no significant differences after treatment with BaP (≤20.00 μmol/L), Aβ1-42 oligomer (20.00, 40.00 mg/L), Aβ1-42 fiber (20.00, 40.00 mg/L) or cotreatment with BaP and Aβ1-42 oligomer or BaP and Aβ1-42 fiber. Compared with the control group, expressions of IDE and NEP in BaP-treated alone group had no obvious change; however, exposure to Aβ1-42 oligomer alone significantly increased the mRNA and protein level of IDE (P<0.05), and the BaP pretreatment could significantly inhibit the up-regulated expressions of IDE by Aβ1-42 oligomer (P<0.05); on the other hand, exposure either to Aβ1-42 fiber alone or under the BaP pretreatment did not change the mRNA and protein level of IDE and NEP obviously.@*CONCLUSION@#On the premise of no significant change of cell survival rate, BaP pretreatment inhibited the up-regulated expressions of IDE in primary mixed neuroglia cells under cotreatment with Aβ oligomer, indicating that BaP may disturb degradation of Aβ oligomer and cause deposition of β-amyloid and further induce cognitive decline and acceleration of Alzheimer.

Colonic Endometriosis / 대한소화기학회지

No abstract available.

Effect of neoadjuvant chemotherapy on stromal CD10 antigens in breast cancer - A preliminary study.

Introduction: CD10 is a zinc-dependent peptidase (metalloproteinase). Stromal CD10 expression in breast cancer correlates with poor prognosis, oestrogen receptor negativity and higher grade. CD10 may be a potential target of new cancer therapies as it is involved in cleavage of doxorubicin. Aim: To evaluate the effect of neo-adjuvant anthracycline-based chemotherapy on status of stromal CD10 antigens in breast cancer. Materials and Methods: Patients with invasive breast cancer scheduled for anthracycline-based neo-adjuvant chemotherapy were included in the study. Tumor stromal CD10 expression was estimated before and after 3 cycles of chemotherapy, and change in its status was correlated with clinical response to chemotherapy. Results: 16 out of the 29 patients had strong CD10 expression; in these 16 patients, 14 (87.5%) were hormone receptor negative, and 14 (87.5%) had HER-2/neu overexpression. Stromal CD10 expression remained same in 13 out of 29 cases (44.83%) after chemotherapy. There was a change in CD10 expression in the remaining 16 cases (55.17%); in 13 cases (44.83%) it decreased from its pre-chemotherapy status, while its expression increased in 3 cases (10.34%). In cases of complete and partial clinical response, there was no increase in CD10 expression. Where CD10 expression had increased after chemotherapy, there was either a minor response or no response to chemotherapy. In 13 cases where CD10 expression had decreased, 12 cases had a clinical response to chemotherapy. Conclusions: Strong CD10 expression correlates with hormone receptor negativity and HER-2/neu overexpression. Stromal CD10 expression in breast cancer is not static and changes with neo-adjuvant anthracycline-based chemotherapy. A stable or decrease in CD10 expression correlates with complete or partial clinical response, while an increase in CD10 expression appears to correlate with poor clinical response. A larger series is required to determine the clinical significance of these changes. As stromal CD10 expression and its change with chemotherapy may have a prognostic significance, they should be documented in breast cancer patients before and after chemotherapy.

Expression of immunohistochemical markers in patients with AIDS-related lymphoma

AIDS-related lymphomas (ARL) present high biological heterogeneity. For better characterization of this type of lymphoma, the objectives of the present study were to evaluate the expression of immunohistochemical markers of cell differentiation (CD10, Bcl-6, MUM-1) and determine cell origin profile according to Hans' classification of diffuse large B-cell lymphoma in AIDS patients. This study included 72 consecutive patients with ARL diagnosed at the University Hospital, Universidade Federal do Rio de Janeiro (UFRJ) and at the Brazilian Instituto Nacional de Câncer (INCA) from 2000 to 2006. The morphologic distribution of the lymphomas was the following: 61 percent were diffuse large B-cell lymphomas (DLBCLs), 15 percent were Burkitt's lymphomas, 13 percent were plasmablastic lymphomas, 10 percent were high-grade lymphomas and 1 percent was follicular lymphoma. The positivity for each immunohistochemical marker in DLBCLs, Burkitt's lymphoma and plasmablastic lymphoma was respectively: CD20, 84 percent, 100 percent, and 0; CD10, 55 percent, 100 percent, and 0; Bcl-6, 45 percent, 80 percent, and 0; MUM-1, 41 percent, 20 percent, and 88 percent. A higher positivity of CD20 (84 percent x 56 percent, p = 0.01) was found in DLBCL compared to non-DLBCL; in Burkitt's lymphomas a higher positivity of CD10 (100 percent x 49 percent, p = 0.04) and Bcl-6 (80 percent x 39 percent, p = 0.035) were found compared to non-Burkitt's lymphomas. Germinal center (GC) profile was detected in 60 percent of DLBCLs. Our study suggests particular findings in ARL, as the most frequent phenotype was GC, different from HIV-negative patients.

Concurrent hepatic adenomatoid tumor and hepatic hemangioma: a case report

A 45-year-old male with alleged asymptomatic hepatic hemangioma of 4 years duration had right upper-quadrant pain and was referred to a tertiary hospital. Computed tomography and magnetic resonance imaging scans revealed a hypervascular mass of about 7 cm containing intratumoral multilobulated cysts. A preoperative liver biopsy was performed, but this failed to provide a definitive diagnosis. The patient underwent a partial hepatectomy of segments IV and VIII. The histologic findings revealed multifocal proliferation of flattened or cuboidal epithelioid cells and a highly vascular edematous stroma. Immunohistochemistry findings demonstrated that the epithelioid tumor cells were positive for cytokeratin (AE1/AE3), vimentin, calretinin, and cytokeratin 5/6, and were focally positive for CD10, and negative for WT1 and CD34, all of which support their mesothelial origin. Immunohistochemistry for a mesothelial marker should be performed for determining the presence of an adenomatoid tumor when benign epithelioid cells are seen.

Bcl-2 protein frequency in patients with high-risk diffuse large B-cell lymphoma / Frequência da proteína Bcl-2 em pacientes com linfoma difuso de grandes células B de alto risco

CONTEXT AND OBJECTIVE: Gene expression and immunohistochemical profiling of diffuse large B-cell lymphoma (DLBCL) have revealed important prognostic subgroups: germinal center B-cell-like (GCB-like) DLBCL and activated B cell-like (ABC-like) DLBCL. Although few reports on high-risk DLBCL are available, the prognosis for the GCB-like subgroup has been shown to be better than that of the ABC-like subgroup. The role of Bcl-2 as a predictor of survival in DLBCL cases is unclear and its expression varies between the two subgroups of DLBCL. In this study, we analyzed the frequency and prognostic impact of Bcl-2 protein expression in high-risk DLBCL cases. DESIGN AND SETTING: Retrospective cohort study among DLBCL patients treated at Hospital das Clínicas, Faculdade de Medicina da Universidade de São Paulo (HC-FMUSP). METHODS: The prognostic impact of the expression of the proteins CD10, Bcl-6, MUM1 (multiple myeloma oncogene-1) and Bcl-2 on high-risk DLBCL cases was evaluated by means of immunohistochemistry. Seventy-three patients aged 18-60 years were evaluated for all these markers. RESULTS: Twenty-four cases (32.9 percent) were GCB-like and 49 (67.1 percent) were ABC-like, with no difference regarding complete remission, disease-free survival or overall survival rates. Twenty-seven patients (37 percent) showed Bcl-2 expression, which was the only independent factor predicting a worse prognosis for overall survival according to multivariate analysis. CONCLUSION: Bcl-2 protein was expressed in 37 percent of the high-risk DLBCL patients, without any difference between the ABC-like DLBCL and GCB-like DLBCL cases.

CONTEXTO E OBJETIVO: A expressão gênica e imunoistoquímica do linfoma difuso de grandes células B (LDGCB) vem permitindo a identificação de importantes subgrupos prognósticos: LDGCB do centro germinativo (CG) e LDGCB de células B ativadas (CBA). Entretanto, existem poucos dados disponíveis com LDGCB de alto risco, sendo o prognóstico dos LDGCB do CG melhor que os LDGCB de CBA. A participação do Bcl-2 como preditor de sobrevida nos LDGCB não é clara e sua expressão é variável entre os dois subgrupos de LDGCB. Neste estudo é avaliada a frequência e o prognóstico da expressão da proteína Bcl-2 em LDGCB de alto risco. TIPO DE ESTUDO E LOCAL: Estudo de coorte retrospectivo realizado entre portadores de LDGCB tratados no Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo. MÉTODOS: Foi avaliado o impacto prognóstico da expressão das proteínas CD10, Bcl-6, MUM1 (multiple myeloma oncogene-1) e Bcl-2 por imunoistoquímica em LDGCB de alto risco. Foram avaliados, para todos os marcadores, 73 pacientes com idade de 18 a 60 anos. RESULTADOS: Vinte e quatro (32,9 por cento) pacientes foram classificados como LDGCB do CG e 49 (67,1 por cento) como LDGCB de CBA, sem diferença nas taxas de remissão completa, sobrevida livre de doença e sobrevida global. Vinte e sete (37 por cento) apresentaram expressão de Bcl-2, o qual foi o único fator preditivo independente de pior prognóstico de sobrevida global à análise multivariada. CONCLUSÃO: A expressão da proteína Bcl-2 ocorreu em 37 por cento dos portadores de LDGCB de alto risco, sem diferença entre os subgrupos de LDGCB do CG ou de CBA.

Immunophenotypic Features of Granulocytes, Monocytes, and Blasts in Myelodysplastic Syndromes / 대한진단검사의학회지

BACKGROUND: Despite the diagnostic utility of immunophenotyping for myelodysplastic syndromes (MDS), it has not been widely performed, and reports on this are absent in Korea. We aimed to evaluate the immunophenotypic features of non-blastic granulocytes, monocytes, and blasts in patients with MDS and non-clonal disorders using routine flow cytometry (FCM). Moreover, we evaluated the phenotypic abnormalities of mature cells in leukemic patients. METHODS: Marrow aspirates from 60 patients, including 18 with MDS, 18 with leukemia, and 24 with non-clonal disorders (control group), were analyzed using FCM. Blasts, non-blast myeloid cells, and monocytes were gated based on CD45 expression and side scatter (SSC). The phenotypes were then compared among the 3 groups. RESULTS: Compared to non-clonal disorders, the granulocytic lineages of MDS showed decreased SSC (P=0.005), increased CD45 intensity (P=0.020), decreased CD10-positive granulocytes (P= 0.030), and a higher CD56-positive rate (P=0.005). It is noteworthy that similar results were obtained in the leukemia group, and these findings were not related to the phenotypes of the leukemic cells. Using blast and monocytic gating, useful parameters for generating a differential diagnosis were not found. CONCLUSIONS: Gating the granulocytic region is a relatively easy method for MDS immunophenotyping. Among the parameters studied, SSC, CD10, and CD56 were the most useful for differentiating MDS from non-clonal disorders. While immunophenotypic changes in MDS appear to be useful for differentiating MDS from non-clonal disorders, these changes were also noted in the mature cells of leukemic patients.

The use of Fluorescence Resonance Energy Transfer (FRET) peptidesfor measurement of clinically important proteolytic enzymes

Proteolytic enzymes have a fundamental role in many biological processes and are associated with multiple pathological conditions. Therefore, targeting these enzymes may be important for a better understanding of their function and development of therapeutic inhibitors. Fluorescence Resonance Energy Transfer (FRET) peptides are convenient tools for the study of peptidases specificity as they allow monitoring of the reaction on a continuous basis, providing a rapid method for the determination of enzymatic activity. Hydrolysis of a peptide bond between the donor/acceptor pair generates fluorescence that permits the measurement of the activity of nanomolar concentrations of the enzyme. The assays can be performed directly in a cuvette of the fluorimeter or adapted for determinations in a 96-well fluorescence plate reader. The synthesis of FRET peptides containing ortho-aminobenzoic acid (Abz) as fluorescent group and 2, 4-dinitrophenyl (Dnp) or N-(2, 4-dinitrophenyl)ethylenediamine (EDDnp) as quencher was optimized by our group and became an important line of research at the Department of Biophysics of the Federal University of São Paulo. Recently, Abz/Dnp FRET peptide libraries were developed allowing high-throughput screening of peptidases substrate specificity. This review presents the consolidation of our research activities undertaken between 1993 and 2008 on the synthesis of peptides and study of peptidases specificities.

As enzimas proteolíticas têm um papel fundamental em muitos processos biológicos e estão associadas a vários estados patológicos. Por isso, o estudo da especificidade das peptidases pode ser importante para uma melhor compreensão da função destas enzimas e para o desenvolvimento de inibidores. Os substratos com supressão intramolecular de fluorescência constituem uma excelente ferramenta, pois permitem o monitoramento da reação de forma contínua, proporcionando um método prático e rápido para a determinação da atividade enzimática. A hidrólise de qualquer ligação da cadeia peptídica entre o grupo doador e o grupo supressor gera fluorescência que permite detectar concentração nanomolar de enzima. Os ensaios podem ser acompanhados diretamente na cubeta ou adaptados para determinações de fluorescência em leitoras de placa. A síntese dos peptídeos com supressão intramolecular de fluorescência contendo o grupo fluorescente Abz (orto-aminobenzóico) e o grupo supressor EDDnp (N-[2, 4-dinitrofenil]-etilenodiamino ou Dnp (2, 4-dinitrophenyl) foi otimizada pelo nosso grupo e tornou-se uma importante linha de pesquisa no Departamento de Biofísica da Universidade Federal de São Paulo. Recentemente, foram desenvolvidas bibliotecas de peptídeos fluorogênico contendo Abz/Dnp como grupo doador/supressor trazendo um grande avanço no estudo de especificidade das peptidases. Esta revisão apresenta o trabalho desenvolvido pelo nosso grupo entre 1993 e 2008 sobre a síntese de peptídeos e o estudo da especificidade de peptidases.

A Case of Atypical Solid-pseudopapillary Tumor of the Pancreas / 대한소화기학회지

Solid-psudopapillary tumor is an uncommon pancreatic neoplasm of low malignant potential that most frequently affect young woman. Solid-psudopapillary tumor are histologically, clinically, and prognostically quite distinct from the more common ductal adenocarcinoma. Recently, we experienced a 36-year-old male who was suspected to have extrapancreatic tumor based on atypical radiologic imaging study, young age, and male sex, and finally diagnosed as solid-psudopapillary tumor on immunohistochemical stain examination. We report this case with review of the relevant literatures.

Useful markers for differential diagnosis of oncocytoma, chromophobe renal cell carcinoma and conventional renal cell carcinoma.

Renal oncocytoma, conventional RCC (granular cell type) and chromophobe RCC have different prognosis. Sometimes differentiation between them is difficult in HandE slides. In a 5-year study of 128 renal tumors, we selected 76 cases [30 conventional RCC (CRCC), 16 papillary RCC, 21 chromophobe RCC (ChRCC), 8 oncocytoma, 1 collecting duct carcinoma (cdc)] and staining with Hale's colloidal iron, CK7, CK8, CK18, CK19, CK20, Vimentin, EMA, CD10 and RCC marker were done. No significant difference was seen between renal tumor subtypes with CK8, CK18, CK19, CK20 and EMA. The most useful markers were Vimentin, CK7, CD10, RCC marker and Hale's colloidal iron. Hale's colloidal iron staining with diffuse reticular fine cytoplasmic pattern was present in ChRCCs, but was absent in other subtypes and oncocytomas. Vimentin, CK7, CD10, RCC marker and Hale's colloidal iron can be used for the differential diagnosis of problematic epithelial tumors of kidney (CRCC, ChRCC and oncocytoma) - i.e. ChRCC: Vimentin, CD10 and RCC marker - negative, CK7 - positive and positive diffuse fine reticular cytoplasmic pattern of Hale's colloidal iron; oncocytoma: Vimentin, CK7, RCC marker and CD10 - negative and Hale's colloidal iron - negative; CRCC: CK7 - negative, Vimentin, CD10 and RCC marker - positive and Hale's colloidal iron - negative.

CD10 in diffuse uterine leiomyomatosis: a case report along with a few comments on histogenesis.

Diffuse uterine leiomyomatosis (DUL) is a rare entity with an unknown etiopathogenesis. A 24 years old female presented with abdominal discomfort and menorrhagia. Clinical and ultrasonographic examination revealed an enlarged uterus. The hysterectomy specimen showed a symmetrically enlarged uterus with a bosselated external surface. The cut surface showed multiple nodules of varying sizes diffusely involving the myometrium. Microscopically, the nodules were leiomyomas of varying degrees of cellularity. Some of the leiomyomas showed an increased vascularity either in the form of congeries of blood vessels with a lobular arrangement or occasionally as foci of 2-3 vessels. The vessels were surrounded by whorls of spindle cells. On immunohistochemistry the leiomyomas expressed vimentin, smooth muscle actin (SMA), desmin and CD10: the cells whorling around the blood vessels expressed vimentin, SMA and focally desmin and were negative for CD10 and HMB-45. The aim of this paper is to document that CD10 is expressed in diffuse uterine leiomyomatosis and discuss the histogenesis of DUL.

Pure malignant myoepitheliomas of the breast: an immunohistochemical study.

Myoepithelioma of breast are extremely rare. We report two cases of pure malignant myoepithelioma of the breast, utilising light microscopic and immunohistochemical methods for diagnosis. Both the cases presented as breast lump. Hematoxylin and Eosin (H&E) stained microscopic sections revealed a predominantly spindle cell tumor. Immunohistochemical work up was done. Case number one expressed positivity for vimentin, Smooth Muscle Actin (SMA), S-100 and CD10. Case number two expressed positivity for Vimentin, CD10 and p63. This led to the diagnoses of malignant myoepithelioma in both of them. Documentation of such cases prospectively and from archival material, using immunohistochemistry, is of extreme importance to assess the prevalence, various phenotypic patterns, long-term biological behaviour and to establish management protocols for malignant myoepithelioma.

Hepatocyte Expressions in Hepatocellular Carcinomas, Gastrointestinal Neoplasms, and Non-neoplastic Gastrointestinal Mucosa: its Role as a Diagnostic Marker

We performed immunohistochemical staining against Hepatocyte (Hep) and CD10 antibodies in 75 hepatocellular carcinoma (HCC), 50 cholangiocarcinomas, 49 colorectal adenocarcinomas, and 308 gastric adenocarcinomas by tissue array method. We also evaluated the various non-neoplastic adult tissues and fetal digestive organs. Hep was expressed in 80% of HCCs, and HCCs without Hep expression were more likely to have a higher Edmondson & Steiner grade than HCCs with Hep expression (p=0.004). In non-HCCs, 16% of cholangiocarcinomas, 8.2% of colorectal carcinomas, and 44.2% of gastric carcinomas expressed Hep. Gastric carcinomas with Hep expression were significantly associated with early gastric carcinomas (p<0.001). In non-neoplastic tissues, Hep was found expressed in normal hepatocytes, small intestinal mucosa, and intestinal metaplasia of the stomach. Fetal hepatocytes expressed Hep after 19 weeks of gestation. CD10 was detected in 46.7% (35/75) of HCCs, and canalicular staining pattern was predominant in HCCs. In conclusion, the expression of Hep and CD10 may help to distinguish HCCs from non-HCCs.

A new fluorometric assay for neutral endopeptidase (EC 3.4.24.11)

An intramolecularly quenched fluorogenic peptide structurally related to Leu-enkephalin, Abz-GGdFLRRV-EDDnp, was selectively hydrolyzed at the R-V bond by neutral endopeptidase (NEP, enkephalinase, neprilysin, EC 3.4.24.11) with kinetic parameters (Km = 3 µM,Kcat = 127 / min and Kcat / Km = 42 / min µM) similar to those of Leu-enkephalin. The specificity of the assay for NEP was demostrated by incubating Abz-GGdFLRRV-EDDnp with a kidney homogenate and with crude membrane preparations of brain and lung. For all three homogenates the complementary fragments Abz-GGdFLRRnp accounted for more than 95 percent of the products wich were totally inhibited by 1 µM thiorphan, a highly specific NEP inhibitor. A continuous fluorometric assay for only 5 min was sufficient to quantify the NEP activity with a minimum sensitivity of 5 ng of purified NEP or the equivalent enzymatic activity in crude tissue preparations.