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Typical hemolytic uremic syndrome (HUS) is mainly caused by Shiga toxin-producing Escherichia coli (STEC) releasing Shiga toxin 2 (Stx2). Two different structures of this AB5 toxin have been described: uncleaved, with intact B and A chains, and cleaved, with intact B and a nicked A chain consisting of two fragments, A1 and A2, connected by a disulfide bond. Despite having the same toxic effect on sensitive cells, the two forms differ in their binding properties for circulating cells, serum components and complement factors, thus contributing to the pathogenesis of HUS differently. The outcome of STEC infections and the development of HUS could be influenced by the relative amounts of uncleaved or cleaved Stx2 circulating in patients' blood. Cleaved Stx2 was identified and quantified for the first time in four out of eight STEC-infected patients' sera by a method based on the inhibition of cell-free translation. Cleaved Stx2 was present in the sera of patients with toxins bound to neutrophils and in two out of three patients developing HUS, suggesting its involvement in HUS pathogenesis, although in association with other bacterial or host factors. [ABSTRACT FROM AUTHOR]

Clomiphene citrate (CC) in male hypogonadism increases testosterone (T) and estrogen levels by stimulating pituitary gonadotropin release. Our group confirmed these hormonal changes in a randomized, cross-over, double-blind trial of CC versus placebo in addition to metformin, conducted in 21 obese dysmetabolic men with low T levels. However, we hypothesize that based on its mechanism of action, CC may directly or indirectly affect adrenal steroidogenesis. The aim of this sub-study was to better understand the changes in steroid levels and metabolism induced by CC treatment. We assessed 17α-hydroxypregnelone (17αOH-P5), dehydroepiandrosterone (DHEA), progesterone (P4), 17α-hydroxyprogesterone (17αOH-P4), androstenedione (A), T, dihydrotestosterone (DHT), estrone (E1), 17β-estradiol (E2), 11-deoxycortisol (11 S), cortisol (F), and cortisone (E) by LC-MS/MS, and corticosteroid binding globulin (CBG) by ELISA, before and after each treatment. In addition, free-F and steroid product/precursor ratios were calculated. We observed a significant change in serum levels induced by CC compared with placebo for 17αOH-P4, DHT, T, E2, E1, F, E, and CBG, but not free-F. In addition, compared to placebo, CC induced higher 17αOH-P4/P4, E2/E1, 17αOH-P4/17αOH-P5, A/17αOH-P4, T/A, E1/A, F/11 S, and F/E ratios. Therefore, besides the CC stimulating effect on testis steroidogenesis, our study showed increased F, E, but not free-F, levels, indicating changes in steroid metabolism rather than adrenal secretion stimulation. The steroid profiling also revealed the CC stimulation of the Δ5 rather than the Δ4 pathway, thus indicating considerable testicular involvement in the increased androgen secretion. [ABSTRACT FROM AUTHOR]

A 62-year-old woman complaining of severe malabsorption was diagnosed with celiac disease based on the findings of flat, small intestinal mucosa and HLA-DQ2 positivity, although celiac serology was negative. This diagnosis was questioned due to the lack of clinical and histological improvement after a long period of strict gluten-free diet. The detection of enterocyte autoantibodies guided to the correct diagnosis of autoimmune enteropathy, leading to a complete recovery of the patient following an appropriate immunosuppressive treatment. Autoimmune enteropathy should be considered in the differential diagnosis of malabsorption with severe villous atrophy, including those cases with negative celiac-related serology. [ABSTRACT FROM AUTHOR]

Mutations of genes encoding the subunits of the succinate dehydrogenase (SDH) complex were described in KIT/PDGFRA wild-type GIST separately in different reports. In this study, we simultaneously sequenced the genome of all subunits, SDHA, SDHB, SDHC, and SDHD in a larger series of KIT/PDGFRA wild-type GIST in order to evaluate the frequency of the mutations and explore their biological role. SDHA, SDHB, SDHC, and SDHD were sequenced on the available samples obtained from 34 KIT/PDGFRA wild-type GISTs. Of these, in 10 cases, both tumor and peripheral blood (PB) were available, in 19 cases only tumor, and in 5 cases only PB. Overall, 9 of the 34 patients with KIT/PDGFRA wild-type GIST carried mutations in one of the four subunits of the SDH complex (six patients in SDHA, two in SDHB, one in SDHC). WB and immunohistochemistry analysis showed that patients with KIT/PDGFRA wild-type GIST who harbored SDHA mutations exhibited a significant downregulation of both SDHA and SDHB protein expression, with respect to the other GIST lacking SDH mutations and to KIT/PDGFRA-mutated GIST. Clinically, four out of six patients with SDHA mutations presented with metastatic disease at diagnosis with a very slow, indolent course. Patients with KIT/PDGFRA wild-type GIST may harbor germline and/or de novo mutations of SDH complex with prevalence for mutations within SDHA, which is associated with a downregulation of SDHA and SDHB protein expression. The presence of germline mutations may suggest that these patients should be followed up for the risk of development of other cancers. [ABSTRACT FROM AUTHOR]

Inflammation promotes colorectal carcinogenesis. Tumour growth often generates a hypoxic environment in the inner tumour mass. We here report that, in colon cancer cells, the expression of the pro-inflammatory enzyme cyclooxygenase-2 (COX-2) associates with that of the hypoxia response gene carbonic anhydrase-IX (CA-IX). The COX-2 knockdown, achieved by the stable infection of a COX-2 specific short harpin RNA interference (shCOX-2), down-regulates CA-IX gene expression. In colorectal cancer (CRC) cells, PGE2, the main COX-2 gene products, promotes CA-IX gene expression by ERK1/2 activation. In normoxic environment, shCOX-2 infected/CA-IX siRNA transfected CRC cells show a reduced level of active metalloproteinase-2 (MMP-2) that associates with a decreased extracellular matrix invasion capacity. In presence of hypoxia, COX-2 gene expression and PGE2 production increase. The knockdown of COX-2/CA-IX blunts the survival capability of CRC cells in hypoxia. At a high cell density, a culture condition that creates a mild pericellular hypoxic environment, the expression of COX-2/CA-IX genes is increased and triggers the invasive potential of colon cancer cells. In human colon cancer tissues, COX-2/CA-IX protein expression levels, assessed by Western blot and immunohistochemistry, correlate each other and increase with tumour stage. In conclusion, these data indicate that COX-2/CA-IX interplay promotes the aggressive behaviour of CRC cells. [ABSTRACT FROM AUTHOR]

High serum levels of IL-6 correlate with poor outcome in breast cancer patients. However, no data are available on the relationship between IL-6 and mammary stem/progenitor cells, which may fuel the genesis of breast cancer in vivo. Herein, we address this issue in the MCF-7 breast cancer cell line and in primary human mammospheres (MS), multicellular structures enriched in stem/progenitor cells of the mammary gland. MS from node invasive breast carcinoma tissues expressed IL-6 mRNA at higher levels than did MS from matched non-neoplastic mammary glands. In addition, IL-6 mRNA was detected only in basal-like breast carcinoma tissues, an aggressive breast carcinoma variant showing stem cell features. IL-6 treatment triggered Notch-3-dependent upregulation of the Notch ligand Jagged-1 and promotion of MS and MCF-7-derived spheroid growth. Moreover, IL-6 induced Notch-3-dependent upregulation of the carbonic anhydrase IX gene and promoted a hypoxia-resistant/invasive phenotype in MCF-7 cells and MS. Finally, autocrine IL-6 signaling relied upon Notch-3 activity to sustain the aggressive features of MCF-7-derived hypoxia-selected cells. In conclusion, these data support the hypothesis that IL-6 induces malignant features in Notch-3-expressing stem/progenitor cells from human ductal breast carcinoma and normal mammary gland. [ABSTRACT FROM AUTHOR]