Testosterone, dihydrotestosterone and oestradiol levels in postmenopausal breast cancer tissues

The ability of breast tumours to synthesize hormones is well recognized, and local production of sex steroids is thought to play a role in breast cancer growth. We measured the intratumour and circulating levels of testosterone, dihydrotestosterone (DHT) and oestradiol in 35 histologically confirmed carcinomatous mammary tissues obtained at breast surgery from 34 postmenopausal patients, age 50–85 years. Intra-tissue steroids were extracted with ethanol:acetone (1:1; v/v), defatted with 70% methanol in water, and extracted with ether. Steroids, from tissue and serum, were separated by partition chromatography on celite columns and were measured by RIA. Intratumour testosterone and DHT concentrations were significantly correlated, after the exclusion of an outlier (r_s = 0.71; P = 0.0001). No association was found between oestradiol and either of the two androgens. Mean oestradiol and DHT concentrations were significantly higher in tissue than in blood (P = 0.0001). Mean testosterone levels in tissues did not significantly differ from those measured in blood. Our data suggest that at least a part of intratissue DHT is produced locally from testosterone. The meaning of high oestradiol and DHT levels in cancer tissue still needs to be defined.     

Immunohistochemistry in the evaluation of neovascularization in tumor xenografts

Angiogenesis, or neovascularization, is known to play an important role in the neoplastic progression leading to metastasis. CD31 or Factor VIII-related antigen (F VIII RAg) immunohistochemistry is widely used in experimental studies for quantifying tumor neovascularization in immunocompromised animal models implanted with transformed human cell lines. Quantification, however, can be affected by variations in the methodology used to measure vascularization including antibody selection, antigen retrieval (AR) pretreatment, and evaluation techniques. To examine this further, we investigated the microvessel density (MVD) and the intensity of microvascular staining among five different human tumor xenografts and a mouse syngeneic tumor using anti-CD31 and F VIII RAg immunohistochemical staining. Different AR methods also were evaluated. Maximal retrieval of CD31 was achieved using 0.5 M Tris (pH 10) buffer, while maximum retrieval of F VIII RAg was achieved using 0.05% pepsin treatment of tissue sections. For each optimized retrieval condition, anti-CD31 highlighted small vessels better than F VIII RAg. Furthermore, the MVD of CD31 was significantly greater than that of F VIII RAg decorated vessels (p<0.001). The choice of antibody and AR method has a significant affect on immunohistochemical findings when studying angiogenesis. One also must use caution when comparing studies in the literature that use different techniques and reagents.     

Resveratrol as a Pan-HDAC Inhibitor Alters the Acetylation Status of Jistone Proteins in Human-Derived Hepatoblastoma Cells

The polyphenolic alcohol resveratrol has demonstrated promising activities for the prevention and treatment of cancer. Different modes of action have been described for resveratrol including the activation of sirtuins, which represent the class III histone deacetylases (HDACs). However, little is known about the activity of resveratrol on the classical HDACs of class I, II and IV, although these classes are involved in cancer development or progression and inhibitors of HDACs (HDACi) are currently under investigation as promising novel anticancer drugs. We could show by in silico docking studies that resveratrol has the chemical structure to inhibit the activity of different human HDAC enzymes. In vitro analyses of overall HDAC inhibition and a detailed HDAC profiling showed that resveratrol inhibited all eleven human HDACs of class I, II and IV in a dose-dependent manner. Transferring this molecular mechanism into cancer therapy strategies, resveratrol treatment was analyzed on solid tumor cell lines. Despite the fact that hepatocellular carcinoma (HCC) is known to be particularly resistant against conventional chemotherapeutics, treatment of HCC with established HDACi already has shown promising results. Testing of resveratrol on hepatoma cell lines HepG2, Hep3B and HuH7 revealed a dose-dependent antiproliferative effect on all cell lines. Interestingly, only for HepG2 cells a specific inhibition of HDACs and in turn a histone hyperacetylation caused by resveratrol was detected. Additional testing of human blood samples demonstrated a HDACi activity by resveratrol ex vivo. Concluding toxicity studies showed that primary human hepatocytes tolerated resveratrol, whereas in vivo chicken embryotoxicity assays demonstrated severe toxicity at high concentrations. Taken together, this novel pan-HDACi activity opens up a new perspective of resveratrol for cancer therapy alone or in combination with other chemotherapeutics. Moreover, resveratrol may serve as a lead structure for chemical optimization of bioavailability, pharmacology or HDAC inhibition.     

The nucleoside diphosphate kinase activity of DRnm23 is not required for inhibition of differentiation and induction of apoptosis in 32Dcl3 myeloid precursor cells

DRnm23 belongs to a multigene family which includes nm23-H1, the first bona fide metastasis suppressor gene, nm23-H2, nm23-H4, and nm23-H5. Like nm23-H1, nm23-H2, and nm23-H4, DRnm23 possesses nucleoside diphosphate kinase (NDPK) activity. Upon overexpression in myeloid precursor 32Dcl3 cells, DRnm23 inhibits granulocytic differentiation and promotes apoptosis. Two specific mutants of DRnm23 (H134Q and S136P), at residues required for the NDPK activity, inhibit differentiation and promote apoptosis of 32Dcl3 cells. By contrast, substitution of serine 61 with proline (S61P) or deletion of the RGD domain (DeltaRGD) abrogates the effects of wild-type DRnm23. Like wild-type DRnm23, all four mutants show a predominantly mitochondrial subcellular localization. These studies indicate that the enzymatic activity of DRnm23 is not required for the effects observed in 32Dcl3 cells. Moreover, the inability of the S61P and DeltaRGD DRnm23 mutants to inhibit differentiation and promote apoptosis may be due to defective protein-protein interactions at the mitochondria, the predominant site of DRnm23 subcellular localization.     

Dietary input of microbes and host genetic variation shape among-population differences in stickleback gut microbiota

To explain differences in gut microbial communities we must determine how processes regulating microbial community assembly (colonization, persistence) differ among hosts and affect microbiota composition. We surveyed the gut microbiota of threespine stickleback (Gasterosteus aculeatus) from 10 geographically clustered populations and sequenced environmental samples to track potential colonizing microbes and quantify the effects of host environment and genotype. Gut microbiota composition and diversity varied among populations. These among-population differences were associated with multiple covarying ecological variables: habitat type (lake, stream, estuary), lake geomorphology and food- (but not water-) associated microbiota. Fish genotype also covaried with gut microbiota composition; more genetically divergent populations exhibited more divergent gut microbiota. Our results suggest that population level differences in stickleback gut microbiota may depend more on internal sorting processes (host genotype) than on colonization processes (transient environmental effects).     

Caspase-mediated cleavage of the exosome subunit PM/Scl-75 during apoptosis

Recent studies have implicated the dying cell as a potential reservoir of modified autoantigens that might initiate and drive systemic autoimmunity in susceptible hosts. A number of subunits of the exosome, a complex of 3'→5' exoribonucleases that functions in a variety of cellular processes, are recognized by the so-called anti-PM/Scl autoantibodies, found predominantly in patients suffering from an overlap syndrome of myositis and scleroderma. Here we show that one of these subunits, PM/Scl-75, is cleaved during apoptosis. PM/Scl-75 cleavage is inhibited by several different caspase inhibitors. The analysis of PM/Scl-75 cleavage by recombinant caspase proteins shows that PM/Scl-75 is efficiently cleaved by caspase-1, to a smaller extent by caspase-8, and relatively inefficiently by caspase-3 and caspase-7. Cleavage of the PM/Scl-75 protein occurs in the C-terminal part of the protein at Asp369 (IILD³⁶⁹↓G), and at least a fraction of the resulting N-terminal fragments of PM/Scl-75 remains associated with the exosome. Finally, the implications of PM/Scl-75 cleavage for exosome function and the generation of anti-PM/Scl-75 autoantibodies are discussed.     

Responses to dehydration in tadpoles of Physalaemus nattereri (Anura: Leptodactylidae)

Hydrobiologia - In species with complex life cycles, such as anuran amphibians, several traits are influenced by ecological factors during ontogeny. The anuran Physalaemus nattereri shows an...     

Identification of DNA sequence variation in Campylobacter jejuni strains associated with the Guillain-Barré syndrome by high-throughput AFLP analysis

Background  

Campylobacter jejuni is the predominant cause of antecedent infection in post-infectious neuropathies such as the Guillain-Barré (GBS) and Miller Fisher syndromes (MFS). GBS and MFS are probably induced by molecular mimicry between human gangliosides and bacterial lipo-oligosaccharides (LOS). This study describes a new C. jejuni-specific high-throughput AFLP (htAFLP) approach for detection and identification of DNA polymorphism, in general, and of putative GBS/MFS-markers, in particular.