Different sensitivity to apoptosis in cells of monocytic or lymphocytic origin chronically infected with human immunodeficiency virus type-1

Apoptotic death of CD4+ T lymphocytes is a major cause of the immunodeficiency caused by human immunodeficiency virus (HIV), but it is still unclear how this process precisely occurs. To characterize a potentially useful cellular model, we have analyzed the tendency of chronically HIV-infected CD4+ human cell lines of different origin to undergo apoptosis. We studied ACH-2 and U1 lines, derived from the CD4+ T-cell A301 and the promonocytic U937 cell lines, respectively, and induced apoptosis via several stimuli that trigger different pathways. Their capacity to regulate plasma membrane CD95 expression and to produce soluble CD95 was also analyzed. Using staurosporine, TNF-alpha plus cycloheximide, and gamma-radiations, we observed that ACH-2 were more sensitive to programmed cell death than A301, while U1 were less sensitive than U937. Both infected cell types had a lower sensitivity to CD95-induced apoptosis; the analysis of changes in mitochondrial membrane potential corroborated these observations. Plasma membrane CD95 was similarly regulated in all cell types, which, however, presented a different capacity to produce soluble CD95 molecules. Our in vitro results may offer a new perspective for developing further studies on the pathogenesis of HIV infection. A chronically infected cell line of lymphocytic origin is more susceptible to apoptosis than its parental cell type, while infected monocytic cells are less sensitive than their uninfected counterpart. Thus, it is possible to hypothesize that one of the reasons by which circulating monocytes survive and represent a viral reservoir is the capacity of HIV to decrease the sensitivity to apoptosis of this cell type. However, further studies on ex-vivo collected fresh cells, as well as on other cell lines, are urgently needed to confirm such hypothesis.     

Role of connexin37 and connexin40 in vascular development

Mice lacking both connexin37 (Cx37) and connexin40 (Cx40), gap junction proteins expressed in vascular endothelium, die perinatally with pronounced vascular abnormalities. Early vasculogenesis proceeds normally, but by E18.5 Cx37(-/-)Cx40(-/-) animals display vessel dilatation and congestion as well as localized hemorrhages in skin, testis, intestines, and lungs. Abnormal vascular channels are present in the testis, often forming cavernous hemangioma-like defects. Unusually large, distended vessels are also present in the submucosa and lamina propria of the intestine. Ablation of Cx40 has a greater effect on endothelial dye-transfer than ablation of Cx37, and the effect of Cx40 ablation is age-dependent. Only in embryonic aortas lacking both Cx37 and Cx40 is there a complete loss of endothelial coupling. Surprisingly, elimination of Cx40 results in a large drop in aortic endothelial Cx37 on western blots, and deletion of Cx37 also reduces endothelial Cx40 levels. In contrast, in the medial layer, both Cx37 and Cx43 increase when Cx40 is ablated. These studies indicate that Cx37 and Cx40 are collectively critical for endothelial communication and provide evidence of an important role for gap junctions in vascular development. In addition, Cx37 and Cx40 appear to be mutually dependent on each other for normal expression in vascular endothelium.     

One eye but no vision: cave fish with induced eyes do not respond to light

One of the most intriguing questions in evolutionary biology is the degree to which behavior is a necessary consequence of morphology. We explore this issue by examining phototactic behavior in epigean (eyed surface-dwelling) and troglomorphic (blind cave) forms of the teleost Astyanax fasciatus whose eyes were modified during embryogenesis by removing one or both lens vesicles from the epigean form or by transplanting the lens vesicle from an epigean fish into the optic cup of a blind cave form. Lens removal results in eye degeneration and blindness in adult epigean fish, whereas lens transplantation stimulates growth of the eye, inducing the development of optic tissues in the normally eyeless adult cave fish. Photoresponsiveness was examined by placing fish in an aquarium with one half illuminated and the other half dark and scoring their presence in the illuminated or dark half. Both the eyeless epigean fish and cave fish with induced eyes are indifferent to the illumination whereas the surface forms are scotophilic, suggesting that optic development and phototactic behavior are decoupled.     

Soil-borne <Emphasis Type="Italic">Penicillium</Emphasis> spp. and other microfungi as efficient degraders of the explosive RDX (hexahydro-1,3,5-trinitro-1,3,5-triazine)

Soil microfungi belonging to the genera Aspergillus, Coniothyrium, Paecilomyces, Penicillium and Trichoderma, as well as wood-and litter-decomposing basidiomycetes, were able to degrade the explosive RDX (hexahydro-1,3,5-trinitro-1,3,5-triazine) co-metabolically, but were unable to utilize it as a sole carbon or nitrogen source. The most efficient RDX-degrading microfungi were characterized morphologically and by analysis of the ITS region of the ribosomal RNA gene cluster as Penicillium janczewskii and an unidentifiable Penicillium sp. with uniseriate phialides. Both species catalysed 80–100 % disappearance of RDX in a liquid defined medium. RDX degradation was inhibited by the presence of 30 mM NH₄ ⁺ but not by 40 mM NO₃ ⁻. In basidiomycetes but not Penicillium spp., RDX degradation was greatly reduced when biomass pregrown at 23 °C was incubated with RDX at 15 °C. Because of their production of copious conidial inoculum, simple growth requirements and ability to degrade RDX at reduced temperature, Penicillium spp. show promise for the bioremediation of RDX-contaminated groundwater.     

The role of nitrate reductase in the degradation of the explosive RDX (hexahydro-1,3,5-trinitro-1,3,5-triazine) by <Emphasis Type="Italic">Penicillium</Emphasis> sp. AK96151

In liquid culture on a defined growth medium, Penicillium sp. AK96151 efficiently degraded the explosive hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX, hexogen), causing > 80 % disappearance after 10 d. RDX degradation was reduced to a basal level (< 15 % degraded after 10 d) by the presence of > 150 μM ammonium ions or when the molybdenum component of the medium was replaced by sodium tungstate. An equivalent effect of ammonium, molybdenum and tungsten was observed in protoplasts of this fungus assayed for nitrate reductase activity. This enzyme was not inhibited by RDX itself. The involvement of a nitrate reductase in RDX degradation by Penicillium has practical implications for bioremediation strategies which are discussed.     

Proteins from bovine tissues and biological fluids: defining a reference electrophoresis map for liver,kidney, muscle,plasma and red blood cells

A number of high resolution two-dimensional electrophoresis (2-DE) reference maps for bovine tissues and biological fluids have been determined for animals in basal state. Among the 1863 distinct protein features detected in samples of liver, kidney, muscle, plasma and red blood cells, 509 species were identified and associated to 209 different genes. Difficulties in the identification were related to the poorly characterized Bos taurus genome and were solved by a combined matrix-assisted laser desorption/ionisation-mass spectrometry and liquid chromatography-electrospray ionization tandem mass spectrometry approach. The experimental output allowed us to establish a 2-DE database accessible through the World Wide Web network at the URL address (http://www.iabbam.na.cnr.it/Biochem). These reference maps may serve as a tool in future veterinary medical studies aimed at the evaluation of changes in protein repertoire for altered animal physiological conditions and infectious diseases, to the definition of molecular markers for novel diagnostic kits and vaccines, as well as the characterization of protein modifications in bovine materials following technological processes used in the food industry.     

The presence of a chromatin boundary appears to shield a transgene in tobacco from RNA silencing

We present isogenic transgenic tobacco lines that carry at a given chromosomal position a beta-glucuronidase (GUS) reporter gene either with or without the presence of the matrix-associated region known as the chicken lysozyme A element. Plants were generated with the Cre-lox site-specific recombination system using heterospecific lox sites. Analysis of GUS gene expression in plant populations demonstrates that the presence of the A element can shield against RNA silencing of the GUS gene. Protection was observed in two of three independent tobacco transformants. Plants carrying an A element 5' of the GUS gene always had stable GUS activity, but upon removal of this A element, the GUS gene became silenced over time in two lines, notably when homozygous.     

Peripheral benzodiazepine receptor ligand-melphalan conjugates for potential selective drug delivery to brain tumors

To gain insight into the strategy to target PBR ligand-drug conjugates to brain tumors, novel N-imidazopyridinacetyl-melphalan conjugates and the corresponding ethyl esters have been prepared and evaluated for their cytotoxicity in melphalan-sensitive human (SF126, SF188) and rat (RG-2) glioma cell lines. These conjugates exhibited PBR binding affinity with IC(50) values ranging from 57 and 2614 nM. By a computational approach it can be predicted that these conjugates possess significant brain penetration. The stability of the conjugates in 0.05 M phosphate buffer at pH 7.4 and, in some cases, in dilute human serum solution was determined. All the ethyl ester derivatives were stable in 0.05 M phosphate buffer at pH 7.4 and their half-lives exceeded 28 h. Conversely, under the same conditions, the corresponding acids were found to undergo a fast cleavage within a few minutes. HPLC-MS analysis of the mixture from degradation in buffer and physiological medium of the representative cases allowed the identification of their main degradation products. A plausible degradation pathway accounting for the available experimental data is presented.