Umbilical cord blood stem cells: Towards a proteomic approach

The first umbilical cord blood (UCB) transplant to a sibling with Fanconi's anaemia in 1988 represented a breakthrough in the field of transplantation. Thereon, several transplants have been performed with UCB-derived hematopoietic stem cells (HSCs) and a plethora of studies have investigated the plasticity of UCB-derived stem and progenitor cells. However, these studies have not been hitherto translated into clinical trials and, although UCB is routinely used as an alternative source of HSCs, no substantial advances have been made in the field of clinical regenerative medicine. The real deal is the lack of knowledge about the molecular processes governing the events of differentiation which transform immature UCB stem cells into terminally-committed hematopoietic, muscle, bone and nervous cells. In order to fill this void, several studies have been recently focused on the identification of the peculiar proteomic profile of UCB-derived stem cells.Hereby, we concisely review recent proteomic surveys addressing UCB-derived stem and progenitor cells.Notably, comparative studies detected a wider spectrum of proteins in immature cells rather than in more differentiated populations, as if maturation events could represent a bottleneck to protein expression. Future research projects should try to shed light on these processes and their completion could pave the way for unprecedented treatments.     

Evaluation of the House Fly Musca domestica as a Mechanical Vector for an Anthrax

Anthrax is a disease of human beings and animals caused by the encapsulated, spore-forming, Bacillus anthracis. The potential role of insects in the spread of B. anthracis to humans and domestic animals during an anthrax outbreak has been confirmed by many studies. Among insect vectors, the house fly Musca domestica is considered a potential agent for disease transmission. In this study, laboratory-bred specimens of Musca domestica were infected by feeding on anthrax-infected rabbit carcass or anthrax contaminated blood, and the presence of anthrax spores in their spots (faeces and vomitus) was microbiologically monitored. It was also evaluated if the anthrax spores were able to germinate and replicate in the gut content of insects. These results confirmed the role of insects in spreading anthrax infection. This role, although not major, given the huge size of fly populations often associated with anthrax epidemics in domestic animals, cannot be neglected from an epidemiological point of view and suggest that fly control should be considered as part of anthrax control programs.     

Synthesis and biological assessment of simplified analogues of the potent microtubule stabilizer (+)-discodermolide

An efficient, convergent and stereocontrolled synthesis of simplified analogues of the potent antimitotic agent (+)-discodermolide has been achieved and several small libraries have been prepared. In all the libraries, the discodermolide methyl groups at C14 and C16 and the C7 hydroxy group were removed and the lactone was replaced by simple esters. Other modifications introduced in each series of analogues were related to C11, C17 and C19 of the natural product. Key elements of the synthetic strategy included (a) elaboration of the main subunits from a common intermediate and (b) fragment couplings using Wittig reactions to install the (Z)-olefins. Library components were analyzed for microtubule-stabilizing actions in vitro, for displacement of [3H]paclitaxel from its binding site on tubulin, for antiproliferative activity against human carcinoma cells, and for cell signaling and mitotic spindle alterations by a multiparameter fluorescence cell-based screening technique. The results show that even significant structural simplification can lead to analogues with actions related to microtubule targeting.     

Presenilin mutations linked to familial Alzheimer's disease reduce endoplasmic reticulum and Golgi apparatus calcium levels

Presenilin-1 and -2 (PS1 and PS2) mutations, the major cause of familial Alzheimer's disease (FAD), have been causally implicated in the pathogenesis of neuronal cell death through a perturbation of cellular Ca(2+) homeostasis. We have recently shown that, at variance with previous suggestions obtained in cells expressing other FAD-linked PS mutations, PS2-M239I and PS2-T122R cause a reduction and not an increase in cytosolic Ca(2+) rises induced by Ca(2+) release from stores. In this contribution we have used different cell models: human fibroblasts from controls and FAD patients, cell lines (SH-SY5Y, HeLa, HEK293, MEFs) and rat primary neurons expressing a number of PS mutations, e.g. P117L, M146L, L286V, and A246E in PS1 and M239I, T122R, and N141I in PS2. The effects of FAD-linked PS mutations on cytosolic Ca(2+) changes have been monitored either by using fura-2 or recombinant cytosolic aequorin as the probe. Independently of the cell model or the employed probe, the cytosolic Ca(2+) increases, caused by agonist stimulation or full store depletion by drug treatment, were reduced or unchanged in cells expressing the PS mutations. Using aequorins, targeted to the endoplasmic reticulum or the Golgi apparatus, we here show that FAD-linked PS mutants lower the Ca(2+) content of intracellular stores. The phenomenon was most prominent in cells expressing PS2 mutants, and was observed also in cells expressing the non-pathogenic, "loss-of-function" PS2-D366A mutation. Taken as a whole, our findings, while confirming the capability of presenilins to modify Ca(2+) homeostasis, suggest a re-evaluation of the "Ca(2+) overload" hypothesis in AD and a new working hypothesis is presented.     

Specific expression of galanin in the peri-infarct zone after permanent focal cerebral ischemia in the rat

Galanin (Gal) is a neuropeptide with supposed neurotrophic-like action. In the present study, expression of Gal has been investigated in the core and peri-infarct zone at 1, 4, 24 and 72 h after middle cerebral artery occlusion (MCAo) in the rat. Three days after MCAo a small but consistent number of morphological intact Gal-positive neuronal cells were observed in the peri-infarct zone. Gal-positive cells were barely detectable in the infarct and peri-infarct zone at 24 h. No Gal immunopositive cells were detected in brain subjected to 1 and 4 h of ischemia. Gal immunoreactivity was also detected in myelinated fibers 4 and 24 h after focal ischemia. Gal may be a peptide with neurotrophic and plasticity functions under stress conditions.     

D-4F decreases brain arteriole inflammation and improves cognitive performance in LDL receptor-null mice on a Western diet

LDL receptor-null mice on a Western diet (WD) have inflammation in large arteries and endothelial dysfunction in small arteries, which are improved with the apolipoprotein A-I mimetic D-4F. The role of hyperlipidemia in causing inflammation of very small vessels such as brain arterioles has not previously been studied. A WD caused a marked increase in the percent of brain arterioles with associated macrophages (microglia) (P < 0.01), which was reduced by oral D-4F but not by scrambled D-4F (ScD-4F; P < 0.01). D-4F (but not ScD-4F) reduced the percent of brain arterioles associated with CCL3/macrophage inflammatory protein-1alpha (P < 0.01) and CCL2/monocyte chemoattractant protein-1 (P < 0.001). A WD increased (P < 0.001) brain arteriole wall thickness and smooth muscle alpha-actin, which was reduced by D-4F but not by ScD-4F (P < 0.0001). There was no difference in plasma lipid levels, blood pressure, or arteriole lumen diameter with D-4F treatment. Cognitive performance in the T-maze continuous alternation task and in the Morris Water Maze was impaired by a WD and was significantly improved with D-4F but not ScD-4F (P < 0.05). We conclude that a WD induces brain arteriole inflammation and cognitive impairment that is ameliorated by oral D-4F without altering plasma lipids, blood pressure, or arteriole lumen size.     

Bortezomib induces in HepG2 cells IκBα degradation mediated by caspase-8

Abstact The present paper demonstrates that the proteasome inhibitor bortezomib, which behaves as an apoptotic agent in hepatoma HepG2 cells, caused in these cells a decrease in IκBα level and a consequent increase in NF-κB activity. The effect already appeared at 4 h of treatment and preceded the onset of apoptosis which was observed at 24 h. Our results demonstrate that bortezomib-induced IκBα degradation occurred in conjunction with the activation of caspase-8; moreover, the decrease in IκBα level was prevented in a dose-dependent manner by the addition of z-IETD, a specific inhibitor of caspase-8. Bortezomib caused the same effects in non-tumor Chang liver cells, which were not susceptible to the apoptotic effect of the drug. Our results also show that other proteases, such as caspase-3 and calpains, exerted only a limited effect on IκBα degradation. These findings suggest that caspase-8 can be involved in the control of IκBα level. In addition, the activation of caspase-8 can exert, at least in the first phase of treatment with bortezomib, a protective effect in both HepG2 and Chang liver cells, favouring the activation of the survival factor NF-κB     

Cellulase production by trichoderma harzianum in static and mixed solid-state fermentation reactors under nonaseptic conditions

Cellulase production from lignocellulosic materials was studied in solid-state cultivation by both static and mixed techniques under nonaseptic conditions. The effects of fermentation conditions, such as moisture content, pH, temperature, and aeration, on cellulase production by Trichoderma harzianum using a mixture of wheat straw (80%) and bran (20%) were investigated. With a moisture content of 74% and a pH of 5.8., 18 IU filter paper activity and 198 IU endoglucanase activity/g initial substrate content were obtained in 66 h. The extension from static column cultivation to stirred tank reactor of 65 L capacity gave similar yields of cellulase.