Molecular characterisation of a novel family of cysteine-rich proteins of Toxoplasma gondii and ultrastructural evidence of oocyst wall localisation

Among apicomplexan parasites, the coccidia and Cryptosporidium spp. are important pathogens of livestock and humans, and the environmentally resistant stage (oocyst) is essential for their transmission. Little is known of the chemical and molecular composition of the oocyst wall. Currently, the only parasite molecules shown to be involved in oocyst wall formation are the tyrosine-rich proteins gam56, gam82 and gam230 of Eimeria spp. and the cysteine-rich proteins COWP1 and COWP8 of Cryptosporidium parvum. In the present study, we searched the ToxoDB database for the presence of putative Toxoplasma gondii oocyst wall proteins (OWPs) and identified seven candidates, herein named TgOWP1 through TgOWP7, showing homology to the Cryptosporidium COWPs. We analysed a cDNA library from partially sporulated oocysts of T. gondii and cloned the full-length cDNAs encoding TgOWP1, TgOWP2 and TgOWP3, which consist of 499, 462 and 640 amino acids, respectively. The three proteins share 24% sequence identity with each other and a markedly similar overall structure, based on the presence of an N-terminal leader peptide followed by tandem duplications of a six-cysteine amino acid motif closely related to the Type I repeat of COWPs. Using antisera to recombinant TgOWP1, TgOWP2 and TgOWP3, we showed by Western blot that these molecules are expressed in T. gondii oocysts but are not detectable in tachyzoites. The solubilisation of TgOWP1–3 strictly depended on the presence of reducing agents, consistent with a likely involvement of these proteins in multimeric complexes mediated by disulphide bridges. Immunofluorescence analysis allowed the localisation of TgOWP1, TgOWP2 and TgOWP3 to the oocyst wall. Additionally, using immunoelectron microscopy and the 1G12 monoclonal antibody, TgOWP3 was specifically detected in the outer layer of the oocyst wall, thus representing the first validated molecular marker of this structure in T. gondii.     

Neurogenic-committed human pre-adipocytes express CYP1A isoforms

Stem cell models offer an opportunity both for therapeutic use and for the assessment of alternative in vitro models. Human lipoaspirate is a source of adult stem cells (pre-adipocytes), which are able to differentiate into various phenotypes, such as neurogenic lineage. Here, we analyse the suitability of these in vitro models in screening exogenous compounds, such as environmental pollutants, that may affect adipose cells and neurogenic development. To evaluate neurogenic differentiation, we analysed expression of cholinergic system and acetylcholinesterase immunoreactivity. Heterocyclic derivatives of polycyclic aromatic hydrocarbons (PAHs) are often significant components of environmental contaminants. As they contain inducers of cytochrome P450 1A1 (CYP1A1), we explored the activity of CYP1A1-related enzymes, i.e. 7-ethoxycoumarin- and 7-ethoxyresorufin-O-deethylase (ECOD and EROD) in both cell systems in basal conditions and after exposure to non-cytotoxic doses of β-naphthoflavone (BNF), a well-known PAH-type inducer. Both cell models showed basal and inducible levels of ECOD. Analysis of CYP1A1 protein expression and EROD-related enzyme activity confirmed the inducibility of the CYP1A1 isoform by BNF. These results demonstrate that mesenchymal adult stem cells can constitute innovative models. We therefore propose the use of pre-adipocytes and their neurogenic derivates to evaluate the cytotoxic/biological effects of unintended exposure to contaminants.     

Skin potential and EMG changes induced by cutaneous electrical stimulation. I. Normal man in arousing and non-arousing environment.

Skin potential and EMG responses induced in normal man by electrical stimuli applied to the skin were recorded in the four limbs in order to study somato-sympathetic and somato-motor reflexes. Different patterns of responses were observed in different conditions: alarm, habituation, sensitization and arousal. During alarm, sensitization and arousal, the responses were present in the four limbs; during habituation, the responses were only present in the stimulated and in the contralateral limb. Three sensory thresholds to cutaneous electrical stimulation were identified in habituated subjects: tactile, tingling and pain. Cutaneous and EMG responses appeared at tingling threshold. A relationship between skin potential level and skin potential response was observed.     

Therapy with bromocriptine and behavior of various hormones in psoriasis patients

The activity of bromocriptin has been clinically tested on 18 patients suffering from psoriasis. The plasma levels of Human Growth Hormone (HGH), Human Adrenocorticotropic Hormone (ACTH), Thyroid Stimulating Hormone (TSH), Prolactin (PL), Aldosterone and Cortisol were investigated in these 18 patients along with 35 untreated psoriatic patients and 19 normal subjects. Bromocriptin was shown to be effective in 13 of our 18 psoriatic patients. Plasma levels of HGH, ACTH, and Aldosterone, measured in all 53 psoriatic patients, were found to be higher than normal in 11, 26 and 40 patients respectively (HGH and Aldosterone: p less than 0,005; ACTH: p less than 0,001).     

Outdoor cultivation ofArthrospira platensis during autumn and winter in temperate climates

Arthrospira (Spirulina) platensis M2 was grown outdoors in 50-mm diameter tubular reactors under the climatic conditions of central Italy (Florence) from September to December 1995 and in March 1996. Except for September, the cultures temperature was regulated. Mean productivities of 0.83, 0.44 and 0.61 g dry wt L^–1 d^–1 were achieved in autumn (September–October), winter (November–December) and March, respectively. In autumn and winter, the photosynthetic efficiency of the cultures and the degree of correlation between productivity and solar irradiance were significantly greater than in summer. The effect of cell density and aeration rate on productivity was evaluated in September. The productivity of cultures operated at high supra-optimal population density was about 30% less at high aeration rate (1.0 LL^–1 min^–1), and 50% less at standard aeration rate (0.17 LL^–1 min^–1), than that of control cultures kept at optimal population density and standard aeration rate. The reduction of productivity in high-density cultures was due to lower daylight output rates and higher night biomass losses (the latter were particularly relevant under standard aeration conditions). The main factor limiting productivity in closed reactors during autumn was the night temperature. Heating the cultures during daylight hours on sunny days did not cause any significant increase of the yields, since under sunlight the unheated cultures also reached the optimal temperature for growth early in the morning. On cloudy days, the day-time temperature of the unheated cultures remained well below the optimum, however this had only a limited effect on productivity since algal growth was mainly light-limited.     

Organization of efferent extraspinal sympathetic pathways in Rana esculenta

Origin and distribution of pre- and postsynaptic fibres in the sympathetic trunk of Rana esculenta (from ganglion 3 to ganglion 10) have been investigated by means of extracellular recordings. Two systems conducting efferent information appear to exist: 1) a system made of faster conducting fibres (B group pre- and postsynaptic fibres); presynaptic fibres originating from a very high monosegmental source (4th spinal root); postsynaptic fibres leave the sympathetic chain plurisegmentally (rami communicantes 5-10); 2) a system made of slower conducting fibres (C group pre- and postsynaptic fibres) originating plurisegmentally from spinal roots 5-8. Intracellular recordings have shown that: 1) integrative processes take place in the amphibian sympathetic trunk, as in the mammalian one, but are quantitatively lesser. Homonomous (B-B) and heteronomous (B-C) convergence has been observed in B neurons, and also the convergence of a collateral of a C postsynaptic axon on B neurons. 2) posthumous depolarizations are present: these are events modulating the activity of sympathetic neurons. In B neurons posthumous depolarization follows orthodromic responses, and a late posthumous depolarization can be seen in B and C neurons following either ortho- or antidromic stimulation.     

As integrated culture system for outdoor production of microalgae and cyanobacteria

Cultivation of microalgae in closed photobioreactors suffers from high temperatures, whereas in open raceway ponds the optimal temperature is seldom reached. With an integrated device coupling a raceway pond with a suitably sized and positioned closed system (alveolar panel), the heat accumulated in the panel was efficiently transferred to the pond and a near-optimum temperature regimen for Arthrospira platensis was maintained, with no need for any additional cooling device. The productivity obtained in the integrated system was higher than the sum of the productivities (g reactor⁻¹ day⁻¹) of the pond and panel systems operating separately. This revised version was published online in September 2006 with corrections to the Cover Date.