Structure-function strategies to improve the pharmacological value of animal toxins.

ISBN : 978-0-12-369442-3Animal venoms are rich sources of bioactive compounds that possess obvious pharmacological, therapeutic and/or biotechnological values. A majority of these compounds are peptides that mainly target enzymes, membrane receptors or ion channels. These peptides are most often in a size range that allows their production in vitro by chemical synthesis or genetic engineering. Unfortunately, they rarely display the required characteristics in terms of selectivity, affinity, stability and targeting with regard to the desired application. In recent years, a number of structural approaches or strategies have been developed to improve the intrinsic potential of venom peptides. They are reviewed herein for their effectiveness

Expression of HLA-G in human cornea, an immune-privileged tissue.

Human leukocyte antigen (HLA)-G retains the capacity to modulate immune responses, favoring the establishment of tolerance in solid-tissue allotransplants. To better understand the mechanisms that promote corneal allograft survival, we investigated whether HLA-G was an immunoregulatory factor involved in corneal immunology. We therefore sought HLA-G expression in corneal tissues. Corneal transplantation consists in replacing the center of a diseased cornea with normal corneal tissue. Two corneal parts are not used in such surgery: diseased central corneal tissue and peripheral normal cornea. For this study, we used healthy corneas obtained from deceased donors and diseased corneas obtained from patients with pseudophakic bullous keratopathy or keratoconus who had undergone corneal transplantation. Immunohistochemical analysis carried out on the cryopreserved corneas showed a positive immunohistochemical staining with anti-HLA-G, anti-HLA-A, -B, and -C, and anti-HLA class I monoclonal antibodies. Staining was obtained for keratocytes, epithelial cells, and endothelial cells from both healthy and pathologic human corneas, revealing the presence of HLA class I proteins, including HLA-G. HLA-G transcripts were detected in normal cornea by reverse transcriptase-polymerase chain reaction with a classical pattern of alternative splicing. The detection of HLA-G protein in adult corneas leads to the conclusion that this protein may contribute to the maintenance of the privileged immune status of cornea

Cytotoxic effect on lymphocytes of Tat from human immunodeficiency virus (HIV-1)

AbstractThe human immunodeficiency virus type 1 (HIV-1) genome codes for trans-activator Tat, an 86-residue protein whose expression is critical for viral replication. Full-length Tat and Tat peptides from HIV-1 were chemically synthesized using optimized solid phase technique. Synthetic Tat2 in86, was found not only to inhibit antigen-induced human peripheral blood lymphocyte (PBL) proliferation in vitro, as described by Viscidi et al. [1989, Science 246, 1606-1608], but also mitogen-induced PBL proliferation, with 50% inhibition obtained at 0.9 and 8 μ;M, respectively. To assess the mechanism by which Tat exert its inhibitory effect, we analysed its interaction and effect on CD4+-cells. Direct fluorescence and indirect immunofluorescence assays analysed by flow cytometry showed that fluorescein isothiocyanate-labeled and -unlabeled Tat interact (>0.2 μ;M) with CD4-expressing lymphoid cells (CEM cell line). Experiments of chromium-51 release and Trypan blue exclusion on these tumor cells in vitro have demonstrated the capacity of Tat to modify cellular membrane permeability and cell viability, in a dose-dependent manner. The use of Tat peptides revealed that those containing the Tat basic region from 49 to 57 were able to bind to the cell membrane and to exhibit a cytotoxic activity on lymphocytes. Together, the data suggest that the potential cytotoxicity of Tat on lymphocytes could be directly implicated in virus-induced immune dysfunction observed in HIV-1 infected patients

Evaluation of the lethal potency of scorpion and snake venoms and comparison between intraperitoneal and intravenous injection routes.

International audienceScorpion stings and snake bites are major health hazards that lead to suffering of victims and high mortality. Thousands of injuries associated with such stings and bites of venomous animals occur every year worldwide. In North Africa, more than 100,000 scorpion stings and snake bites are reported annually. An appropriate determination of the 50% lethal doses (LD50) of scorpion and snake venoms appears to be an important step to assess (and compare) venom toxic activity. Such LD50 values are also commonly used to evaluate the neutralizing capacity of specific anti-venom batches. In the present work, we determined experimentally the LD50 values of reference scorpion and snake venoms in Swiss mice, and evaluated the influence of two main venom injection routes (i.e., intraperitoneal (IP) versus intravenous (IV)). The analysis of experimental LD50 values obtained with three collected scorpion venoms indicates that Androctonus mauretanicus (Am) is intrinsically more toxic than Androctonus australis hector (Aah) species, whereas the latter is more toxic than Buthus occitanus (Bo). Similar analysis of three representative snake venoms of the Viperidae family shows that Cerastes cerastes (Cc) is more toxic than either Bitis arietans (Ba) or Macrovipera lebetina (Ml) species. Interestingly, the venom of Elapidae cobra snake Naja haje (Nh) is far more toxic than viper venoms Cc, Ml and Ba, in agreement with the known severity of cobra-related envenomation. Also, our data showed that viper venoms are about three-times less toxic when injected IP as compared to IV, distinct from cobra venom Nh which exhibited a similar toxicity when injected IP or IV. Overall, this study clearly highlights the usefulness of procedure standardization, especially regarding the administration route, for evaluating the relative toxicity of individual animal venoms. It also evidenced a marked difference in lethal activity between venoms of cobra and vipers, which, apart from the nature of toxins, might be attributed to the rich composition of high molecular weight enzymes in the case of viper venoms

Influence of the consumption pattern of magnesium from magnesium-rich mineral water on magnesium bioavailability

It is generally considered that the absorption of Mg is inversely related to the ingested dose. The objective of the present study was to determine if the mode of administration (bolus v. consumption throughout the day) could influence Mg bioavailability from Mg-rich natural mineral water comparing the same nutritional Mg amount (126mg). Using a 2d cross-over design, twelve healthy men were asked to drink 1·5 litres Mg-rich mineral water either as 2×750ml or 7×212ml throughout the day. Two stable isotopes (25Mg and 26Mg) were used to label the water in order to distinguish both regimens. Fractional apparent Mg absorption was determined by faecal monitoring and Mg retention was determined by measuring urinary excretion of Mg isotopes. Higher Mg absorption (50·7 (sd 12·7) v. 32·4 (sd 8·1) %; P=0·0007) and retention (47·5 (sd 12·9) v. 29·0 (sd 7·5) %; P=0·0008) from Mg-rich mineral water were observed when it was consumed in seven servings compared with larger servings. Thus, regular water consumption throughout the day is an effective way to increase Mg bioavailability from Mg-rich mineral wate

Down-Regulation of ECRG4, a Candidate Tumor Suppressor Gene, in Human Breast Cancer

INTRODUCTION: ECRG4/C2ORF40 is a potential tumor suppressor gene (TSG) recently identified in esophageal carcinoma. Its expression, gene copy number and prognostic value have never been explored in breast cancer. METHODS: Using DNA microarray and array-based comparative genomic hybridization (aCGH), we examined ECRG4 mRNA expression and copy number alterations in 353 invasive breast cancer samples and normal breast (NB) samples. A meta-analysis was done on a large public retrospective gene expression dataset (n = 1,387) in search of correlations between ECRG4 expression and histo-clinical features including survival. RESULTS: ECRG4 was underexpressed in 94.3% of cancers when compared to NB. aCGH data revealed ECRG4 loss in 18% of tumors, suggesting that DNA loss is not the main mechanism of underexpression. Meta-analysis showed that ECRG4 expression was significantly higher in tumors displaying earlier stage, smaller size, negative axillary lymph node status, lower grade, and normal-like subtype. Higher expression was also associated with disease-free survival (DFS; HR = 0.84 [0.76-0.92], p = 0.0002) and overall survival (OS; HR = 0.72 [0.63-0.83], p = 5.0E-06). In multivariate analysis including the other histo-clinical prognostic features, ECRG4 expression remained the only prognostic factor for DFS and OS. CONCLUSIONS: Our data suggest that ECRG4 is a candidate TSG in breast cancer, the expression of which may help improve the prognostication. If functional analyses confirm this TSG role, restoring ECRG4 expression in the tumor may represent a promising therapeutic approach

Maurocalcine and domain A of the II-III loop of the dihydropyridine receptor Cav 1.1 subunit share common binding sites on the skeletal ryanodine receptor.

International audienceMaurocalcine is a scorpion venom toxin of 33 residues that bears a striking resemblance to the domain A of the dihydropyridine voltage-dependent calcium channel type 1.1 (Cav1.1) subunit. This domain belongs to the II-III loop of Cav1.1, which is implicated in excitation-contraction coupling. Besides the structural homology, maurocalcine also modulates RyR1 channel activity in a manner akin to a synthetic peptide of domain A. Because of these similarities, we hypothesized that maurocalcine and domain A may bind onto an identical region(s) of RyR1. Using a set of RyR1 fragments, we demonstrate that peptide A and maurocalcine bind onto two discrete RyR1 regions: fragments 3 and 7 encompassing residues 1021-1631 and 3201-3661, respectively. The binding onto fragment 7 is of greater importance and was thus further investigated. We found that the amino acid region 3351-3507 of RyR1 (fragment 7.2) is sufficient for these interactions. Proof that peptide A and maurocalcine bind onto the same site is provided by competition experiments in which binding of fragment 7.2 to peptide A is inhibited by preincubation with maurocalcine. Moreover, when expressed in COS-7 cells, RyR1 carrying a deletion of fragment 7 shows a loss of interaction with both peptide A and maurocalcine. At the functional level, this deletion abolishes the maurocalcine induced stimulation of [3H]ryanodine binding onto microsomes of transfected COS-7 cells without affecting the caffeine and ATP responses

: Maurocalcine transduction into cells

International audienceMaurocalcine (MCa) is a 33-amino-acid residue peptide toxin isolated from the scorpion Scorpio maurus palmatus. External application of MCa to cultured myotubes is known to produce Ca2+ release from intracellular stores. MCa binds directly to the skeletal muscle isoform of the ryanodine receptor, an intracellular channel target of the endoplasmic reticulum, and induces long lasting channel openings in a mode of smaller conductance. Here we investigated the way MCa proceeds to cross biological membranes to reach its target. A biotinylated derivative of MCa was produced (MCa(b)) and complexed with a fluorescent indicator (streptavidine-cyanine 3) to follow the cell penetration of the toxin. The toxin complex efficiently penetrated into various cell types without requiring metabolic energy (low temperature) or implicating an endocytosis mechanism. MCa appeared to share the same features as the so-called cell-penetrating peptides. Our results provide evidence that MCa has the ability to act as a molecular carrier and to cross cell membranes in a rapid manner (1-2 min), making this toxin the first demonstrated example of a scorpion toxin that translocates into cells

Emerging diseases of wildlife in Europe : Lessons to draw to prevent a resurgence of avian influenza

The incursion of Highly Pathogenic Avian Influenza among wildfowl in Europe in 2006 was a new illustration of the health risk presented by wildlife to humans and domestic animals. To help anticipate similar incursions and avoid the pitfalls of poor communication, this paper describes how this risk was analysed and managed in the past for other wildlife diseases in Europe. The author also proposes a general methodology to anticipate such events.La contamination de l'avifaune sauvage en Europe, en 2006, par des foyers d'Influenza Aviaire Hautement Pathogène (IAHP) a été une nouvelle illustration du risque sanitaire que représente la faune sauvage pour l'homme ou les animaux domestiques. Afin de mieux anticiper des incursions similaires et éviter les dérives liées à une communication mal conduite, cet article décrit la façon dont le risque a été étudié et géré dans le passé pour d'autres maladies de la faune sauvage en Europe. L'auteur propose aussi une méthodologie générale pour anticiper de tels événements