Ethoxyformylation of histidine residues in equine growth hormone

Reactivity of histidine residues in equine growth hormone to ethoxyformic anhydride was studied. The existence of two kinetically different sets was demonstrated: one of them including only the slow reacting histidine 169 (k = 0.164 min-1) and the other containing fast reacting histidines 19 and 21 (k = 0.892 min-1). A correlation between the decrease in the capacity to compete with 125I-labeled hormone for rat liver binding sites and the degree of ethoxyformylation of the fast group was found. Circular dichroism studies indicated no significant conformational changes in the protein with all three residues modified. These results fully agree with those obtained for bovine growth hormone which is further evidence supporting the vinculation of histidines 19 and/or 21 with the binding site of these hormones to their specific receptors.     

Tag loss is a minor limiting factor in sea turtle tagging programs relying on distant tag returns: the case of Mediterranean loggerhead sea turtles

As in many other species, tagging has been routinely conducted for decades in over a hundred sea turtle capture-mark-recapture (CMR) programs worldwide. Tag loss is a key limiting factor because it violates the main assumption in CMR models; however, very few estimates of tag loss exist, and we provide here a review. No published estimations of tag loss are available for the Mediterranean, in spite of intensive tagging since the 1980s. This study aims to provide an estimation of tag loss in loggerhead turtles tagged in the Mediterranean. We modeled 64 tag returns out of ca. 2200 loggerhead turtles tagged at Mediterranean foraging grounds, with mark-recapture intervals up to 11.5 years, in order to estimate a daily tag loss probability of the most used tag applied to the most common turtle species in the region. Five models were evaluated through maximum likelihood estimation. The model with the best fit described a tag loss initially high and then decreasing to a lower asymptote, which is probably due to some defective tag applications. The resulting tag loss (0.15 in the first year and 0.31 after 5 years) was comparable or even lower than those from other areas and/or species and predictions indicate that double tagging can make a turtle identifiable for a long period. Hence, in our tagging program and probably in similar ones as well, tag loss appears to be the less important of the factors affecting tag returns, and efforts in other directions are more likely to improve CMR results.     

CARD14/CARMA2sh and TANK differentially regulate poly(I:C)–induced inflammatory reaction in keratinocytes

CARD14/CARMA2sh (CARMA2sh) is a scaffold protein whose mutations are associated with the onset of human genetic psoriasis and other inflammatory skin disorders. Here we show that the immunomodulatory adapter protein TRAF family member-associated NF-κB activator (TANK) forms a complex with CARMA2sh and MALT1 in a human keratinocytic cell line. We also show that CARMA2 and TANK are individually required to activate the nuclear factor κB (NF-κB) response following exposure to polyinosinic-polycytidylic (poly [I:C]), an agonist of toll-like receptor 3. Finally, we present data indicating that TANK is essential for activation of the TBK1/IRF3 pathway following poly (I:C) stimulation, whereas CARMA2sh functions as a repressor of it. More important, we report that two CARMA2sh mutants associated with psoriasis bind less efficiently to TANK and are therefore less effective in suppressing the TBK1/IRF3 pathway. Overall, our data indicate that TANK and CARMA2sh regulate TLR3 signaling in human keratinocytes, which could play a role in the pathophysiology of psoriasis.     

Environment, inflammation, and cancer

Rudolf Virchow postulated that a critical feature of tumors was the presence of leukocytes, providing the first indication that inflammation may play a role in tumorigenesis. We now have a wealth of experimental and clinical data demonstrating a clear relationship between inflammatory responses and the roles they play at different stages of tumor development. The details of the dynamic relationship between tumor cells and specific subtypes of immune cells and mesenchymal cells are being revealed as critical to cancer progression which has led to the development of potential new targets for cancer treatment. This review describes some of the key molecular and cellular events demonstrating the critical role of inflammation on promoting tumorigenesis with attention on novel therapeutics and their potential clinical success.     

Peroxisomes as novel players in cell calcium homeostasis

Ca2+ concentration in peroxisomal matrix ([Ca2+](perox)) has been monitored dynamically in mammalian cells expressing variants of Ca2+-sensitive aequorin specifically targeted to peroxisomes. Upon stimulation with agonists that induce Ca2+ release from intracellular stores, peroxisomes transiently take up Ca2+ reaching peak values in the lumen as high as 50-100 microm, depending on cell types. Also in resting cells, peroxisomes sustain a Ca2+ gradient, [Ca2+](perox) being approximately 20-fold higher than [Ca2+] in the cytosol ([Ca2+](cyt)). The properties of Ca2+ traffic across the peroxisomal membrane are different from those reported for other subcellular organelles. The sensitivity of peroxisomal Ca2+ uptake to agents dissipating H+ and Na+ gradients unravels the existence of a complex bioenergetic framework including V-ATPase, Ca2+/H+, and Ca2+/Na+ activities whose components are yet to be identified at a molecular level. The different [Ca2+](perox) of resting and stimulated cells suggest that Ca2+ could play an important role in the regulation of peroxisomal metabolism.     

Fresh-frozen human bone graft to repair defect after mandibular giant follicular cyst removal: a case report

The purpose of this article is to report the clinical, radiographical and histological findings about a case of a young woman affected by a mandibular giant follicular cyst. Conservative tumor resection was followed by immediate reconstructive treatment using fresh frozen human bone graft, instead of autologous bone graft, as material for bone regeneration. Follicular cyst is a benign, non invasive lesion with slow but progressive growth. Radiological and histological examination of the lesion confirmed the presence of a follicular cyst which underwent biopsy evaluation before enucleation. According to literature, conservative treatment was performed with optimal prognosis. At 24?months post surgery no evidence of recurrency was objective; the CT scan revealed optimal bone formation inside the reconstructed site. The use of fresh frozen bone allograft, thanks to its osteoinductive and osteoconductive properties, can be a safe choice for reconstruction of bone defects after jaw cysts removal.     

Isolation and characterization of <Emphasis Type="Italic">Clostridium difficile</Emphasis> from shellfish and marine environments

This pilot study was carried out to evaluate the occurrence of Clostridium difficile in marine environments and in edible shellfish. Samples of seawater, sediment, and zooplankton were collected at five sampling stations in the Gulf of Naples. Six samples of edible shellfish, furthermore, were obtained: two from mussel farms and four from wholesalers. The isolation and the characterization of C. difficile strains were carried out using selective media and molecular techniques, respectively. C. difficile was isolated from nine of the 21 samples investigated. Shellfish and zooplankton showed the highest prevalence of positive samples. No C. difficile was detected in marine sediment. Majority of the C. difficile isolates were toxin A/B positive. Six known different PCR ribotypes (003, 005, 009, 010, 056, and 066) were identified, whereas one strain may represent a new PCR ribotype. C. difficile may be present in the marine environment in Southern Italy, including shellfish and zooplankton. This study is reporting the isolation of C. difficile from zooplankton, clams, and mussels and pointing out a new possible route to exposure to C. difficile of healthy individuals in the community.     

In vivo tyrosine phosphorylation sites of activated ephrin-B1 and ephB2 from neural tissue

EphB2 is a receptor tyrosine kinase of the Eph family and ephrin-B1 is one of its transmembrane ligands. In the embryo, EphB2 and ephrin-B1 participate in neuronal axon guidance, neural crest cell migration, the formation of blood vessels, and the development of facial structures and the inner ear. Interestingly, EphB2 and ephrin-B1 can both signal through their cytoplasmic domains and become tyrosine-phosphorylated when bound to each other. Tyrosine phosphorylation regulates EphB2 signaling and likely also ephrin-B1 signaling. Embryonic retina is a tissue that highly expresses both ephrin-B1 and EphB2. Although the expression patterns of EphB2 and ephrin-B1 in the retina are different, they partially overlap, and both proteins are substantially tyrosine-phosphorylated. To understand the role of ephrin-B1 phosphorylation, we have identified three tyrosines of ephrin-B1 as in vivo phosphorylation sites in transfected 293 cells stimulated with soluble EphB2 by using mass spectrometry and site-directed mutagenesis. These tyrosines are also physiologically phosphorylated in the embryonic retina, although the extent of phosphorylation at each site may differ. Furthermore, many of the tyrosines of EphB2 previously identified as phosphorylation sites in 293 cells (Kalo, M. S., and Pasquale, E. B. (1999) Biochemistry 38, 14396-14408) are also phosphorylated in retinal tissue. Our data underline the complexity of ephrin-Eph bidirectional signaling by implicating many tyrosine phosphorylation sites of the ligand-receptor complex.