Murine CD93 (C1qRp) contributes to the removal of apoptotic cells in vivo but is not required for C1q-mediated enhancement of phagocytosis

Human CD93 (known as C1qRp) has been shown to be a phagocytic receptor involved in the in vitro C1q-dependent enhancement of phagocytosis. However, binding of CD93 to C1q and its function remain controversial. In this study, we have generated CD93-deficient mice (CD93(-/-)) to investigate its biological role(s). The CD93(-/-) mice were viable and showed no gross abnormalities in their development. Thioglycolate-elicited peritoneal macrophages deficient in CD93 showed a similar enhancement in complement- and FcgammaR-dependent uptake of RBC to the wild-type macrophages when plated on C1q-coated surfaces suggesting that the lack of this receptor had no effect on these C1q-mediated events. There was no impairment in either complement- or FcgammaR-dependent phagocytic assays in vivo. By contrast, the CD93(-/-) mice had a significant phagocytic defect in the clearance of apoptotic cells in vivo (human Jurkat T cells and murine thymocytes: p=0.0006 and p=0.0079, respectively) compared with strain-matched controls. However, in vitro, the CD93(-/-) macrophages showed similar engulfment of apoptotic cells to wild-type macrophages. Furthermore, no supporting evidence for a role of CD93 as an adhesion molecule was found using intravital microscopy or analyzing peritoneal cell recruitment in response to three different inflammatory stimuli (thioglycolate, zymosan A, and IL-1beta). Thus, our findings indicate that murine CD93 is expressed on the peritoneal macrophage, especially on thioglycolate-elicited cells, but does not appear to play a key role in C1q-mediated enhancement of phagocytosis or in the intercellular adhesion events tested. However, our results suggest that it may contribute to the in vivo clearance of dying cells.     

Paleoenvironmental conditions preceding the Messinian Salinity Crisis in the Central Mediterranean: Integrated data from the Upper Miocene Trave section (Italy)

Integrated data of calcareous plankton and benthic foraminifers from the pre-evaporitic interval of Trave section (Central Italy) allowed the reconstruction of surface and bottom-water conditions in the Central Mediterranean during the interval from 7.61 to 6.33 Ma, preceding the Messinian Salinity Crisis.Our data point out a three-step paleoenvironmental evolution. During the first stage (7.61-7.02 Ma) benthic foraminiferal assemblages depict stable, well-oxygenated and ventilated bottom-water conditions, while the surface water records variable temperature and high nutrient conditions, probably associated with strong seasonality. The second stage (7.02-6.70 Ma) points to unfavourable bottom-water condition, triggered by deep-sea stagnation. This is witnessed by a significant decrease in oxygen concentration and biotic diversity, and by the presence of stress-tolerant taxa. A general warming of the surface water and a strongly stratified water column, characterized by an expanded mixed layer, are also recorded.From 6.70 Ma onwards (third stage), a prominent change to more restricted, low-oxygenated, hypersaline conditions at the sea floor is testified by the total disappearance of deep-dwelling planktonic foraminifers and the increasing abundance of stress-tolerant species. Calcareous plankton reflects high instability of the surface water in terms of nutrients, temperature and salinity. During this stage the environmental deterioration reaches intermediate depths in the water column.The initial change toward a step-wise isolation of the Central Mediterranean bottom-waters is probably related to a general warming, responsible for a first slowing-down of the vertical circulation, favouring stratification of surface and intermediate waters and stagnation of bottom-waters. This warming is related to the restricted connection between the Mediterranean Sea and the Atlantic Ocean, which occurred since 7.146 Ma.In the Trave section, the isolation of bottom-waters most likely occurred at the same time as in other Mediterranean sections. However, due to the presence of a hiatus it cannot be excluded that it occurred with a delay of ~ 100 kyr, probably related to the shallower paleodepth of the basin.     

Cytoskeletal mechanics in adherent human airway smooth muscle cells: probe specificity and scaling of protein-protein dynamics

We probed elastic and loss moduli in the adherent human airway smooth muscle cell through a variety of receptor systems, each serving as a different molecular window on cytoskeletal dynamics. Coated magnetic microbeads were attached to the cell surface via coating-receptor binding. A panel of bead coatings was investigated: a peptide containing the sequence RGD, vitronectin, urokinase, activating antibody against ₁-integrin, nonactivating antibody against ₁-integrin, blocking antibody against ₁-integrin, antibody against ₁-integrin, and acetylated low-density lipoprotein. An oscillatory mechanical torque was applied to the bead, and resulting lateral displacements were measured at baseline, after actin disruption by cytochalasin D, or after contractile activation by histamine. As expected, mechanical moduli depended strongly on bead type and bead coating, differing at the extremes by as much as two orders of magnitude. In every case, however, elastic and loss moduli increased with frequency f as a weak power law, f ^x–1. Moreover, with few exceptions, data could be scaled such that elastic and frictional responses depended solely on the power law exponent x. Taken together, these data suggest that power law behavior represents a generic feature of underlying protein-protein dynamics. actin; cytoskeleton; magnetic twisting cytometry; scale free; viscoelasticity     

Characterization,Toxicity, and Optimization for the Growth and Production of Bacteriocin-like Substances by Lactobacillus curvatus

Probiotics and Antimicrobial Proteins - This study aimed to characterize, evaluate toxicity and optimize the conditions for the growth and production of bacteriocin-like substances by Lactobacillus...     

Complement C1q activates canonical Wnt signaling and promotes aging-related phenotypes

Wnt signaling plays critical roles in development of various organs and pathogenesis of many diseases, and augmented Wnt signaling has recently been implicated in mammalian aging and aging-related phenotypes. We here report that complement C1q activates canonical Wnt signaling and promotes aging-associated decline in tissue regeneration. Serum C1q concentration is increased with aging, and Wnt signaling activity is augmented during aging in the serum and in multiple tissues of wild-type mice, but not in those of C1qa-deficient mice. C1q activates canonical Wnt signaling by binding to Frizzled receptors and subsequently inducing C1s-dependent cleavage of the ectodomain of Wnt coreceptor low-density lipoprotein receptor-related protein 6. Skeletal muscle regeneration in young mice is inhibited by exogenous C1q treatment, whereas aging-associated impairment of muscle regeneration is restored by C1s inhibition or C1qa gene disruption. Our findings therefore suggest the unexpected role of complement C1q in Wnt signal transduction and modulation of mammalian aging.     

Fine structure of leydig and sertoli cells in the testis of immature and mature spotted ray Torpedo marmorata

An ultrastructural investigation revealed the presence of true Leydig cells in the testis of sexually mature specimens of Torpedo marmorata. They showed the typical organization of steroid-hormone-producing cells, which, however, changed as spermatocysts approached maturity. In fact, they appeared as active cells among spermatocysts engaged in spermatogenesis, while in regions where spermiation occurred, they progressively regressed resuming the fibroblastic organization typically present in the testis of immature specimens. Such observations strongly suggest that these cells might be engaged in steroidogenesis and actively control spermatogenesis. Sertoli cells, too, appeared to play a role in spermatogenesis control, since, like Leydig cells, they showed the typical aspect of steroidogenic cells. In addition, the presence of gap junctions between Sertoli cells suggests that their activity might be coordinated. After sperm release, most Sertoli cells were modified and, finally, degenerated, but few of them changed into round cells (cytoplasts) or round cell remnants, which continued their steroidogenic activity within the spermatocyst and the genital duct lumen. From the present observations, it can be reasonably concluded that, in T. marmorata, spermatogenesis depends on both Leydig and Sertoli cells, and, as postulated by Callard (1991), in cartilaginous fish, the function of the Leydig cells as producers of steroids might be more recent and subsequent to that of Sertoli cells. In this regard, it is noteworthy that, in immature males, when Leydig cells showed a fibroblastic organization, Sertoli cells already displayed the typical organization of a steroidogenic cell.     

Molecular Support for the Placement of Saiga and Procapra in Antilopinae (Artiodactyla, Bovidae)

The evolutionary history of the bovid subfamily Antilopinae is unclear. Traditionally, this subfamily is subdivided into two tribes: Neotragini (dwarf antelopes) and Antilopini (gazelles and their relatives). Here, we report new sequences for the 12S and 16S rRNA genes in the enigmatic antilopine taxa Procapra gutturosa and Saiga tatarica and analyze the phylogenetic relationships of these taxa relative to other antilopines. Our study demonstrates the close affinity of the saiga antelope to Gazella despite the conventional systematic allocation of Saiga to the Caprinae subfamily. The second member of the Saigini tribe, Pantholops hodgsoni (Tibetan gazelle), falls within Caprinae. In all of our analyses, Procapra gutturosa occupied a basal position in the Antilopinae clade or was a sister-group to the dwarf antelope Madoqua. This suggests early separation of Procapra from other antelopes.     

STAT1 regulates lipopolysaccharide- and TNF-alpha-dependent expression of transporter associated with antigen processing 1 and low molecular mass polypeptide 2 genes in macrophages by distinct mechanisms

Transporter associated with Ag processing 1 and low molecular mass polypeptide 2 (LMP2) are essential for class I MHC function and share a common bidirectional promoter. In murine bone marrow-derived macrophages, LPS and TNF-alpha induced Tap1 and up-regulated Lmp2, which is constitutively expressed at low levels. These two genes are induced by LPS and TNF-alpha with distinct kinetics, at 6 and 12-24 h, respectively. Using macrophages derived from the TNF-alpha receptors of knockout mice, we found that induction by LPS is not due to the autocrine production of TNF-alpha. In macrophages from STAT-1 knockout mice, neither LPS nor TNF-alpha induced the expression of Tap1 or Lmp2. The shared promoter contains several areas that can be controlled by STAT-1, such as the proximal and distal IFN-gamma activation site (GAS) boxes in the direction of the Tap1 gene. By making deletions of the promoter, we determined that only the proximal GAS box is required for LPS induction of Tap1 and Lmp2. In contrast, TNF-alpha induction of these two genes is dependent on the IFN regulatory factor-1 and NF-kappaB boxes, and not on the GAS box. Our experiments using gel shift analysis and Abs indicated that STAT1 binds to the GAS box in nuclear extracts from LPS-treated macrophages. The nuclear extracts obtained from macrophages treated with TNF-alpha bound to the IFN regulatory factor-1 and NF-kappaB boxes. These results show that LPS and TNF-alpha regulate the induction of Tap1 and Lmp2 through STAT1, but use distinct areas of the promoter.     

Near infrared spectroscopy as a tool for the determination of eumelanin in human hair

Eumelanins are brown-black pigments present in the hair and in the epidermis which are acknowledged as protection factors against cell damage caused by ultraviolet radiation. The quantity of eumelanin present in hair has recently been put forward as a means of identifying subjects with a higher risk of skin tumours. For epidemiological studies, chromatographic methods of determining pyrrole-2,3,5-tricarboxylic acid (PTCA; the principal marker of eumelanin) are long, laborious and unsuitable for screening large populations. We suggest near infrared (NIR) spectroscopy as an alternative method of analysing eumelanin in hair samples. PCTA was determined on 93 samples of hair by means of oxidizing with hydrogen peroxide in a basic environment followed by chromatographic separation. The same 93 samples were then subjected to NIR spectrophotometric analysis. The spectra were obtained in reflectance mode on hair samples which had not undergone any preliminary treatment, but had simply been pressed and placed on the measuring window of the spectrophotometer. The PTCA values obtained by means of HPLC were correlated with the near infrared spectrum of the respective samples. A correlation between the PTCA values obtained by means of HPLC and the PTCA values obtained from an analysis of the spectra was obtained using the principal component regression (PCR) algorithm. The correlation obtained has a coefficient of regression (R(2)) of 0.89 and a standard error of prediction (SEP) of 13.8 for a mean value of 108.6 ng PTCA/mg hair. Some considerations about the accuracy of the obtained correlation and the main sources of error are made and some validation results are shown.     

Concentrative uptake of cyclic ADP-ribose generated by BST-1+ stroma stimulates proliferation of human hematopoietic progenitors

Cyclic ADP-ribose (cADPR) is an intracellular calcium mobilizer generated from NAD(+) by the ADP-ribosyl cyclases CD38 and BST-1. cADPR, both exogenously added and paracrinally produced by a CD38(+) feeder layer, has recently been demonstrated to stimulate the in vitro proliferation of human hemopoietic progenitors (HP) and also the in vivo expansion of hemopoietic stem cells. The low density of BST-1 expression on bone marrow (BM) stromal cells and the low specific activity of the enzyme made it unclear whether cADPR generation by a BST-1(+) stroma could stimulate HP proliferation in the BM microenvironment. We developed and characterized two BST-1(+) stromal cell lines, expressing an ectocellular cyclase activity similar to that of BST-1(+) human mesenchymal stem cells, the precursors of BM stromal cells. Long term co-culture of cord blood-derived HP over these BST-1(+) feeders determined their expansion. Influx of paracrinally generated cADPR into clonogenic HP was mediated by a concentrative, nitrobenzylthioinosine- and dipyridamole-inhibitable nucleoside transporter, this providing a possible explanation to the effectiveness of the hormone-like concentrations of the cyclic nucleotide measured in the medium conditioned by BST-1(+) feeders. These results suggest that the BST-1-catalyzed generation of extracellular cADPR, followed by the concentrative uptake of the cyclic nucleotide by HP, may be physiologically relevant in normal hemopoiesis.