Human kallikrein 13 expression in normal tissues: an immunohistochemical study.

The human tissue kallikrein 13 gene (KLK13), encoding for hK13 protein, was recently cloned and characterized. Here we describe the immunohistochemical (IHC) localization of hK13 in normal human tissues and compare it with the expression of two other kallikreins, hK6 and hK10. We performed the streptavidin-biotin IHC method on 204 paraffin blocks from archival, current, and autopsy material prepared from almost every normal human tissue, using a polyclonal and a monoclonal hK13 antibody. The staining was cytoplasmic and both antibodies yielded similar results. The hK13 protein was revealed in a variety of tissues, mainly in glandular epithelia. Other epithelia that expressed hK13 included the urothelium, the spermatic epithelium, and the epithelium of the choroid plexus. hK13 was intensely immunoexpressed by some endocrine organs, such as the adenohypophysis, the thyroid gland, the parathyroid glands, the adrenal medulla, the Leydig cells of the testis, and the cells of the endocrine pancreas. Immunoreactivity was also observed in the primordial follicles, the corpus luteum, and sparse luteinized cells in the stroma of the ovary, the trophoblastic cells of the placenta, the Hassall's corpuscles of the thymus, and chondrocytes. Nerves and ganglia of the peripheral nervous system, and both neurons and glial cells in the central nervous system, were positive. In short, hK13 was expressed by many glandular epithelia, some endocrine organs, and some specialized epithelia and cells. Comparison of these data with hK6 and hK10 expression suggests that the three kallikreins have a similar IHC pattern in normal human tissues.     

Synthesis and free radical scavenging activity of some new spiropyranocoumarins

A series of novel spiro-substituted 4-hydroxypyranocoumarins and their corresponding dihydropyrano cis-diols has been synthesized. Among them the spiroadamantylpyranocoumarin and the diols can interact with the stable free radical 1,1-diphenyl-2-picrylhydrazyl and scavenge superoxide anions generated in the xanthine-xanthine oxidase system.     

Microfacies and cycle stacking pattern in Liassic peritidal carbonate platform strata, Gavrovo-Tripolitza platform, Peloponnesus, Greece

Platform carbonate sediments of Liassic age cropping out in the area of the Pigadi-Fokianos Gulf (SE of Leonidion, Peloponnesus) have been investigated in order to determine their depositional environment. Facies analysis allowed the recognition of several microfacies types and their cyclic stacking pattern. The carbonates were deposited in a restricted inner platform environment (lagoon-peritidal domain) and are arranged into small-scale shallowing-upward cycles. Palaeosol horizons containing typical pedogenic features are developed on the top of the peritidal facies or are directly superimposed on subtidal deposits, forming diagenetic caps. This implies repeated sea-level fluctuations and periodic emersion episodes. The presence of orbitally forced cyclicity though is mostly probable, cannot be clearly documented by the available data. The studied carbonates are comparable with other coeval analogous peritidal cycles of the same age along the southern margin of the Tethys.     

Mechanisms and consequences of agonist-induced talin recruitment to platelet integrin alphaIIbbeta3

Platelet aggregation requires agonist-induced alphaIIbbeta3 activation, a process mediated by Rap1 and talin. To study mechanisms, we engineered alphaIIbbeta3 Chinese hamster ovary (CHO) cells to conditionally express talin and protease-activated receptor (PAR) thrombin receptors. Human PAR1 or murine PAR4 stimulation activates alphaIIbbeta3, which was measured with antibody PAC-1, indicating complete pathway reconstitution. Knockdown of Rap1-guanosine triphosphate-interacting adaptor molecule (RIAM), a Rap1 effector, blocks this response. In living cells, RIAM overexpression stimulates and RIAM knockdown blocks talin recruitment to alphaIIbbeta3, which is monitored by bimolecular fluorescence complementation. Mutations in talin or beta3 that disrupt their mutual interaction block both talin recruitment and alphaIIbbeta3 activation. However, one talin mutant (L325R) is recruited to alphaIIbbeta3 but cannot activate it. In platelets, RIAM localizes to filopodia and lamellipodia, and, in megakaryocytes, RIAM knockdown blocks PAR4-mediated alphaIIbbeta3 activation. The RIAM-related protein lamellipodin promotes talin recruitment and alphaIIbbeta3 activity in CHO cells but is not expressed in megakaryocytes or platelets. Thus, talin recruitment to alphaIIbbeta3 by RIAM mediates agonist-induced alphaIIbbeta3 activation, with implications for hemostasis and thrombosis.     

Spatial organization of the glucosinolate-myrosinase system in brassica specialist aphids is similar to that of the host plant.

Secondary metabolites are important in plant defence against pests and diseases. Similarly, insects can use plant secondary metabolites in defence and, in some cases, synthesize their own products. The paper describes how two specialist brassica feeders, Brevicoryne brassicae (cabbage aphid) and Lipaphis erysimi (turnip aphid) can sequester glucosinolates (thioglucosides) from their host plants, yet avoid the generation of toxic degradation products by compartmentalizing myrosinase (thioglucosidase) into crystalline microbodies. We propose that death, or damage, to the insect by predators or disease causes disruption of compartmentalized myrosinase, which results in the release of isothiocyanate that acts as a synergist for the alarm pheromone E-beta-farnesene.     

Kinetics of Methyl t-Butyl Ether Cometabolism at Low Concentrations by Pure Cultures of Butane-Degrading Bacteria

Butane-oxidizing Arthrobacter (ATCC 27778) bacteria were shown to degrade low concentrations of methyl t-butyl ether (MTBE; range, 100 to 800 μg/liter) with an apparent half-saturation concentration (K_s) of 2.14 mg/liter and a maximum substrate utilization rate (k_c) of 0.43 mg/mg of total suspended solids per day. Arthrobacter bacteria demonstrated MTBE degradation activity when grown on butane but not when grown on glucose, butanol, or tryptose phosphate broth. The presence of butane, tert-butyl alcohol, or acetylene had a negative impact on the MTBE degradation rate. Neither Methylosinus trichosporium OB3b nor Streptomyces griseus was able to cometabolize MTBE.     

A G1‐like state allows HIV‐1 to bypass SAMHD1 restriction in macrophages

An unresolved question is how HIV‐1 achieves efficient replication in terminally differentiated macrophages despite the restriction factor SAMHD1. We reveal inducible changes in expression of cell cycle‐associated proteins including MCM2 and cyclins A, E, D1/D3 in macrophages, without evidence for DNA synthesis or mitosis. These changes are induced by activation of the Raf/MEK/ERK kinase cascade, culminating in upregulation of CDK1 with subsequent SAMHD1 T592 phosphorylation and deactivation of its antiviral activity. HIV infection is limited to these G1‐like phase macrophages at the single‐cell level. Depletion of SAMHD1 in macrophages decouples the association between infection and expression of cell cycle‐associated proteins, with terminally differentiated macrophages becoming highly susceptible to HIV‐1. We observe both embryo‐derived and monocyte‐derived tissue‐resident macrophages in a G1‐like phase at frequencies approaching 20%, suggesting how macrophages sustain HIV‐1 replication in vivo. Finally, we reveal a SAMHD1‐dependent antiretroviral activity of histone deacetylase inhibitors acting via p53 activation. These data provide a basis for host‐directed therapeutic approaches aimed at limiting HIV‐1 burden in macrophages that may contribute to curative interventions.     

Identification of novel cystinuria mutations and polymorphisms in SLC3A1 and SLC7A9 genes: absence of SLC7A10 gene mutations in cystinuric patients

Cystinuria represents 3% of nephrolithiasis in humans with an overall prevalence of 1 in 7,000 neonates. Two genes have been reported to account for the genetic basis of cystinuria, the SLC3A1 and the SLC7A9. Recently, the possible involvement of the SLC7A10 gene in the genetic basis of the disorder was also reported. In the present study, we found a total of 15 mutations in 20 Greek cystinuric patients. Eight mutations are novel, 4 in the SLC3A1: F266S, T351I, R456C, and N516D, and 4 in the SLC7A9: 479-1G>C, Y232C, D233E, and 1399+1G>T. Furthermore, 2 polymorphisms were identified in the SLC3A1 gene and 16 polymorphic variants were also found in the SLC7A9 gene of which the 235+18C>A, 604+10G>A, and 604+24T>C are novel. Finally, no mutation was found in the SLC7A10 gene in all patients. Only, the novel 634+8C>G and the previously reported 913-11C+T polymorphisms were identified in the SLC7A10 gene. In conclusion, a spectrum of SLC3A1 and SLC7A9 mutations are responsible for the genetic basis of cystinuria in Greek patients.     

Synthesis, characterization, and evaluation as transfection reagents of double-hydrophilic star copolymers: effect of star architecture

Five star polymers of the ionizable hydrophilic 2-(dimethylamino)ethyl methacrylate (DMAEMA) and the nonionic hydrophilic methoxy hexa(ethylene glycol) methacrylate (HEGMA) were prepared by group transfer polymerization (GTP) using ethylene glycol dimethacrylate (EGDMA) as coupling agent. In particular, four isomeric star copolymers, one heteroarm, two star block and one statistical star, with 90% mol DMAEMA and 10% mol HEGMA, plus one star homopolymer of DMAEMA with degrees of polymerization of the arms equal to 20 were synthesized. The polymers were characterized in terms of their molar masses (MMs) and compositions using gel permeation chromatography (GPC) and proton nuclear magnetic resonance (1H NMR) spectroscopy, respectively. The hydrodynamic diameters in water indicated some aggregation for all the star polymers except for the statistical copolymer star, while the pK values of the DMAEMA units were around 7 for all star polymers. All the star polymers were evaluated for their ability to transfect human cervical HeLa cancer cells with the modified plasmid pRLSV40 bearing the enhanced green fluorescent protein (EGFP) as the reporter gene. All four star copolymers showed decreased toxicity compared to that of the DMAEMA star homopolymer for the same amounts of star polymer tested. The star block copolymer with outer DMAEMA blocks exhibited the highest overall transfection efficiency, 11%, compared to that of all the star polymers examined in this study. This efficiency was the same as that of the commercially available transfection reagent SuperFect.     

Butterfly,spider, and plant communities in different land-use types in Sardinia,Italy

Butterfly, spider, and plant species richness and diversity were investigated in five different land-use types in Sardinia. In 16 one-hectare plots we measured a set of 15 environmental variables to detect the most important factors determining patterns of variation in species richness, particularly endemicity. The studied land-use types encompassed homogeneous and heterogeneous shrublands, shrublands with tree-overstorey, Quercus forest and agricultural land. A total of 30 butterfly species, among which 10 endemics, and 50 spider (morpho)species, were recorded. Butterfly and spider community composition differed according to land-use type. The main environmental factors determining diversity patterns in butterflies were the presence of flowers and trees. Spiders reacted mainly to habitat heterogeneity and land-use type. Traditional land-use did not have adverse effects on the diversity of butterflies, spiders, or plants. The number of endemic butterfly species per treatment increased with total species richness and altitude. Butterfly and spider richness did not co-vary across the five land-use types. Butterflies were, however, positively associated with plant species richness and elevation, whereas spiders were not. Conclusively, butterflies did not appear to be good indicators for spider diversity and species richness at the studied sites.