Pseudomonas aeruginosa delays Kupffer cell death via stabilization of the X-chromosome-linked inhibitor of apoptosis protein

Kupffer cells are important for bacterial clearance and cytokine production during infection. We have previously shown that severe infection with Pseudomonas aeruginosa ultimately results in loss of Kupffer cells and hepatic bacterial clearance. This was associated with prolonged hepatic inflammation. However, there is a period of time during which there is both preserved hepatic bacterial clearance and increased circulating TNF-alpha. We hypothesized that early during infection, Kupffer cells are protected against TNF-alpha-induced cell death via activation of survival pathways. KC13-2 cells (a clonal Kupffer cell line) were treated with P. aeruginosa (strain PA103), TNF-alpha, or both. At early time points, TNF-alpha induced caspase-mediated cell death, but PA103 did not. When we combined the two exposures, PA103 protected KC13-2 cells from TNF-alpha-induced cell death. PA103, in the setting of TNF exposure, stabilized the X-chromosome-linked inhibitor of apoptosis protein (XIAP). Stabilization of XIAP can occur via PI3K and Akt. We found that PA103 activated Akt and that pretreatment with the PI3K inhibitor, LY294002, prevented PA103-induced protection against TNF-alpha-induced cell death. The effects of LY294002 included decreased levels of XIAP and increased amounts of cleaved caspase-3. Overexpression of Akt mimicked the effects of PA103 by protecting cells from TNF-alpha-induced cell death and XIAP cleavage. Transfection with a stable, nondegradable XIAP mutant also protected cells against TNF-alpha-induced cell death. These studies demonstrate that P. aeruginosa delays TNF-alpha-induced Kupffer cell death via stabilization of XIAP.     

Non-contiguous finished genome sequence and description of Oceanobacillus massiliensis sp. nov.

Oceanobacillus massiliensis strain N’Diop^T sp. nov. is the type strain of O. massiliensis sp. nov., a new species within the genus Oceanobacillus. This strain, whose genome is described here, was isolated from the fecal flora of a healthy patient. O. massiliensis is an aerobic rod. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 3,532,675 bp long genome contains 3,519 protein-coding genes and 72 RNA genes, including between 6 and 8 rRNA operons.     

Late steps of ribosome assembly in E. coli are sensitive to a severe heat stress but are assisted by the HSP70 chaperone machine

The late stages of 30S and 50S ribosomal subunits biogenesis have been studied in a wild-type (wt) strain of Escherichia coli (MC4100) subjected to a severe heat stress (45-46°C). The 32S and 45S ribosomal particles (precursors to 50S subunits) and 21S ribosomal particles (precursors to 30S subunits) accumulate under these conditions. They are authentic precursors, not degraded or dead-end particles. The 21S particles are shown, by way of a modified 3'5' RACE procedure, to contain 16S rRNA unprocessed, or processed at its 5' end, and not at the 3' end. This implies that maturation of 16S rRNA is ordered and starts at its 5'-terminus, and that the 3'-terminus is trimmed at a later step. This observation is not limited to heat stress conditions, but it also can be verified in bacteria growing at a normal temperature (30°C), supporting the idea that this is the general pathway. Assembly defects at very high temperature are partially compensated by plasmid-driven overexpression of the DnaK/DnaJ chaperones. The ribosome assembly pattern in wt bacteria under a severe heat stress is therefore reminiscent of that observed at lower temperatures in E. coli mutants lacking the chaperones DnaK or DnaJ.     

Differential sensitivity of two insect GABA-gated chloride channels to dieldrin,fipronil and picrotoxinin

In the central nervous system of both vertebrates and invertebrates inhibitory neurotransmission is mainly achieved through activation of γ-aminobutyric acid (GABA) receptors. Extensive studies have established the structural and pharmacological properties of vertebrate GABA receptors. Although the vast majority of insect GABA-sensitive responses share some properties with vertebrate GABAA receptors, peculiar pharmacological properties of these receptors led us to think that several GABA-gated chloride channels are present in insects. We describe here the pharmacological properties of two GABA receptor subtypes coupled to a chloride channel on dorsal unpaired median (DUM) neurones of the adult male cockroach. Long applications of GABA induce a large biphasic hyperpolarization, consisting of an initial transient hyperpolarization followed by a slow phase of hyperpolarization that is not quickly desensitized. With GABA, the transient hyperpolarization is sensitive to picrotoxinin, fipronil and dieldrin whereas the slow response is insensitive to these insecticides.When GABA is replaced by muscimol and cis-4-aminocrotonic acid (CACA) a biphasic hyperpolarization consisting of an initial transient hyperpolarization followed by a sustained phase is evoked which is blocked by picrotoxinin and fipronil. Exposure to dieldrin decreases only the early phase of the muscimol and CACA-induced biphasic response, suggesting that two GABA-gated chloride channel receptor subtypes are present in DUM neurones. This study describes, for the first time, a dieldrin resistant component different to the dieldrin- and picrotoxinin-resistant receptor found in several insect species.     

Spectroscopic investigations of U(VI) species sorbed by the green algae Chlorella vulgaris

The green alga Chlorella vulgaris has the ability to bind high amounts of uranium(VI) in the pH range from 3 to 6. At pH 3 up to 40% of the uranium are bound by the algal cells. The uranium removal is almost complete at pH 5 and 6 under the given experimental conditions. Scanning electron microscopy and laser-induced fluorescence spectroscopy were used to characterize uranyl species formed in the selected pH range. The micrographs show a regular distribution of U(VI) on the cell surface. Fluorescence spectroscopic investigations of formed algal uranyl complexes indicate that the binding of U(VI) to carboxyl groups plays a dominating role at pH 3, whereas a minor impact of organic phosphate compounds on the U(VI) sorption cannot be excluded. In contrast, at pH 5 and 6 the phosphate groups are mainly responsible for the removal and binding of U(VI) by formation of organic and/or inorganic uranyl phosphates.     

Inflammatory signals enhance piezo2-mediated mechanosensitive currents

Heightened nociceptor function caused by inflammatory mediators such as bradykinin (BK) contributes to increased pain sensitivity (hyperalgesia) to noxious mechanical and thermal stimuli. Although it is known that sensitization of the heat transducer TRPV1 largely subserves thermal hyperalgesia, the cellular mechanisms underlying mechanical hyperalgesia have been elusive. The role of the mechanically activated (MA) channel piezo2 (known as FAM38B) present in mammalian sensory neurons is unknown. We test the hypothesis that piezo2 activity is enhanced by BK, an algogenic peptide that induces mechanical hyperalgesia within minutes. Piezo2 current amplitude is increased and inactivation is slowed by bradykinin receptor beta 2 (BDKRB2) activation in heterologous expression systems. Protein kinase A (PKA) and protein kinase C (PKC) agonists enhance piezo2 activity. BDKRB2-mediated effects are abolished by PKA and PKC inhibitors. Finally, piezo2-dependent MA currents in a class of native sensory neurons are enhanced 8-fold by BK via PKA and PKC. Thus, piezo2 sensitization may contribute to PKA- and PKC-mediated mechanical hyperalgesia.     

Caffeine or melatonin effects on sleep and sleepiness after rapid eastward transmeridian travel.

We measured the effects of slow-release caffeine (SRC) and melatonin (Mlt) on sleep and daytime sleepiness after a seven-time zone eastbound flight. In a double-blind, randomized, placebo-controlled study, each of three groups of nine subjects was given either 300 mg SRC on recovery day 1 (D1) to D5 (0800) or 5 mg Mlt on preflight D-1 (1700), flight day D0 (1600), and from D1 to D3 (2300), or placebo (Pbo) at the same times. Nighttime sleep was evaluated by polysomnography and daytime sleepiness from measurements of sleep latencies and continuous wrist actigraphy. Compared with baseline, we found a significant rebound of slow-wave sleep on night 1 (N1) to N2 under Pbo and Mlt and a significant decrease in rapid eye movement sleep on N1 (Pbo) and N1-N3 (Mlt). Sleepiness was objectively increased under Pbo (D1-D6) and Mlt (D1-D3). SRC reduced sleepiness but also tended to affect sleep quality until the last drug day. In conclusion, both drugs have positive effects on some jet lag symptoms after an eastbound flight: SRC on daytime sleepiness, and Mlt on sleep.