Beyond reduction of atherosclerosis: PON2 provides apoptosis resistance and stabilizes tumor cells

Major contributors to atherosclerosis are oxidative damage and endoplasmic reticulum (ER) stress-induced apoptosis; both of which can be diminished by the anti-oxidative protein paraoxonase-2 (PON2). ER stress is also relevant to cancer and associated with anti-cancer treatment resistance. Hence, we addressed, for the first time, whether PON2 contributes to tumorigenesis and apoptotic escape. Intriguingly, we found that several human tumors upregulated PON2 and such overexpression provided resistance to different chemotherapeutics (imatinib, doxorubicine, staurosporine, or actinomycin) in cell culture models. This was reversed after PON2 knock-down. Remarkably, just deficiency of PON2 caused apoptosis of selective tumor cells per se, demonstrating a previously unanticipated oncogenic function. We found a dual mechanistic role. During ER stress, high PON2 levels lowered redox-triggered induction of pro-apoptotic CHOP particularly via the JNK pathway, which prevented mitochondrial cell death signaling. Apart from CHOP, PON2 also diminished intrinsic apoptosis as it prevented mitochondrial superoxide formation, cardiolipin peroxidation, cytochrome c release, and caspase activation. Ligand-stimulated apoptosis by TRAIL or TNFα remained unchanged. Finally, PON2 knock-down caused vast reactive oxygen species formation and stimulated JNK-triggered CHOP expression, but inhibition of JNK signaling did not prevent cell death, demonstrating the pleiotropic, dominating anti-oxidative effect of PON2. Therefore, targeting redox balance is powerful to induce selective tumor cell death and proposes PON2 as new putative anti-tumor candidate

Femtosecond x-ray diffraction from an aerosolized beam of protein nanocrystals

We demonstrate near-atomic-resolution Bragg diffraction from aerosolized single granulovirus crystals using an x-ray free-electron laser. The form of the aerosol injector is nearly identical to conventional liquid-microjet nozzles, but the x-ray-scattering background is reduced by several orders of magnitude by the use of helium carrier gas rather than liquid. This approach provides a route to study the weak diffuse or lattice-transform signal arising from small crystals. The high speed of the particles is particularly well suited to upcoming MHz-repetition-rate x-ray free-electron lasers

Mapping the Complete Reaction Path of a Complex Photochemical Reaction

We probe the dynamics of dissociating CS2 molecules across the entire reaction pathway upon excitation. Photoelectron spectroscopy measurements using laboratory-generated femtosecond extreme ultraviolet pulses monitor the competing dissociation, internal conversion, and intersystem crossing dynamics. Dissociation occurs either in the initially excited singlet manifold or, via intersystem crossing, in the triplet manifold. Both product channels are monitored and show that despite being more rapid, the singlet dissociation is the minor product and that triplet state products dominate the final yield. We explain this by consideration of accurate potential energy curves for both the singlet and triplet states. We propose that rapid internal conversion stabilises the singlet population dynamically, allowing for singlet-triplet relaxation via intersystem crossing and efficient formation of spin-forbidden dissociation products on longer timescales. The study demonstrates the importance of measuring the full reaction pathway for defining accurate reaction mechanisms

Paraoxonase 2 protein is spatially expressed in the human placenta and selectively reduced in labour

Humans parturition involves interaction of hormonal, neurological, mechanical stretch and inflammatory pathways and the placenta plays a crucial role. The paraoxonases (PONs 1–3) protect against oxidative damage and lipid peroxidation, modulation of endoplasmic reticulum stress and regulation of apoptosis. Nothing is known about the role of PON2 in the placenta and labour. Since PON2 plays a role in oxidative stress and inflammation, both features of labour, we hypothesised that placental PON2 expression would alter during labour. PON2 was examined in placentas obtained from women who delivered by cesarean section and were not in labour and compared to the equivalent zone of placentas obtained from women who delivered vaginally following an uncomplicated labour. Samples were obtained from 12 sites within each placenta: 4 equally spaced apart pieces were sampled from the inner, middle and outer placental regions. PON2 expression was investigated by Western blotting and real time PCR. Two PON2 forms, one at 62 kDa and one at 43 kDa were found in all samples. No difference in protein expression of either isoform was found between the three sites in either the labour or non-labour group. At the middle site there was a highly significant decrease in PON2 expression in the labour group when compared to the non-labour group for both the 62 kDa form (p = 0.02) and the 43 kDa form (p = 0.006). No spatial differences were found within placentas at the mRNA level in either labour or non-labour. There was, paradoxically, an increase in PON2 mRNA in the labour group at the middle site only. This is the first report to describe changes in PON2 in the placenta in labour. The physiological and pathological significance of these remains to be elucidated but since PON2 is anti-inflammatory further studies are warranted to understand its role

Deficiency of Antioxidative Paraoxonase 2 (Pon2) Leads to Increased Number of Phenotypic LT-HSCs and Disturbed Erythropoiesis

Background. Long-term hematopoietic stem cells (LT-HSCs) reside in bone marrow niches with tightly controlled reactive oxygen species (ROS) levels. ROS increase results into LT-HSC differentiation and stem cell exhaustion. Paraoxonase 2 (PON2) has been shown to be important for ROS control. Objectives. We investigate the effects of inactivation of the PON2 gene on hematopoietic cell differentiation and activity. Methods and Results. In young mice with inactivated Pon2 gene (Pon2-/-, -/- BM outcompeted WT BM at early time points. ROS levels were significantly increased in Pon2-/- whole BM, but not in Pon2-/- LT-HSCs. In more differentiated stages of hematopoiesis, Pon2 deficiency led to a misbalanced erythropoiesis both in physiologic and stress conditions. In older mice (>9 months), Pon2 depletion caused an increase in LT-HSCs as well as increased levels of granulocyte/macrophage progenitors (GMPs) and myeloid skewing, indicating a premature aging phenotype. No significant changes in ROS levels in old Pon2-/- LT- and short-term (ST-) HSCs were observed, but a significant reduction of spontaneous apoptotic cell death was measured. RNA-seq analysis in Pon2-/- LT-HSCs identified overrepresentation of genes involved in the C-X-C chemokine receptor type 4 (Cxcr4) signaling, suggesting compensatory mechanisms to overcome ROS-mediated accelerated aging in hematopoietic progenitor cells. Conclusions. In summary, our current data indicate that PON2 is involved in the regulation of HSC functions

A European study on decellularized homografts for pulmonary valve replacement: initial results from the prospective ESPOIR Trial and ESPOIR Registry data\u2020

OBJECTIVES: Decellularized pulmonary homografts (DPH) have shown excellent results for pulmonary valve replacement. However, controlled multicentre studies are lacking to date.METHODS: Prospective European multicentre trial evaluating DPH for pulmonary valve replacement. Matched comparison of DPH to bovine jugular vein (BJV) conduits and cryopreserved homografts (CH) considering patient age, type of heart defect and previous procedures.RESULTS: In total, 121 patients (59 female) were prospectively enrolled (August 2014-December 2016), age 21.3 +/- 14.4 years, DPH diameter 24.4 +/- 2.8 mm. No adverse events occurred with respect to surgical handling; there were 2 early deaths (30 + 59 years) due to myocardial failure after multi-valve procedures and no late mortality (1.7% mortality). After a mean follow-up of 2.2 +/- 0.6 years, the primary efficacy end points mean peak gradient (16.1 +/- 12.1 mmHg) and regurgitation (mean 0.25 +/- 0.48, grade 0-3) were excellent. One reoperation was required for recurrent subvalvular stenosis caused by a pericardial patch and 1 balloon dilatation was performed on a previously stented LPA. 100% follow-up for DPH patients operated before or outside the trial (n = 114) included in the ESPOIR Registry, age 16.6 +/- 10.4 years, diameter 24.1 +/- 4.2 mm, follow-up 5.1 +/- 3.0 years. The combined DPH cohort, n = 235, comprising both Trial and Registry data showed significantly better freedom from explantation (DPH 96.7 +/- 2.1%, CH 84.4 +/- 3.2%, P = 0.029 and BJV 82.7 +/- 3.2%, P = 0.012) and less structural valve degeneration at 10 years when matched to CH, n = 235 and BJV, n = 235 (DPH 61.4 +/- 6.6%, CH 39.9 +/- 4.4%, n.s., BJV 47.5 +/- 4.5%, P = 0.029).CONCLUSIONS: Initial results of the prospective multicentre ESPOIR Trial showed DPH to be safe and efficient. Current DPH results including Registry data were superior to BJV and CH.Thoracic Surger

New aerodynamic lens injector for single particle diffractive imaging

An aerodynamic lens injector was developed specifically for the needs of single-particle diffractive imaging experiments at free-electron lasers. Its design allows for quick changes of injector geometries and focusing properties in order to optimize injection for specific individual samples. Here, we present results of its first use at the FLASH free-electron-laser facility. Recorded diffraction patterns of polystyrene spheres are modeled using Mie scattering, which allowed for the characterization of the particle beam under diffractive-imaging conditions and yield good agreement with particle-trajectory simulations

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Thoracic Surger

A Novel Inhibitor of Human La Protein with Anti-HBV Activity Discovered by Structure-Based Virtual Screening and In Vitro Evaluation

Background: Over 350 million people worldwide are infected with hepatitis B virus (HBV), a major cause of liver failure and hepatocellular carcinoma. Current therapeutic agents are highly effective, but are also associated with development of viral resistance. Therefore, strategies for identifying other anti-HBV agents with specific, but distinctive mechanisms of action are needed. The human La (hLa) protein, which forms a stabilizing complex with HBV RNA ribonucleoprotein to promote HBV replication, is a promising target of molecular therapy. Aims: This study aimed to discover novel inhibitors of hLa that could inhibit HBV replication and expression. Methods: A multistage molecular docking approach was used to screen a Specs database and an in-house library against hLa binding sites. Sequential in vitro evaluations were performed to detect potential compounds with high scores in HepG2.2.15 cells. Results: Of the 26 potential compounds with high scores chosen for experimental verification, 12 had HBV DNA inhibition ratios of less than 50 % with P,0.05. Six had significant inhibition of HBV e antigen (HBeAg) levels, and 13 had significant inhibition of HBV surface antigen (HBsAg) levels by in vitro assays. Compounds HBSC-11, HBSC-15 and HBSC-34 (HBSC is system prefix for active compounds screened by the library) were selected for evaluation. HBSC-11 was found to have an obvious inhibitory effect on hLa transcription and expression