Late onset cardiovascular dysfunction in adult mice resulting from galactic cosmic ray exposure.

The complex and inaccessible space radiation environment poses an unresolved risk to astronaut cardiovascular health during long-term space exploration missions. To model this risk, healthy male c57BL/6 mice aged six months (corresponding to an astronaut of 34 years) were exposed to simplified galactic cosmic ray (GCR5-ion; 5-ion sim) irradiation at the NASA Space Radiation Laboratory (NSRL) at Brookhaven National Laboratories (BNL). Multi-modal cardiovascular functional assessments performed longitudinally and terminally revealed significant impairment in cardiac function in mice exposed to GCR5-ion compared to unirradiated controls, gamma irradiation, or single mono-energetic ions (56Fe or 16O). GCR5-ion-treated mice exhibited increased arterial elastance likely mediated by disruption of elastin fibers. This study suggests that a single exposure to GCR5-ion is associated with deterioration in cardiac structure and function that becomes apparent long after exposure, likely associated with increased morbidity and mortality. These findings represent important health considerations when preparing for successful space exploration.

The Relationship Between Atrial Fibrillation, Mitral Regurgitation, and Heart Failure Subtype: The ARIC Study.

BACKGROUND: Atrial fibrillation (AF) and mitral regurgitation (MR) are closely interrelated in the setting of heart failure (HF). Here we investigate the prevalence and prognostic significance of AF in patients with acute decompensated HF (ADHF) stratified by MR severity. METHODS AND RESULTS: The Atherosclerosis Risk in Communities Study investigated ADHF hospitalizations in residents greater than or equal to 55 years of age in 4 US communities. ADHF cases were stratified by MR severity (none/mild or moderate/severe) and HF subtype (HF with reduced [HFrEF] or preserved [HFpEF] ejection fraction). The odds of AF in patients with increasing MR severity was estimated using multivariable logistic regression, adjusting for age, race, sex, diabetes, hypertension, coronary artery disease, hemodialysis, stroke, and anemia. Cox regression models were used to assess the association of AF with 1-year mortality in patients with HFpEF and HFrEF, stratified by MR severity and adjusted as described, also adjusting for the year of hospitalization. From 2005 to 2014, there were 3,878 ADHF hospitalizations (17,931 weighted). AF was more likely in those with higher MR severity regardless of HF subtype; more so in HFpEF (odds ratio [OR] 1.38, 95% confidence interval [CI], 1.31-1.45) than in HFrEF (OR, 1.19, 95% CI, 1.13-1.25) (interaction P [by HF subtype] < .01). When stratified by HF type, association between AF and 1-year mortality was noted in patients with HFpEF (OR, 1.28, 95% CI 1.04-1.56) but not HFrEF (OR 0.96, 95% CI 0.79-1.16) (interaction by EF subtype, P = .02). CONCLUSIONS: In patients with ADHF, AF prevalence increased with MR severity and this effect was more pronounced in HFpEF compared with HFrEF. AF was associated with an increased 1-year mortality only in patients with HFpEF and concomitant moderate/severe MR. REGISTRATION: NCT00005131, https://clinicaltrials.gov/ct2/show/NCT00005131.

The use of an inexpensive processing aid device (the Mouse PAD) to facilitate rodent tissue banking.

The generation of an experimental animal model often requires considerable investment of both time and money. Typically, investigators are interested in specific organs and when experimental animals are euthanized, tissues that are not the focus of the research project are discarded. However, the remaining tissues from these animals could provide valuable scientific information if efficient, error-proof and economical approaches to collect and biobank them were available. We have developed a device that, when incorporated into our tissue processing workflow, allows for high-throughput collection and processing of multiple rodent organ systems. This device, the mouse Processing Aid Device, or mouse PAD, helps to standardize organ collection and increase its efficiency.

A normothermic ex vivo organ perfusion delivery method for cardiac transplantation gene therapy.

Clinically, both percutaneous and surgical approaches to deliver viral vectors to the heart either have resulted in therapeutically inadequate levels of transgene expression or have raised safety concerns associated with extra-cardiac delivery. Recent developments in the field of normothermic ex vivo cardiac perfusion storage have now created opportunities to overcome these limitations and safety concerns of cardiac gene therapy. This study examined the feasibility of ex vivo perfusion as an approach to deliver a viral vector to a donor heart during storage and the resulting bio distribution and expression levels of the transgene in the recipient post-transplant. The influence of components (proprietary solution, donor blood, and ex vivo circuitry tubing and oxygenators) of the Organ Care System (OC) (TransMedics, Inc., Andover MA) on viral vector transduction was examined using a cell-based luciferase assay. Our ex vivo perfusion strategy, optimized for efficient Adenoviral vector transduction, was utilized to deliver 5 × 1013 total viral particles of an Adenoviral firefly luciferase vector with a cytomegalovirus (CMV) promotor to porcine donor hearts prior to heterotopic implantation. We have evaluated the overall levels of expression, protein activity, as well as the bio distribution of the firefly luciferase protein in a series of three heart transplants at a five-day post-transplant endpoint. The perfusion solution and the ex vivo circuitry did not influence viral vector transduction, but the serum or plasma fractions of the donor blood significantly inhibited viral vector transduction. Thus, subsequent gene delivery experiments to the explanted porcine heart utilized an autologous blood recovery approach to remove undesired plasma or serum components of the donor blood prior to its placement into the circuit. Enzymatic assessment of luciferase activity in tissues (native heart, allograft, liver etc.) obtained post-transplant day five revealed wide-spread and robust luciferase activity in all regions of the allograft (right and left atria, right and left ventricles, coronary arteries) compared to the native recipient heart. Importantly, luciferase activity in recipient heart, liver, lung, spleen, or psoas muscle was within background levels. Similar to luciferase activity, the luciferase protein expression in the allograft appeared uniform and robust across all areas of the myocardium as well as in the coronary arteries. Importantly, despite high copy number of vector genomic DNA in transplanted heart tissue, there was no evidence of vector DNA in either the recipient's native heart or liver. Overall we demonstrate a simple protocol to achieve substantial, global gene delivery and expression isolated to the cardiac allograft. This introduces a novel method of viral vector delivery that opens the opportunity for biological modification of the allograft prior to implantation that may improve post-transplant outcomes.

MRGPR-mediated activation of local mast cells clears cutaneous bacterial infection and protects against reinfection.

Mast cells (MCs) are strategically distributed at barrier sites and prestore various immunocyte-recruiting cytokines, making them ideal targets for selective activation to treat peripheral infections. Here, we report that topical treatment with mastoparan, a peptide MC activator (MCA), enhances clearance of Staphylococcus aureus from infected mouse skins and accelerates healing of dermonecrotic lesions. Mastoparan functions by activating connective tissue MCs (CTMCs) via the MRGPRX2 (Mas-related G protein-coupled receptor member X2) receptor. Peripheral CTMC activation, in turn, enhances recruitment of bacteria-clearing neutrophils and wound-healing CD301b+ dendritic cells. Consistent with MCs playing a master coordinating role, MC activation also augmented migration of various antigen-presenting dendritic cells to draining lymph nodes, leading to stronger protection against a second infection challenge. MCAs therefore orchestrate both the innate and adaptive immune arms, which could potentially be applied to combat peripheral infections by a broad range of pathogens.

Transition Metal Functionalization of P4 Using a Diarylgermylene Anchor.

The manipulation of white phosphorus (P4) has been a long-standing challenge for chemists. While the holy grail remains at finding a method to catalytically activate and functionalize P4 to yield new organophosphorus compounds, fundamental research lies in developing procedures to control the reactivity of elemental phosphorus. In this work, Lewis acidic transition metal moieties M(CO)5 (M = Cr, Mo, W) and AuCl react with P4 derivatized with a low valent germanium compound. For both M(CO)5 and AuCl, bis-functionalized products can be formed; however the monosubstituted derivatives are found to be more stable, and the decomposition can be monitored by 31P{1H} NMR spectroscopy. The selective reactivity of white phosphorus, once a P-P bond has been activated, is a key step in yielding new organophosphorus compounds.

Reversible, photoinduced activation of P4 by low-coordinate main group compounds.

Two unique systems based on low-coordinate main group elements that activate P4 are shown to quantitatively release the phosphorus cage upon short exposure to UV light. This reactivity marks the first reversible reactivity of P4, and the germanium system can be cycled 5 times without appreciable loss in activity. Theoretical calculations reveal that the LUMO is antibonding with respect to the main group element-phosphorus bonds and bonding with respect to reforming the P4 tetrahedron, providing a rationale for this unprecedented activity, and suggesting that the process is tunable based on the substituents.

Activation of gaseous PH3 with low coordinate diaryltetrylene compounds.

The reaction of phosphine gas with a low coordinate diaryl germylene or diarylstannylene results in both oxidative addition and arene elimination at the group 14 atom. The products were characterised by (31)P NMR spectroscopy and X-ray crystallography, and represent the first P-H bond activation by a heavy group 14 element compound.

Mechanisms of reactions of open-shell, heavier group 14 derivatives with small molecules: n-π* back-bonding in isocyanide complexes, C-H activation under ambient conditions, CO coupling, and ancillary molecular interactions.

The main themes of this review are the mechanisms of the reactions of germanium and tin analogues of carbenes with isocyanides, CO, ammonia, and related molecules. The treatment of Ge(Ar(Me6))2 (Ar(Me6) = C6H3-2,6(C6H2-2,4,6-Me3)2) with MeNC or Bu(t)NC afforded 1:1 complexes, but the increase in the electron density at germanium leads to C-H activation at the isocyanide methyl or tert-butyl substituents. For MeNC, the initial adduct formation is followed by a migratory insertion of the MeNC carbon into a Ge-C(ipso) bond of an aryl substituent. The addition of excess MeNC led to sequential insertions of two further MeNC molecules. The third insertion led to methylisocyanide methyl group C-H activation, to afford an azagermacyclopentadienyl species. The Bu(t)NC complex (Ar(Me6))2GeCNBu(t) spontanously transforms into (Ar(Me6))2Ge(H)CN and isobutene with C-H activation of the Bu(t) substituent. The germylene Ge(Ar(Me6))(Ar(Pr(i)4)) [Ar(Pr(i)4) = C6H3-2,6(C6H3-2,6-Pr(i)2)2] reacted with CO to afford α-germyloxyketones. The initial step is the formation of a 1:1 complex, followed by migratory insertion into the Ge-C bond of the Ar(Pr(i)4) ligand to give Ar(Me6)GeC(O)Ar(Pr(i)4). Insertion of a second CO gave Ar(Me6)GeC(O)C(O)Ar(Pr(i)4), which rearranges to afford α-germyloxyketone. No reaction was observed for Sn(Ar(Me6))2 with RNC (R = Me, Bu(t)) or CO. Spectroscopic (IR) results and density functional theory (DFT) calculations showed that the reactivity can be rationalized on the basis of Ge-C (isocyanide or CO) Ge(n) → π* (ligand) back-bonding. The reaction of Ge(Ar(Me6))2 and Sn(Ar(Me6))2 with ammonia or hydrazines initially gave 1:1 adducts. However, DFT calculations show that there are ancillary N-H---N interactions with a second ammonia or hydrazine, which stabilizes the transition state to form germanium(IV) hydride (amido or hydrazido) products. For tin, arene elimination is favored by a buildup of electron density at the tin, as well as the greater polarity of the Sn-C(ipso) bond. Germanium(IV) products were observed upon reaction of Ge(Ar(Me6))2 with acids, whereas reactions of Sn(Ar(Me6))2 with acids did not give tin(II) products. In contrast to reactions with NH3, there is no buildup of negative charge at tin upon protonation, and its subsequent reaction with conjugate bases readily affords the tin(IV) products.

Mechanistic study of stepwise methylisocyanide coupling and C-H activation mediated by a low-valent main group molecule.

An experimental and DFT investigation of the mechanism of the coupling of methylisocyanide and C-H activation mediated by the germylene (germanediyl) Ge(Ar(Me6))2 (Ar(Me6) = C6H3-2,6(C6H2-2,4,6-Me3)2) showed that it proceeded by initial MeNC adduct formation followed by an isomerization involving the migratory insertion of the MeNC carbon into the Ge-C ligand bond. Addition of excess MeNC led to sequential insertions of two further MeNC molecules into the Ge-C bond. The insertion of the third MeNC leads to methylisocyanide methyl group C-H activation to afford an azagermacyclopentadienyl species. The X-ray crystal structures of the 1:1 (Ar(Me6))2GeCNMe adduct, the first and final insertion products (Ar(Me6))GeC(NMe)Ar(Me6) and (Ar(Me6))GeC(NHMe)C(NMe)C(Ar(Me6))NMe were obtained. The DFT calculations on the reaction pathways represent the first detailed mechanistic study of isocyanide oligomerization by a p-block element species.

A germanium isocyanide complex featuring (n → π*) back-bonding and its conversion to a hydride/cyanide product via C-H bond activation under mild conditions.

Reaction of the diarylgermylene Ge(Ar(Me(6)))(2) [Ar(Me(6)) = C(6)H(3)-2,6-(C(6)H(2)-2,4,6-(CH(3))(3))(2)] with tert-butyl isocyanide gave the Lewis adduct species (Ar(Me(6)))(2)GeCNBu(t), in which the isocyanide ligand displays a decreased C-N stretching frequency consistent with an n → π* back-bonding interaction. Density functional theory confirmed that the HOMO is a Ge-C bonding combination between the lone pair of electrons on the germanium atom and the C-N π* orbital of the isocyanide ligand. The complex undergoes facile C-H bond activation to produce a new diarylgermanium hydride/cyanide species and isobutene via heterolytic cleavage of the N-Bu(t) bond.

Activation of olefins with low-valent gallium compounds under ambient conditions.

The reactions of Ar'GaGaAr' (Ar' = C(6)H(3)-2,6-(C(6)H(3)-2,6-(i)Pr(2))(2)) with alkenes revealed the addition of two olefins per Ar'GaGaAr' under ambient conditions for ethylene, propene, 1-hexene and styrene but no reactions with more hindered or cyclic olefins.

MOZ and MOZ-CBP cooperate with NF-kappaB to activate transcription from NF-kappaB-dependent promoters.

OBJECTIVE: Monocytic zinc finger (MOZ) maintains hematopoietic stem cells and, upon fusion to the coactivator CREB-binding protein (CBP), induces acute myeloid leukemia (AML). Leukemic stem cells in AML often exhibit excessive signal-dependent activity of the transcription factor nuclear factor (NF)-kappaB. Because aberrant interaction between NF-kappaB and coactivators represents an alternative mechanism for enhancing NF-kappaB activity, we evaluated whether MOZ and MOZ-CBP cooperate with NF-kappaB to activate transcription from NF-kappaB-dependent promoters. METHODS: The ability of MOZ, MOZ mutants, and MOZ-CBP to enhance expression of NF-kappaB-dependent promoters was tested in reporter studies. The interaction between MOZ and NF-kappaB was evaluated by both coimmunoprecipitation and glutathione S-transferase pulldown assays. RESULTS: MOZ activates transcription from the NF-kappaB-dependent interleukin-8 promoter; interestingly, this effect is markedly enhanced by CBP. Although MOZ has less potent transcriptional activity than MOZ-CBP, both proteins cooperate with steroid receptor coactivator-1 to activate transcription. MOZ also induces multiple NF-kappaB-dependent viral promoters. Importantly, MOZ associates in a protein complex with the p65 subunit of NF-kappaB and interacts directly with p65 in vitro. Transcriptional activity of MOZ requires its C-terminal domain, which is absent from MOZ-CBP, indicating that the transcriptional activity of MOZ-CBP derives from its CBP sequence. CONCLUSIONS: MOZ interacts with the p65 subunit of NF-kappaB and enhances expression of NF-kappaB-dependent promoters. The more potent transcriptional activity of MOZ-CBP derives from its CBP sequence. Thus, interaction between NF-kappaB and MOZ-CBP may play an important role in the pathogenesis of certain acute myeloid leukemias.