Synthesis of Norbornenyl Polymers with Bioactive Oligopeptides by Ring-Opening Metathesis Polymerization

Synthetic norbornenyl polymers with pendent cell adhesive sequences glycine-arginine-glycine-aspartic acid (GRGD) and serine-arginine-asparagine (SRN) were synthesized by ring-opening metathesis polymerization (ROMP) using newly developed ruthenium initiators. Initially, simpler polymers with pendent glycine, alanine, or penta(ethylene glycol) (EO_5) units attached directly or through ethyl and propyl spacers to various norbornenyl backbones were synthesized using Ru CHPh(Cl)_2(PCy_3)_2 (1) as the initiator. The molecular weights, PDI's, polymerization times, yields, and glass transition temperatures were compared for these polymers. As a result of this comparison, poly(5-norbornene-2-carboxyl) was chosen as the backbone for the more complex oligopeptide containing polymers, and norbornene monomers with pendent EO_5 (21), GRGD (24), and SRN (25) units were made. Monomers 21 and 24 were copolymerized to form a poly(norbornene) containing 9.2 mol % GRGD (26a) using 1 as the initiator. However, incorporating larger amounts of GRGD resulted in extremely low yields of polymers that exhibited bimodal molecular weight distributions. Homopolymers and copolymers with larger amounts of GRGD and SRN were synthesized in good yields (32−92%) with monomodal molecular weight distributions using the newly developed, more active, 2,3-dihydroimidazolylidene initiators, Ru CHPh(Cl)_2(PCy_3)(DHIMes) (2) and Ru CH−CH C(CH_3)_2(Cl)_2(PCp_3)(DHIMes) (3). In this way, EO_5 containing copolymers with 49 mol % GRGD (26b), 53 mol % SRN (27b), or 32 mol % GRGD and 21 mol % SRN (28a) were synthesized, as well as copolymer 28b with 53 mol % GRGD and 47 mol % SRN. To alter the presentation of the GRGD, an EO_5 containing copolymer with a propyl spacer between the GRGD and the backbone (30) was also synthesized

Inhibition of Cell Adhesion to Fibronectin by Oligopeptide-Substituted Polynorbornenes

Polynorbornenes substituted with two different peptide sequences from the RGD-containing integrin cell-binding domain of fibronectin are potent inhibitors of human foreskin fibroblast cell adhesion to fibronectin-coated surfaces. Ring-opening metathesis polymerization (ROMP) using Ru═CHPh(Cl)_2(PCy_3)(DHIMes) (1) as an initiator produced polymers substituted with GRGDS and PHSRN peptide sequences. The inhibitory activity was quantified for these polymers and compared to the free peptides and GRGES-containing controls. A homopolymer substituted with GRGDS peptides was significantly more active than the free GRGDS peptide (IC_(50) of 0.18 ± 0.03 and 1.33 ± 0.20 mM respectively), and the copolymer containing both GRGDS and PHSRN is the most potent inhibitor (IC_(50) of 0.04 ± 0.01 mM). These results demonstrate that significant enhancements of observed biological activity can be obtained from polymeric materials containing more than one type of multivalent ligand and that ROMP is a useful method to synthesize such well-defined copolymers

Sex, War, and Disease: The Role of Parasite Infection on Weapon Development and Mating Success in a Horned Beetle (Gnatocerus cornutus)

While parasites and immunity are widely believed to play important roles in the evolution of male ornaments, their potential influence on systems where male weaponry is the object of sexual selection is poorly understood. We experimentally infect larval broad-horned flour beetles with a tapeworm and study the consequent effects on: 1) adult male morphology 2) male-male contests for mating opportunities, and 3) induction of the innate immune system. We find that infection significantly reduces adult male size in ways that are expected to reduce mating opportunities in nature. The sum of our morphological, competition, and immunological data indicate that during a life history stage where no new resources are acquired, males allocate their finite resources in a way that increases future mating potential

A Companion Cell–Dominant and Developmentally Regulated H3K4 Demethylase Controls Flowering Time in Arabidopsis via the Repression of FLC Expression

Flowering time relies on the integration of intrinsic developmental cues and environmental signals. FLC and its downstream target FT are key players in the floral transition in Arabidopsis. Here, we characterized the expression pattern and function of JMJ18, a novel JmjC domain-containing histone H3K4 demethylase gene in Arabidopsis. JMJ18 was dominantly expressed in companion cells; its temporal expression pattern was negatively and positively correlated with that of FLC and FT, respectively, during vegetative development. Mutations in JMJ18 resulted in a weak late-flowering phenotype, while JMJ18 overexpressors exhibited an obvious early-flowering phenotype. JMJ18 displayed demethylase activity toward H3K4me3 and H3K4me2, and bound FLC chromatin directly. The levels of H3K4me3 and H3K4me2 in chromatins of FLC clade genes and the expression of FLC clade genes were reduced, whereas FT expression was induced and the protein expression of FT increased in JMJ18 overexpressor lines. The early-flowering phenotype caused by the overexpression of JMJ18 was mainly dependent on the functional FT. Our findings suggest that the companion cell–dominant and developmentally regulated JMJ18 binds directly to the FLC locus, reducing the level of H3K4 methylation in FLC chromatin and repressing the expression of FLC, thereby promoting the expression of FT in companion cells to stimulate flowering

On the Action of Cyclosporine A, Rapamycin and Tacrolimus on M. avium Including Subspecies paratuberculosis

BACKGROUND: Mycobacterium avium subspecies paratuberculosis (MAP) may be zoonotic. Recently the "immuno-modulators" methotrexate, azathioprine and 6-MP and the "anti-inflammatory" 5-ASA have been shown to inhibit MAP growth in vitro. We concluded that their most plausible mechanism of action is as antiMAP antibiotics. The "immunosuppressants" Cyclosporine A, Rapamycin and Tacrolimus (FK 506) treat a variety of "autoimmune" and "inflammatory" diseases. Rapamycin and Tacrolimus are macrolides. We hypothesized that their mode of action may simply be to inhibit MAP growth. METHODOLOGY: The effect on radiometric MAP (14)CO(2) growth kinetics of Cyclosporine A, Rapamycin and Tacrolimus on MAP cultured from humans (Dominic & UCF 4) or ruminants (ATCC 19698 & 303) and M. avium subspecies avium (ATCC 25291 & 101) are presented as "percent decrease in cumulative GI" (%-DeltacGI.) PRINCIPAL FINDINGS: The positive control clofazimine has 99%-DeltacGI at 0.5 microg/ml (Dominic). Phthalimide, a negative control has no dose dependent inhibition on any strain. Against MAP there is dose dependent inhibition by the immunosuppressants. Cyclosporine has 97%-DeltacGI by 32 microg/ml (Dominic), Rapamycin has 74%-DeltacGI by 64 microg/ml (UCF 4) and Tacrolimus 43%-DeltacGI by 64 microg/ml (UCF 4) CONCLUSIONS: We show heretofore-undescribed inhibition of MAP growth in vitro by "immunosuppressants;" the cyclic undecapeptide Cyclosporine A, and the macrolides Rapamycin and Tacrolimus. These data are compatible with our thesis that, unknowingly, the medical profession has been treating MAP infections since 1942 when 5-ASA and subsequently azathioprine, 6-MP and methotrexate were introduced in the therapy of some "autoimmune" and "inflammatory" diseases

Three Neutrino Δm2\Delta m^2 scales and Singular Seesaw Mechanism

It is shown that the singular seesaw mechanism can simultaneously explain all the existing data supporting nonzero neutrino masses and mixing. The three mass-squared differences that are needed to accommodate the atmospheric neutrino data (through $\nu_\mu - \nu_s$ oscillation), the solar neutrino data via MSW mechanism (through $\nu_e - \nu_\tau$ oscillation), and the positive result of $\nu_\mu - \nu_e$ oscillation from LSND can be generated by this mechanism, whereas the vacuum oscillation solution to the solar neutrino problem is disfavored. We find that the electron and tau neutrino masses are of order $10^{-3}$ eV, and the muon neutrino and a sterile neutrino are almost maximally mixed to give a mass of order 1 eV. Two heavy sterile neutrinos have a mass of order 1 keV which can be obtained by the double seesaw mechanism with an intermediate mass scale $\sim 10^5$ GeV. A possible origin of such a scale is discussed.Comment: Revtex, 7 pages with 1 epsfig (uuencoded

Impact of primary kidney disease on the effects of empagliflozin in patients with chronic kidney disease: secondary analyses of the EMPA-KIDNEY trial

Background: The EMPA KIDNEY trial showed that empagliflozin reduced the risk of the primary composite outcome of kidney disease progression or cardiovascular death in patients with chronic kidney disease mainly through slowing progression. We aimed to assess how effects of empagliflozin might differ by primary kidney disease across its broad population. Methods: EMPA-KIDNEY, a randomised, controlled, phase 3 trial, was conducted at 241 centres in eight countries (Canada, China, Germany, Italy, Japan, Malaysia, the UK, and the USA). Patients were eligible if their estimated glomerular filtration rate (eGFR) was 20 to less than 45 mL/min per 1·73 m2, or 45 to less than 90 mL/min per 1·73 m2 with a urinary albumin-to-creatinine ratio (uACR) of 200 mg/g or higher at screening. They were randomly assigned (1:1) to 10 mg oral empagliflozin once daily or matching placebo. Effects on kidney disease progression (defined as a sustained ≥40% eGFR decline from randomisation, end-stage kidney disease, a sustained eGFR below 10 mL/min per 1·73 m2, or death from kidney failure) were assessed using prespecified Cox models, and eGFR slope analyses used shared parameter models. Subgroup comparisons were performed by including relevant interaction terms in models. EMPA-KIDNEY is registered with ClinicalTrials.gov, NCT03594110. Findings: Between May 15, 2019, and April 16, 2021, 6609 participants were randomly assigned and followed up for a median of 2·0 years (IQR 1·5–2·4). Prespecified subgroupings by primary kidney disease included 2057 (31·1%) participants with diabetic kidney disease, 1669 (25·3%) with glomerular disease, 1445 (21·9%) with hypertensive or renovascular disease, and 1438 (21·8%) with other or unknown causes. Kidney disease progression occurred in 384 (11·6%) of 3304 patients in the empagliflozin group and 504 (15·2%) of 3305 patients in the placebo group (hazard ratio 0·71 [95% CI 0·62–0·81]), with no evidence that the relative effect size varied significantly by primary kidney disease (pheterogeneity=0·62). The between-group difference in chronic eGFR slopes (ie, from 2 months to final follow-up) was 1·37 mL/min per 1·73 m2 per year (95% CI 1·16–1·59), representing a 50% (42–58) reduction in the rate of chronic eGFR decline. This relative effect of empagliflozin on chronic eGFR slope was similar in analyses by different primary kidney diseases, including in explorations by type of glomerular disease and diabetes (p values for heterogeneity all >0·1). Interpretation: In a broad range of patients with chronic kidney disease at risk of progression, including a wide range of non-diabetic causes of chronic kidney disease, empagliflozin reduced risk of kidney disease progression. Relative effect sizes were broadly similar irrespective of the cause of primary kidney disease, suggesting that SGLT2 inhibitors should be part of a standard of care to minimise risk of kidney failure in chronic kidney disease. Funding: Boehringer Ingelheim, Eli Lilly, and UK Medical Research Council

Multi-messenger observations of a binary neutron star merger

On 2017 August 17 a binary neutron star coalescence candidate (later designated GW170817) with merger time 12:41:04 UTC was observed through gravitational waves by the Advanced LIGO and Advanced Virgo detectors. The Fermi Gamma-ray Burst Monitor independently detected a gamma-ray burst (GRB 170817A) with a time delay of ~1.7 s with respect to the merger time. From the gravitational-wave signal, the source was initially localized to a sky region of 31 deg2 at a luminosity distance of 40+8-8 Mpc and with component masses consistent with neutron stars. The component masses were later measured to be in the range 0.86 to 2.26 Mo. An extensive observing campaign was launched across the electromagnetic spectrum leading to the discovery of a bright optical transient (SSS17a, now with the IAU identification of AT 2017gfo) in NGC 4993 (at ~40 Mpc) less than 11 hours after the merger by the One- Meter, Two Hemisphere (1M2H) team using the 1 m Swope Telescope. The optical transient was independently detected by multiple teams within an hour. Subsequent observations targeted the object and its environment. Early ultraviolet observations revealed a blue transient that faded within 48 hours. Optical and infrared observations showed a redward evolution over ~10 days. Following early non-detections, X-ray and radio emission were discovered at the transient’s position ~9 and ~16 days, respectively, after the merger. Both the X-ray and radio emission likely arise from a physical process that is distinct from the one that generates the UV/optical/near-infrared emission. No ultra-high-energy gamma-rays and no neutrino candidates consistent with the source were found in follow-up searches. These observations support the hypothesis that GW170817 was produced by the merger of two neutron stars in NGC4993 followed by a short gamma-ray burst (GRB 170817A) and a kilonova/macronova powered by the radioactive decay of r-process nuclei synthesized in the ejecta