Collagen membrane functionalized with magnesium oxide via room-temperature atomic layer deposition promotes osteopromotive and antimicrobial properties.

Artificial bone grafting materials such as collagen are gaining interest due to the ease of production and implantation. However, collagen must be supplemented with additional coating materials for improved osteointegration. Here, we report room-temperature atomic layer deposition (ALD) of MgO, a novel method to coat collagen membranes with MgO. Characterization techniques such as X-ray photoelectron spectroscopy, Raman spectroscopy, and electron beam dispersion mapping confirm the chemical nature of the film. Scanning electron and atomic force microscopies show the surface topography and morphology of the collagen fibers were not altered during the ALD of MgO. Slow release of magnesium ions promotes bone growth, and we show the deposited MgO film leaches trace amounts of Mg when incubated in phosphate-buffered saline at 37 °C. The coated collagen membrane had a superhydrophilic surface immediately after the deposition of MgO. The film was not toxic to human cells and demonstrated antibacterial properties against bacterial biofilms. Furthermore, in vivo studies performed on calvaria rats showed MgO-coated membranes (200 and 500 ALD) elicit a higher inflammatory response, leading to an increase in angiogenesis and a greater bone formation, mainly for Col-MgO500, compared to uncoated collagen. Based on the characterization of the MgO film and in vitro and in vivo data, the MgO-coated collagen membranes are excellent candidates for guided bone regeneration.

Identification of 5-(3-(methylsulfonyl)phenyl)-3-(4-(methylsulfonyl)phenyl)-3H-imidazo[4,5-b]pyridine as novel orally bioavailable and metabolically stable antimalarial compound for further exploration.

Malaria continues to be a significant public health problem threatened by the emergence and spread of resistance to artemisinin-based combination therapies and marked half a million deaths in 2016. A new imidazopyridine chemotype has been envisaged through scaffold-hopping approach combined with docking studies for putative-binding interactions with Plasmodium falciparum phosphatidylinositol-4-kinase (PfPI4K) target. The docking results steered to the synthesis of compound 1 [5-(3-(methylsulfonyl)phenyl)-3-(4-(methylsulfonyl)phenyl)-3H-imidazo[4,5-b]pyridine] followed by the in vitro screening for antiplasmodial activity and ADME-PK studies. Combined with potent antimalarial activity of compound 1 (Pf3D7 IC50  = 29 nM) with meager in vitro intrinsic clearance, moderate plasma-protein binding, and acceptable permeability, compound 1 displayed sustained exposure and high oral bioavailability in mice and can thus have the potential as next generation PI4K inhibitor for in vivo studies.

Sustainable Intensification of a Rice-Maize System through Conservation Agriculture to Enhance System Productivity in Southern India.

Integrated management of rice-maize systems is an emerging challenge in southern India due to improper rice residues and tillage management in maize crops. Conservation agriculture (CA) practices such as a reduced tillage and maintaining stubble mulch may hold the potential to increase yields, reduce crop establishment costs and increase farm incomes. A five-year trial was performed to study the effect of different CA and establishment methods in rice on system productivity, profitability, and soil carbon status in a rice-maize system. In the rainy season, the trial consisted of two main treatments: (i) normal manual transplanting and (ii) direct-wet seeding, and three sub-main treatments at different sowing dates with fifteen day intervals. In addition, in the winter season, two tillage treatments (conventional and minimum tillage) were imposed over the rainy season treatments. Both rice and maize were grown under irrigated conditions. The results showed that sowing times at 15 day intervals did not impact the yield significantly. Transplanted rice obtained a significantly higher grain yield during the first four years, but in the last year, the yield was similar in both of the establishment methods. In the winter season, conventional tilled maize recorded a higher cob yield than under the minimum tilled treatment, except for the last year, where both the tillage treatment effects were the same. System productivity of CA-based minimum tilled rice-maize was inferior during the first three years but was superior to the conventionally tilled method in the fourth and fifth year. Pooled analysis revealed that the conventionally tilled rice-maize system resulted in a similar system productivity as that of the CA during the study period. The cost-benefit analysis revealed that transplanted rice and conventionally tilled maize fetched higher net returns of INR 111,074 and INR 101,658/ha, respectively, over the direct-wet seeded rice and CA. In addition, the 15 July rice sown followed by the maize system led to an increase in irrigation water productivity by 15.7%, and the total water (irrigation + rainfall) productivity by 27.1% in the maize crop compared with the 30 July sown system. The CA-based rice-maize system resulted in a significantly higher very labile (0.194%) and labile (0.196%) carbon concentration at a 0-5 cm depth of soil compared to those under the conventional system. Thus, CA can be recommended for southern India and similar agro-ecological tropic and sub-tropic conditions. This system can be followed with appropriate location-specific modification in South-Asian countries, where crop yields and soil health are declining as a result of continuous cereal-cereal crop rotation.

Antinociceptive properties of shikonin: in vitro and in vivo studies.

Shikonin possess a diverse spectrum of pharmacological properties in multiple therapeutic areas. However, the nociceptive effect of shikonin is not largely known. To investigate the antinociceptive potential of shikonin, panel of GPCRs, ion channels, and enzymes involved in pain pathogenesis were studied. To evaluate the translation of shikonin efficacy in vivo, it was tested in 3 established rat pain models. Our study reveals that shikonin has significant inhibitory effect on pan sodium channel/N1E115 and NaV1.7 channel with half maximal inhibitory concentration (IC50) value of 7.6 µmol/L and 6.4 µmol/L, respectively, in a cell-based assay. Shikonin exerted significant dose dependent antinociceptive activity at doses of 0.08%, 0.05%, and 0.02% w/v in pinch pain model. In mechanical hyperalgesia model, dose of 10 and 3 mg/kg (intraperitoneal) produced dose-dependent analgesia and showed 67% and 35% reversal of hyperalgesia respectively at 0.5 h. Following oral administration, it showed 39% reversal at 30 mg/kg dose. When tested in first phase of formalin induced pain, shikonin at 10 mg/kg dose inhibited paw flinching by ∼71%. In all studied preclinical models, analgesic effect was similar or better than standard analgesic drugs. The present study unveils the mechanistic role of shikonin on pain modulation, predominantly via sodium channel modulation, suggesting that shikonin could be developed as a potential pain blocker.

EN3427: a novel cationic aminoindane with long-acting local anesthetic properties.

BACKGROUND: Currently approved local anesthetic drugs provide relatively brief local anesthesia that is appropriate and even desirable in some settings, but an extended duration of action beyond their capabilities would be a distinct benefit in other clinical situations. We implemented a drug discovery program that sought to identify novel local anesthetic molecules that specifically demonstrated a long-acting, preferential action on nociceptor sensory afferents that expressed transient receptor potential (TRP) channels. The hypothesis we tested was whether relatively membrane-impermeant local anesthetic molecules could confer long-lasting anesthesia if neuronal access was facilitated by TRP channel activation. The current work describes in vivo studies on a lead molecule that emerged from the discovery program, EN3427, in several rodent pain models. METHODS: Studies were performed on male Sprague-Dawley rats using 2 models of acute mechanical paw-pinch-evoked and pinprick-evoked nociceptive pain. Behavioral responses to noxious stimuli were assessed at baseline, that is, before any pharmacologic intervention, and at various timepoints after a single perisciatic or subcutaneous administration of either EN3427 alone or in combination with lidocaine. Paw withdrawal thresholds or cutaneous trunci reflexes were quantified, and pre-post drug values were compared statistically with analysis of variance followed by post hoc Dunnett multiple range test. RESULTS: A single perisciatic injection of lidocaine (2%) produced relief of paw-pinch-evoked pain that was significantly different from baseline through to the 1-hour timepoint (Dunnett multiplicity-adjusted P = 0.0081), as assessed using paw withdrawal or vocalization end points. EN3427 (0.2%), in the same model, produced a long-lasting block, with pain thresholds being significantly above baseline through to the 18-hour timepoint (Dunnett multiplicity-adjusted P = 0.0002); the combination of EN3427 (0.2%) plus lidocaine (2%) produced even longer lasting analgesia, with pain thresholds being significantly above baseline through to the 24-hour timepoint (Dunnett multiplicity-adjusted P = 0.0073). Similar results were obtained with use of the pinprick approach. A single subcutaneous injection of lidocaine (2%) produced complete loss of sensation to cutaneous pinprick through 0.5 hours, but sensitivity thresholds were no different to baseline by the 1-hour timepoint, a similar injection of EN3427 alone (0.2%) produced a loss of sensation that was significantly different from baseline through the 8-hour timepoint (Dunnett multiplicity-adjusted P = 0.0045), and the combination of lidocaine (2%) plus EN3427 (0.2%) appeared to further enhance duration of analgesia, although this was significantly different from baseline only through the 10-hour timepoint (Dunnett multiplicity-adjusted P = 0.0048). Analgesic efficacy was dose related; using the combined injection approach, we found that increases in the dose of EN3427 with a fixed 2% lidocaine led to substantially extended analgesia and increasing doses of lidocaine combined with a fixed dose of EN3427 (0.2%) led to only modestly increased duration of action. CONCLUSIONS: The present studies demonstrate that a new molecular entity, EN3427, produces effective and long-lasting analgesia in 2 rodent pain models. The analgesic effects of EN3427 are significantly longer-lasting than lidocaine and are further extended when EN3427 is combined with lidocaine. The results are discussed with respect to a possible lidocaine-mediated TRP channel activation and facilitated neuronal access of EN3427, with subsequent entrapment conferring extended-duration efficacy.

Synonymous codon usage in forty staphylococcal phages identifies the factors controlling codon usage variation and the phages suitable for phage therapy.

The immergence and dissemination of multidrug-resistant strains of Staphylococcus aureus in recent years have expedited the research on the discovery of novel anti-staphylococcal agents promptly. Bacteriophages have long been showing tremendous potentialities in curing the infections caused by various pathogenic bacteria including S. aureus. Thus far, only a few virulent bacteriophages, which do not carry any toxin-encoding gene but are capable of eradicating staphylococcal infections, were reported. Based on the codon usage analysis of sixteen S. aureus phages, previously three phages were suggested to be useful as the anti-staphylococcal agents. To search for additional S. aureus phages suitable for phage therapy, relative synonymous codon usage bias has been investigated in the protein-coding genes of forty new staphylococcal phages. All phages appeared to carry A and T ending codons. Several factors such as mutational pressure, translational selection and gene length seemed to be responsible for the codon usage variation in the phages. Codon usage indeed varied phage to phage. Of the phages, phages G1, Twort, 66 and Sap-2 may be extremely lytic in nature as majority of their genes possess high translational efficiency, indicating that these phages may be employed in curing staphylococcal infections.

Reduced levels of cyclic AMP contribute to the enhanced oxidative stress in vascular smooth muscle cells from spontaneously hypertensive rats.

We have earlier shown that aortic vascular smooth muscle cells (VSMC) from 12-week-old spontaneously hypertensive rats (SHR) exhibited enhanced production of superoxide anion (O(2)(-)) compared with Wistar-Kyoto (WKY) rats. This production was attenuated to control levels by losartan, an angiotensin II (Ang II) AT(1)-receptor antagonist, suggesting that the AT(1) receptor is implicated in enhanced oxidative stress in SHR. Since AT(1) receptor activation signals via adenylyl cyclase inhibition and decreases cAMP levels, it is possible that AT(1) receptor-mediated decreased levels of cAMP contribute to the enhanced production of O(2)(-) in SHR. The present study was undertaken to investigate this possibility. The basal adenylyl cyclase activity as well as isoproterenol and forskolin-mediated stimulation of adenylyl cyclase was significantly attenuated in VSMC from 12-week-old SHR compared with those from WKY rats, whereas Ang II-mediated inhibition of adenylyl cyclase was significantly enhanced by about 70%, resulting in decreased levels of cAMP in SHR. NADPH oxidase activity and the levels of O2- were significantly higher (about 120% and 200%, respectively) in VSMC from SHR than from WKY rats. In addition, the levels of p47(phox) and Nox4 proteins, subunits of NADPH oxidase, were significantly augmented about 35%-40% in VSMC from SHR compared with those from WKY rats. Treatment of VSMC from SHR with 8Br-cAMP, as well as with cAMP-elevating agents such as isoproterenol and forskolin, restored to control WKY levels the enhanced activity of NADPH oxidase and the enhanced levels of O(2)(-), p47(phox), and Nox4. Furthermore, in the VSMC A10 cell line, 8Br-cAMP also restored the Ang II-evoked enhanced production of O(2)(-), NADPH oxidase activity, and enhanced levels of p47(phox) and Nox4 proteins to control levels. These data suggest that decreased levels of cAMP in SHR may contribute to the enhanced oxidative stress in SHR and that increasing the levels of cAMP may have a protective effect in reducing oxidative stress and thereby improve vascular function.

Activation of natriuretic peptide receptor-C attenuates the enhanced oxidative stress in vascular smooth muscle cells from spontaneously hypertensive rats: implication of Gialpha protein.

We have recently shown that vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) exhibit enhanced expression of Gialpha proteins, which was attributed to the enhanced oxidative stress. Since C-ANP(4-23) that specifically interacts with natriuretic peptide C (NPR-C) receptor has been shown to decrease the expression of Gialpha protein in VSMC, the present study was undertaken to examine if C-ANP(4-23) can also decrease the enhanced expression of Gialpha protein in VSMC from SHR and whether it is attributed to its ability to attenuate the enhanced oxidative stress. Aortic VSMC from 12-week-old SHR and their age-matched Wistar-Kyoto (WKY) rats were used for the present studies. VSMC from SHR exhibited enhanced expression of Gialpha-2 and Gialpha-3 proteins, different subunits of NADPH oxidase such as Nox(4) and p(47phox) proteins but not of p(22phox), enhanced production of superoxide anion as well as NADPH oxidase activity as compared to age-matched WKY rats. Treatment of VSMC from SHR with C-ANP(4-23) decreased towards control levels the enhanced expression of Gialpha proteins, enhanced superoxide anion production and enhanced NADPH oxidase activity as well as the enhanced expression of Nox(4) and p(47phox). However, C-ANP(4-23)-induced attenuation of the enhanced level of O(2)(-) and NADPH oxidase activity occurs at 4 h before the decrease in the enhanced expression of p(47phox) that occurs at 16 h of C-ANP(4-23) treatment. The decreased expression of NADPH oxidase in SHR was also associated with further decrease in O(2)(-) and NADPH oxidase activity. Furthermore, treatment of VSMC from SHR with pertussis toxin (PT) decreased the enhanced levels of superoxide anion as well as NADPH oxidase activity; however, the enhanced levels of different subunits of NADPH oxidase were not attenuated by PT treatment. These results suggest that C-ANP(4-23) decreases the enhanced oxidative stress in SHR by attenuating the enhanced expression of Gialpha proteins and also the enhanced levels of NADPH oxidase.