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Palmatine (PAL), a natural isoquinoline alkaloid, possesses extensive biological and pharmaceutical activities, including antioxidative stress, anti-inflammatory, antitumor, neuroprotective, and gastroprotective activities. However, it is unknown whether PAL has a protective effect against ischemic stroke and cerebral ischemia/reperfusion (I/R) injury. In the present study, a transient middle cerebral artery occlusion (MCAO) mouse model was used to mimic ischemic stroke and cerebral I/R injury in mice. Our study demonstrated that PAL treatment ameliorated cerebral I/R injury by decreasing infarct volume, neurological scores, and brain water content. PAL administration attenuated oxidative stress, the inflammatory response, and neuronal apoptosis in mice after cerebral I/R injury. In addition, PAL treatment also decreases hypoxia and reperfusion- (H/R-) induced neuronal injury by reducing oxidative stress, the inflammatory response, and neuronal apoptosis. Moreover, the neuroprotective effects of PAL were associated with the activation of the AMP-activated protein kinase (AMPK)/nuclear factor E2-related factor 2 (Nrf2) pathway, and Nrf2 knockdown offsets PAL-mediated antioxidative stress and anti-inflammatory effects. Therefore, our results suggest that PAL may be a novel treatment strategy for ischemic stroke and cerebral I/R injury.

A polymer-supported terpyridine palladium complex was prepared. The complex was found to promote hydrodechlorination of aryl chlorides with potassium formate in seawater. Generally, reductive cleavage of aryl chlorides using transition metal catalysts is more difficult than that of aryl bromides and iodides (reactivity: I > Br > Cl); however, the results obtained did not follow the general trend. Therefore, we investigated the reaction inhibition agents and found a method to remove these inhibitors. The polymeric catalysts showed high catalytic activity and high reusability for transfer reduction in seawater.

Trp222 of diketoreductase (DKR), an enzyme responsible for reducing a variety of ketones to chiral alcohols, is located at the hydrophobic dimeric interface of the C-terminus. Single substitutions at DKR Trp222 with either canonical (Val, Leu, Met, Phe and Tyr) or unnatural amino acids (UAAs) (4-cyano-L-phenylalanine, 4-methoxy-L-phenylalanine, 4-phenyl-L-phenyalanine, O-tert-butyl-L-tyrosine) inverts the enantiotope preference of the enzyme toward 2-chloro-1-phenylethanone with close side chain correlation. Analyses of enzyme activity, substrate affinity and ternary structure of the mutants revealed that substitution at Trp222 causes a notable change in the overall enzyme structure, and specifically in the entrance tunnel to the active center. The size of residue 222 in DKR is vital to its enantiotope preference. Trp222 serves as a \"gate keeper\" to control the direction of substrate entry into the active center. Consequently, opposite substrate-binding orientations produce respective alcohol enantiomers.

Background The bacterium Acetobacter sp. CCTCC M209061 is a promising whole-cell biocatalyst with exclusive anti-Prelog stereoselectivity for the reduction of prochiral ketones that can be used to make valuable chiral alcohols such as ( R )-4-(trimethylsilyl)-3-butyn-2-ol. Although it has promising catalytic properties, its stability and reusability are relatively poor compared to other biocatalysts. Hence, we explored various materials for immobilizing the active cells, in order to improve the operational stability of biocatalyst. Results It was found that Ca-alginate give the best immobilized biocatalyst, which was then coated with chitosan to further improve its mechanical strength and swelling-resistance properties. Conditions were optimized for formation of reusable immobilized beads which can be used for repeated batch asymmetric reduction of 4′-chloroacetophenone. The optimized immobilized biocatalyst was very promising, with a specific activity of 85% that of the free-cell biocatalyst (34.66 μ mol/min/g dw of cells for immobilized catalyst vs 40.54 μ mol/min/g for free cells in the asymmetric reduction of 4′-chloroacetophenone). The immobilized cells showed better thermal stability, pH stability, solvent tolerance and storability compared with free cells. After 25 cycles reaction, the immobilized beads still retained >50% catalytic activity, which was 3.5 times higher than degree of retention of activity by free cells reused in a similar way. The cells could be recultured in the beads to regain full activity and perform a further 25 cycles of the reduction reaction. The external mass transfer resistances were negligible as deduced from Damkohler modulus Da < <1, and internal mass transfer restriction affected the reduction action but was not the principal rate-controlling step according to effectiveness factors η < 1 and Thiele modulus 0.3< ∅ <1. Conclusions Ca-alginate coated with chitosan is a highly effective material for immobilization of Acetobacter sp. CCTCC M209061 cells for repeated use in the asymmetric reduction of ketones. Only a small cost in terms of the slightly lower catalytic activity compared to free cells could give highly practicable immobilized biocatalyst.

A primary mechanism for activation of innate immunity is recognition of damage or pathogen associated molecular patterns by pattern recognition receptors (PRRs). Nucleic acid is a damage associated molecular pattern molecule that when internalized into a monocyte and recognized by intracellular nucleic acid sensing toll like receptors will cause production of type 1 interferon. The process by which DNA or RNA is delivered into the cytosol of monocytes in systemic lupus erythematosus remains incompletely understood, and therapeutic approaches to prevent DNA-mediated monocyte activation are needed. We identified two mechanisms for internalization of DNA by monocytes. IgG-bound DNA was internalized by interacting with Fc gamma receptor IIa, while high-mobility group box-1 protein-bound DNA was internalized by interacting with the receptor for advanced glycation end products. Both pathways contribute to an inflammatory phenotype in monocytes exposed to serum from patients with SLE. Moreover, both of these pathways can be inhibited by a pentapeptide, DWEYS, which is a DNA mimetope. In one instance DWEYS directly competes with DNA for antibody binding and in the other DWEYS binds high-mobility group box-1 and blocks its interaction with RAGE. Our data highlight distinct pathways involved in nucleic acid enters monocytes in SLE, and identify a potential therapeutic to prevent nucleic acid internalization in SLE.

Mesenchymal stem cells (MSCs) have the function of repairing damaged tissue, which is known to be mediated by the secretome, the collection of secretory materials shed from MSCs. Adjusting the culture conditions of MSCs can lead to a significant difference in the composition of the secretome. It was hypothesized that pre-sensitization of MSCs with specific disease-causing agents could harness MSCs to release the therapeutic materials specialized for the disease. To validate this hypothesis, the present study aimed to generate a 'disease-specific secretome' for hepatitis caused by hepatitis B virus using hepatitis BX antigen (HBx) as a disease-causing material. Secretary materials (HBx-IS) were collected following the stimulation of adipose-derived stem cells (ASCs) with 100-fold diluted culture media of AML12 hepatocytes that had been transfected with pcDNA-HBx for 24 h. An animal model of hepatitis B was generated by injecting HBx into mice, and the mice were subsequently intravenously administered a control secretome (CS) or HBx-IS. Compared with the CS injection, the HBx-IS injection significantly reduced the serum levels of interleukin-6 and tumor necrosis factor-α (pro-inflammatory cytokines). Western blot analysis and immunohistochemistry of the liver specimens revealed that the HBx-IS injection led to a higher expression of liver regeneration-related markers, including hepatocyte growth factor and proliferating cell nuclear antigen, a lower expression of pro-apoptotic markers, such as cleaved caspase 3 and Bim in mouse livers, and a lower expression of pro-inflammatory markers (F4/80 and CD68) compared to the CS injection. HBx-IS exhibited higher liver regenerative, anti-inflammatory and anti-apoptotic properties, particularly in the mouse model of hepatitis B compared to CS. This suggests that the secretome obtained by stimulating ASCs with disease-causing agents may have a more prominent therapeutic effect on the specific disease than the naïve secretome.

Candida albicans (C. albicans ) is an important human commensal and opportunistic fungal pathogen. Secreted aspartyl proteinases (Saps) are a major virulence trait of C. albicans , and among these proteases Sap2 has the highest expression levels. It is possible that antibodies against Sap2 could provide an antifungal effect. In this study, two phages displaying anti-rSap2 single chain variable fragments (scFvs) were screened from human single fold scFv libraries, and their potential therapeutic roles were evaluated using a murine model infected by C. albicans . The in vivo efficacies were assessed by mortality rates, fungal burden and histological examination. Overall survival rates were significantly increased while the colony counts and infectious foci were significantly decreased after treatment with the scFv-phages relative to the control groups. In order to investigate the immune response provoked by scFv-phages, three kinds of cytokines (Th1, Th2 and Th17 types) were measured and a clear immune response was observed. These findings suggest that anti-rSap2 scFv-phages have potential in the therapy of systemic infection caused by C. albicans .

Background Enzymatic NADH or NADPH-dependent reduction is a widely applied approach for the synthesis of optically active organic compounds. The overall biocatalytic conversion usually involves in situ regeneration of the expensive NAD(P)H. Oxidation of formate to carbon dioxide, catalyzed by formate dehydrogenase (EC 1.2.1.2; FDH), presents an almost ideal process solution for coenzyme regeneration that has been well established for NADH. Because isolated FDH is relatively unstable under a range of process conditions, whole cells often constitute the preferred form of the biocatalyst, combining the advantage of enzyme protection in the cellular environment with ease of enzyme production. However, the most prominent FDH used in biotransformations, the enzyme from the yeast Candida boidinii , is usually expressed in limiting amounts of activity in the prime host for whole cell biocatalysis, Escherichia coli . We therefore performed expression engineering with the aim of enhancing FDH activity in an E. coli ketoreductase catalyst. The benefit resulting from improved NADH regeneration capacity is demonstrated in two transformations of technological relevance: xylose conversion into xylitol, and synthesis of ( S )-1-(2-chlorophenyl)ethanol from o -chloroacetophenone. Results As compared to individual expression of C. boidinii FDH in E. coli BL21 (DE3) that gave an intracellular enzyme activity of 400 units/g CDW , co-expression of the FDH with the ketoreductase ( Candida tenuis xylose reductase; XR) resulted in a substantial decline in FDH activity. The remaining FDH activity of only 85 U/g CDW was strongly limiting the overall catalytic activity of the whole cell system. Combined effects from increase in FDH gene copy number, supply of rare tRNAs in a Rosetta strain of E. coli , dampened expression of the ketoreductase, and induction at low temperature (18°C) brought up the FDH activity threefold to a level of 250 U/g CDW while reducing the XR activity by just 19% (1140 U/g CDW ). The E. coli whole-cell catalyst optimized for intracellular FDH activity showed improved performance in the synthesis of ( S )-1-(2-chlorophenyl)ethanol, reflected in a substantial, up to 5-fold enhancement of productivity (0.37 g/g CDW ) and yield (95% based on 100 mM ketone used) as compared to the reference catalyst. For xylitol production, the benefit of enhanced FDH expression was observed on productivity only after elimination of the mass transfer resistance caused by the cell membrane. Conclusions Expression engineering of C. boidinii FDH is an important strategy to optimize E. coli whole-cell reductase catalysts that employ intracellular formate oxidation for regeneration of NADH. Increased FDH-activity was reflected by higher reduction yields of D-xylose and o -chloroacetophenone conversions provided that mass transfer limitations were overcome.

The asymmetric reduction of 2-chloro-1-phenylethanone (1) by seven strains of marine fungi was evaluated and afforded (S)-(-)-2-chloro-1-phenylethanol with, in the best case, an enantiomeric excess of 50% and an isolated yield of 60%. The ability of marine fungi to catalyse the reduction was directly dependent on growth in artificial sea water-based medium containing a high concentration of Cl⁻ (1.2 M). When fungi were grown in the absence of artificial sea water, no reduction of 1 by whole cells was observed. The biocatalytic reduction of 1 was more efficient at neutral rather than acidic pH values and in the absence of glucose as co-substrate.

The excited delirium syndrome (EDS) is a life-threatening condition caused by a variety of factors including drug intoxication and psychiatric illness. Fatal instances of excited delirium frequently come to the attention of the medical examiner/coroner due to the circumstances and potential causes. Excited delirium may include paranoid, aggressive, and incoherent behavior which may lead to an encounter with law enforcement. In some instances, the person may die while in the presence of law enforcement. This circumstance further broadens the potential causes of death particularly as EDS has no pathognomonic autopsy finding. Although the syndrome of excited delirium is sufficient to explain death, other intervening causes need to be considered. These include chest or neck compression during restraint, blunt trauma, and underlying natural disease. Since chest/neck compression, natural disease (e.g., atherosclerosis), blunt trauma, and excited delirium are not mutually exclusive, all may be present in one death. The forensic pathologist’s role is to determine what caused and/or contributed to the death. When attempting to determine the proximate cause of death in instances with multiple potential causes, determining the mechanism of death often is useful. As not all causes of death have pathologically-demonstrable mechanisms of death, examination of the circumstances of the death often are diagnostically important. The main goal of the autopsy of deaths suspected to be due to EDS is to identify (or exclude) intervening diseases or injuries sufficient to explain the death in the context of the investigated circumstances.