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Background Recently, tranexamic acid (TXA) and epsilon aminocaproic acid (EACA) have been applied in total hip arthroplasty (THA). However, doubts in clinicians’ minds about which medicine is more efficient and economical in THA need to be clarified. Therefore, this study compared the efficacy and cost of the intraoperative administration of TXA and EACA per surgery in decreasing perioperative blood transfusion rates in THA. Methods This study enrolled patients who underwent THA between January 2019 to December 2020. A total of 295 patients were retrospectively divided to receive topical combined with intravenous TXA (n = 94), EACA (n = 97) or control (n = 104). The primary endpoints included transfusions, estimated perioperative blood loss, cost per patient and the drop in the haemoglobin and haematocrit levels. Results Patients who received EACA had greater total blood loss, blood transfusion rates, changes in HGB levels and mean cost of blood transfusion per patient (P P Conclusions The application of perioperative antifibrinolytics notably reduces the need for perioperative blood transfusions. What’s more, this study demonstrated that TXA is superior to EACA for decreasing blood loss and transfusion rates while at a lower cost per surgery. These results indicate that TXA may be the optimum antifibrinolytics for THA in Chinese area rather than EACA.

Background: The main goal of our study was to explore the wound-healing property of a novel cerium-containing N-acethyl-6-aminohexanoate acid compound and determine key molecular targets of the compound mode of action in diabetic animals. Methods: Cerium N-acetyl-6-aminohexanoate (laboratory name LHT-8-17) as a 10 mg/mL aquatic spray was used as wound experimental topical therapy. LHT-8-17 toxicity was assessed in human skin epidermal cell culture using (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. A linear wound was reproduced in 18 outbred white rats with streptozotocin-induced (60 mg/kg i.p.) diabetes, planar cutaneous defect was modelled in 60 C57Bl6 mice with streptozotocin-induced (200 mg/kg i.p.) diabetes and 90 diabetic db/db mice. Firmness of the forming scar was assessed mechanically. Skin defect covering was histologically evaluated on days 5, 10, 15, and 20. Tissue TNF-α, IL-1β and IL-10 levels were determined by quantitative ELISA. Oxidative stress activity was detected by Fe-induced chemiluminescence. Ki-67 expression and CD34 cell positivity were assessed using immunohistochemistry. FGFR3 gene expression was detected by real-time PCR. LHT-8-17 anti-microbial potency was assessed in wound tissues contaminated by MRSA. Results: LHT-8-17 4 mg twice daily accelerated linear and planar wound healing in animals with type 1 and type 2 diabetes. The formulated topical application depressed tissue TNF-α, IL-1β, and oxidative reaction activity along with sustaining both the IL-10 concentration and antioxidant capacity. LHT-8-17 induced Ki-67 positivity of fibroblasts and pro-keratinocytes, upregulated FGFR3 gene expression, and increased tissue vascularization. The formulation possessed anti-microbial properties. Conclusions: The obtained results allow us to consider the formulation as a promising pharmacological agent for diabetic wound topical treatment.

Bioplastics produced from microbial source are promising green alternatives to traditional petrochemical-derived plastics. Nonnatural straight-chain amino acids, especially 5-aminovalerate, 6-aminocaproate and 7-aminoheptanoate are potential monomers for the synthesis of polymeric bioplastics as their primary amine and carboxylic acid are ideal functional groups for polymerization. Previous pathways for 5-aminovalerate and 6-aminocaproate biosynthesis in microorganisms are derived from L-lysine catabolism and the citric acid cycle, respectively. Here, we show the construction of an artificial iterative carbon-chain-extension cycle in Escherichia coli for simultaneous production of a series of nonnatural amino acids with varying chain length. Overexpression of L-lysine α-oxidase in E. coli yields 2-keto-6-aminocaproate (2K6AC) as a non-native substrate for the artificial iterative carbon-chain-extension cycle. The chain-extended α-ketoacid products are decarboxylated and oxidized by an α-ketoacid decarboxylase and an aldehyde dehydrogenase, respectively, to yield their corresponding nonnatural straight-chain amino acids. The engineered system demonstrated simultaneous in vitro production of 99.16 mg/L of 5-aminovalerate, 46.96 mg/L of 6-aminocaproate and 4.78 mg/L of 7-aminoheptanoate after 8 h of enzyme catalysis starting from 2K6AC as the substrate. Furthermore, simultaneous production of 2.15 g/L of 5-aminovalerate, 24.12 mg/L of 6-aminocaproate and 4.74 mg/L of 7-aminoheptanoate was achieved in engineered E. coli. This work illustrates a promising metabolic-engineering strategy to access other medium-chain organic acids with –NH2, –SCH3, –SOCH3, –SH, –COOH, –COH, or –OH functional groups through carbon-chain-elongation chemistry., Fundamental Research Funds for the Central Universities of China (Project 106112017CDJXFLX0014), Fundamental Research Funds for the Central Universities of China (Project 2018CDQYHG0010), Tianjin University. Science and Technology Support Program (Grant 15PTCYSY00020), Chinese Academy of Sciences. Research Equipment Program (Grant (YJKYYQ20170023), Chinese Academy of Sciences. Henan Provincial Science and Technology Open Cooperation Project (162106000014)

Current methods for assessing health capital are not accessible to clinicians. To increase accessibility, we evaluated a Brief Adult Health Capital Scale (BAHCS-10) using classical and modern testing theories. With 588 clients, we found an adequate fit for the BAHCS-10[Formula: see text], CFIscaled = 0.949, TLIscaled = 0.935, RMSEA = 0.077, and the SRMR = 0.060. We also found evidence of invariance across race but did find significant non-invariance across some items for gender and age. Future researchers should review items displaying noninvariance and develop optimal cut scores for the BAHCS-10 to further support clinician decision making.

In an attempt to understand the possibility of applications of the fullerene-based systems for transporting various polar compounds like hexamethonium through the blood-brain barrier, we studied the influence of a series of derivatives of fullerene C

From a collection of marine cyanobacteria made in the Coiba National Park along the Pacific coast of the Republic of Panama a novel cyclic depsipeptide, given the trivial name medusamide A, has been isolated and fully characterized. Medusamide A contains four contiguous β-amino acid (2R,3R)-3-amino-2-methylhexanoic acid (Amha) residues. This is the first report of multiple Amha residues and contiguous β-amino acid residues within a single cyclic peptide-type natural product. Stereochemical assignment of the Amha residues was completed following the synthesis of reference standards for this β-amino acid and the subsequent derivatization with Marfey's reagent and LC-MS analysis.

Human arginase I (hARGI) is an important enzyme involved in the urea cycle, its overexpression has been associated to cardiovascular and cerebrovascular diseases. In the last years, several congeneric sets of hARGI inhibitors have been reported with possible beneficial roles for the cardiovascular system. At the same time, crystallographic data have been reported including hARGI&ndash, inhibitor complexes, which can be considered for the design of novel inhibitors. In this work, the structure&ndash, activity relationship (SAR) of C&alpha, substituted 2(S)-amino-6-boronohexanoic acid (ABH) derivatives as hARGI inhibitors was studied by using a three-dimensional quantitative structure&ndash, activity relationships (3D-QSAR) method. The predictivity of the obtained 3D-QSAR model was demonstrated by using internal and external validation experiments. The best model revealed that the differential hARGI inhibitory activities of the ABH derivatives can be described by using steric and electrostatic fields, the local effects of these fields in the activity are presented. In addition, binding modes of the above-mentioned compounds inside the hARGI binding site were obtained by using molecular docking. It was found that ABH derivatives adopted the same orientation reported for ABH within the hARGI active site, with the substituents at C&alpha, exposed to the solvent with interactions with residues at the entrance of the binding site. The hARGI residues involved in chemical interactions with inhibitors were identified by using an interaction fingerprints (IFPs) analysis.

Aim: Zinc has proved its efficacy in many models of ischemia reperfusion (I/R) injury. In this study, we used zinc acexamate (ZAC) as an exogenous source of zinc against renal I/R injury and we investigated whether its protective effects are mediated by the decrease of oxidative stress, inflammation, and mitochondria inducedapoptosis. Methods: Rats were orally pretreated with vehicle or ZAC (10 or 100 mg/kg) 24 h and 30 min prior to 1 h of bilateral renal warm ischemia and 2 h of reperfusion. Results: Our data showed that 10 mg/kg of ZAC, but not 100 mg/kg, improved renal architecture and function. Also, the low dose of ZAC up-regulated antioxidant enzymes activities and glutathione level and decreased lipids and proteins oxidation. Interestingly, the use of ZAC resulted in a significant reduce of pro-inflammatory cytokines (IL-1ß, IL-6 and MCP-1), enhanced mitochondria integrity and decreased expression of the pro-apoptotic protein caspase-9. Conclusion: We conclude that renal I/R induced oxidative stress, inflammation and apoptosis and that the use of ZAC at 10 mg/kg, but not 100 mg/kg, protects rat kidneys from I/R injury by down-regulating these processes., This work was supported by grants from: The Tunisian Ministry of Higher Education and Scientific Research (UR12ES11); SAF-2014- 57674-R, SAF-2015-69944-R from Plan Nacional de I + D, Spain and CIBEREHD; the center grant P50AA011999 Southern California Research Center for ALPD and Cirrhosis funded by NIAAA/NIH; and AGAUR of the Generalitat de Catalunya2014-SGR785.

Histone deacetylases (HDACs) play essential roles in transcription regulation and are valuable theranostic targets. However, there are no activatable fluorescent probes for imaging of HDAC activity in live cells. Here, we develop for the first time a novel activatable two-photon fluorescence probe that enables in situ imaging of HDAC activity in living cells and tissues. The probe is designed by conjugating an acetyl-lysine mimic substrate to a masked aldehyde-containing fluorophore via a cyanoester linker. Upon deacetylation by HDAC, the probe undergoes a rapid self-immolative intramolecular cyclization reaction, producing a cyanohydrin intermediate that is spontaneously rapidly decomposed into the highly fluorescent aldehyde-containing two-photon fluorophore. The probe is shown to exhibit high sensitivity, high specificity, and fast response for HDAC detection in vitro. Imaging studies reveal that the probe is able to directly visualize and monitor HDAC activity in living cells. Moreover, the probe is demonstrated to have the capability of two-photon imaging of HDAC activity in deep tissue slices up to 130 μm. This activatable fluorescent probe affords a useful tool for evaluating HDAC activity and screening HDAC-targeting drugs in both live cell and tissue assays.

The biobased production of adipic acid, a precursor in the production of nylon, is of great interest in order to replace the current petrochemical production route. Glucose-rich lignocellulosic raw materials have high potential to replace the petrochemical raw material. A number of metabolic pathways have been proposed for the microbial conversion of glucose to adipic acid, but achieved yields and titers remain to be improved before industrial applications are feasible. One proposed pathway starts with lysine, an essential metabolite industrially produced from glucose by microorganisms. However, the drawback of this pathway is that several reactions are involved where there is no known efficient enzyme. By changing the order of the enzymatic reactions, we were able to identify an alternative pathway with one unknown enzyme less compared to the original pathway. One of the reactions lacking known enzymes is the reduction of the unsaturated α,β bond of 6-amino-trans-2-hexenoic acid and trans-2-hexenedioic acid. To identify the necessary enzymes, we selected N-ethylmaleimide reductase from Escherichia coli and Old Yellow Enzyme 1 from Saccharomyces pastorianus. Despite successful in silico docking studies, where both target substrates could fit in the enzyme pockets, and hydrogen bonds with catalytic residues of both enzymes were predicted, no in vitro activity was observed. We hypothesize that the lack of activity is due to a difference in electron withdrawing potential between the naturally reduced aldehyde and the carboxylate groups of our target substrates. Suggestions for protein engineering to induce the reactions are discussed, as well as the advantages and disadvantages of the two metabolic pathways from lysine. We have highlighted bottlenecks associated with the lysine pathways, and proposed ways of addressing them.

Antibacterial monomers can combat oral biofilm acids and caries; however, little is known on whether quaternary ammonium monomers (QAMs) would induce drug persistence in oral bacteria. The objectives of this study were to investigate the interactions of Streptococcus mutans (S. mutans) with dimethylaminohexadecyl methacrylate (DMAHDM), and determine for the first time whether DMAHDM could induce persisters in S. mutans. DMAHDM was synthesized using a modified Menschutkin reaction. Dose-dependent killing curves and time-dependent killing curves of planktonic S. mutans and biofilms were determined to evaluate drug persistence, using chlorhexidine (CHX) as control. The inheritability assay, minimum inhibitory concentration (MIC) and live/dead biofilm assay were determined to investigate persister characteristics. DMAHDM matched the killing potency of the gold standard CHX against S. mutans biofilms. DMAHDM and CHX induced drug persistence in S. mutans biofilms but not in planktonic bacteria. S. mutans biofilm persistence was not inheritable in that the tolerance to DMAHDM or CHX of the surviving persisters in the initial population was not transferred to subsequent generations, as displayed by the inheritability assay. The MIC of S. mutans parental strain and induced persisters remained the same. The induced persisters in S. mutans biofilms could be eliminated via higher doses of 300 μg/mL of DMAHDM and CHX. In conclusion, this study showed for the first time that (1) DMAHDM induced persisters only in biofilms, but not in planktonic bacteria; and (2) both DMAHDM-induced and CHX-induced S. mutans persister biofilms could be completely eradicated by even higher concentrations of DMAHDM and CHX. More studies are needed on the induction of persisters in oral biofilms for the development and use of a new generation of antibacterial dental monomers and resins.

Plasminogen (Plg) circulates in the host as two predominant glycoforms. Glycoform I Plg (GI-Plg) contains glycosylation sites at Asn289 and Thr346, whereas glycoform II Plg (GII-Plg) is exclusively glycosylated at Thr346. Surface plasmon resonance experiments demonstrated that Plg binding group A streptococcal M protein (PAM) exhibits comparative equal affinity for GI- and GII-Plg in the "closed" conformation (for GII-Plg, KD = 27.4 nM; for GI-Plg, KD = 37.0 nM). When Plg was in the "open" conformation, PAM exhibited an 11-fold increase in affinity for GII-Plg (KD = 2.8 nM) compared with that for GI-Plg (KD = 33.2 nM). The interaction of PAM with Plg is believed to be mediated by lysine binding sites within kringle (KR) 2 of Plg. PAM-GI-Plg interactions were fully inhibited with 100 mM lysine analogue ε-aminocaproic acid (εACA), whereas PAM-GII-Plg interactions were shown to be weakened but not inhibited in the presence of 400 mM εACA. In contrast, binding to the KR1-3 domains of GII-Plg (angiostatin) by PAM was completely inhibited in the presence 5 mM εACA. Along with PAM, emm pattern D GAS isolates express a phenotypically distinct SK variant (type 2b SK) that requires Plg ligands such as PAM to activate Plg. Type 2b SK was able to generate an active site and activate GII-Plg at a rate significantly higher than that of GI-Plg when bound to PAM. Taken together, these data suggest that GAS selectively recruits and activates GII-Plg. Furthermore, we propose that the interaction between PAM and Plg may be partially mediated by a secondary binding site outside of KR2, affected by glycosylation at Asn289.

Arginase is an enzyme that limits substrate l-arginine bioavailability for the production of nitric oxide by the nitric oxide synthases and produces l-ornithine, which is a precursor for collagen formation and tissue remodeling. We studied the pulmonary vascular effects of arginase inhibition in an established model of repeated systemic bleomycin sulfate administration in neonatal rats that results in pulmonary hypertension and lung injury mimicking the characteristics typical of bronchopulmonary dysplasia. We report that arginase expression is increased in the lungs of bleomycin-exposed neonatal rats and that treatment with the arginase inhibitor amino-2-borono-6-hexanoic acid prevented the bleomycin-induced development of pulmonary hypertension and deposition of collagen. Arginase inhibition resulted in increased l-arginine and l-arginine bioavailability and increased pulmonary nitric oxide production. Arginase inhibition also normalized the expression of inducible nitric oxide synthase, and reduced bleomycin-induced nitrative stress while having no effect on bleomycin-induced inflammation. Our data suggest that arginase is a promising target for therapeutic interventions in neonates aimed at preventing lung vascular remodeling and pulmonary hypertension.

Aberrant excitatory neurotransmission associated with overproduction of glutamate has been implicated in the development of HIV-associated neurocognitive disorders (HAND). The glutamine antagonist 6-diazo-5-oxo-L-norleucine (DON, 14) attenuates glutamate synthesis in HIV-infected microglia/macrophages, offering therapeutic potential for HAND. We show that 14 prevents manifestation of spatial memory deficits in chimeric EcoHIV-infected mice, a model of HAND. 14 is not clinically available, however, because its development was hampered by peripheral toxicities. We describe the synthesis of several substituted N-(pivaloyloxy)alkoxy-carbonyl prodrugs of 14 designed to circulate inert in plasma and be taken up and biotransformed to 14 in the brain. The lead prodrug, isopropyl 6-diazo-5-oxo-2-(((phenyl(pivaloyloxy)methoxy)-carbonyl)amino)hexanoate (13d), was stable in swine and human plasma but liberated 14 in swine brain homogenate. When dosed systemically in swine, 13d provided a 15-fold enhanced CSF-to-plasma ratio and a 9-fold enhanced brain-to-plasma ratio relative to 14, opening a possible clinical path for the treatment of HAND.

Five bacterial strains with the unique ability to utilize low-molecular linear caprolactam olygomers (nylon olygomers) were isolated from soil samples contaminated with industrial wastes of epsilon-caprolactam. Based on the properties studied and also on the analysis of 16S rRNA gene nucleotide sequences, the strains BS2,BS3, BS9, BS38, and BS57 were classified to the general Arthrobacter, Brevibacterium, Microbacteriun, Gulosibacter, and Achromobacter, respectively. All of the strains also utilized 6-aminohexanoic and adipic acids, which are intermidiates of the epsilon-caprolactam catabolism. This indirectly points to the fact that degradation of olygomers in these bacteria occurs via the monomer degradation pathway. The BS9 and BS57 strains utilized only olygomers of the epsilon-caprolactam, while BS2, BS3, and BS38 also degraded epsilon-caprolactam and its homologs, enantolactam and caprylolactam, which differentiates the latter from the previously known degraders of olygomers and suggests the presence in these strains of enzymes with lactam hydrolase activity, in addition to 6-aminohexanoate-dimer hydrolase.

Lysine is frequently a first- or second-limiting amino acid in poultry diets. Improving the efficiency of lysine use for protein synthesis would effectively lower the lysine requirement and decrease feed costs. Understanding how lysine is degraded and how the degradation is regulated would identify potential molecular targets for interventions to decrease lysine degradation. To better understand lysine degradation in poultry, 3 experiments were conducted. In experiment 1, one-day-old chicks were fed 1.07, 1.25, 1.73, or 3.28% dietary lysine for 2 wk. In experiments 2 and 3, fourteen-day-old chicks were fed 1.07 or 1.25% dietary lysine for 2 wk. Measures of liver lysine catabolism including lysine α-ketoglutarate reductase (LKR) and lysine oxidation (LOX) were assessed. The α-aminoadipate δ-semialdehyde synthase (AASS) is a bifunctional enzyme composed of both LKR and saccharopine dehydrogenase activities, and the relative abundance of this protein and mRNA were likewise assessed. Moreover, potential alternative pathways of lysine catabolism that depend on l-amino acid oxidase (AAOX) and on lysyl oxidase (LYLOX) were considered. In experiment 1, chicks fed lysine-deficient diets had decreased (P0.05) LKR activities compared with chicks fed at or above the requirement. However, the lowered LKR activities were not associated with a decreased (P0.05) LOX as measured in vitro. In experiments 2 and 3, chicks 28 d of age did not decrease LKR activity (P0.05) in response to a lysine-deficient diet. No changes in AASS protein abundance or mRNA were detected. Likewise, no differences in the mRNA abundances of AAOX or LYLOX were detected. The activity of AAOX did increase (P0.05) in birds fed a lysine-adequate diets compared with those fed a lysine-deficient diet. Based on kinetic parameters and assumed concentrations, AAOX could account for about 20% of liver lysine oxidation in avians.

The Ugi reaction has been successfully applied to the synthesis of novel arginase inhibitors. In an effort to decrease conformational flexibility of the previously reported series of 2-amino-6-boronohexanoic acid (ABH) analogs 1, we designed and synthesized a series of compounds, 2, in which a piperidine ring is linked directly to a quaternary amino acid center. Further improvement of in vitro activity was achieved by adding two carbon bridge in the piperidine ring, that is, tropane analogs 11. These improvements in activity are rationalized by X-ray crystallography analysis, which show that the tropane ring nitrogen atom moves into direct contact with Asp202 (arginase II numbering). The synthetic routes described here enabled the design of novel arginase inhibitors with improved potency and markedly different physico-chemical properties compared to ABH. Compound 11c represents the most in vitro active arginase inhibitor reported to date.

Objectives To illustrate that matching on provider may exacerbate, not remove, bias. Study Design and Setting The degree of confounding bias depends in part on the proportions of treatment variation that can be ascribed to confounders and to instruments, respectively. This commentary raises the specific example of bias by matching on hospital induced in a study of coronary artery bypass graft surgery patients and illustrates the effect of matching on provider in a constructed example. Results Matching on provider removes a “benign” source of treatment variability, leaving unmeasured confounders as potentially the most important determinants of treatment. Conclusions Researchers need to articulate the presumed source of pseudorandom variation in observational studies and need to take care not to reduce their effect through unnecessary control.

Substitution at the alpha center of the known human arginase inhibitor 2-amino-6-boronohexanoic acid (ABH) is acceptable in the active site pockets of both human arginase I and arginase II. In particular, substituents with a tertiary amine linked via a two carbon chain show improved inhibitory potency for both enzyme isoforms. This potency improvement can be rationalized by X-ray crystallography, which shows a water-mediated contact between the basic nitrogen and the carboxylic acid side chain of Asp200, which is situated at the mouth of the active site pocket of arginase II (Asp181 in arginase I). We believe that this is the first literature report of compounds with improved arginase inhibitory activity, relative to ABH, and represents a promising starting point for further optimization of in vitro potency and the identification of better tool molecules for in vivo investigations of the potential pathophysiological roles of arginases.

A first synthetic effort toward total synthesis of highly potent solomonamides is disclosed. An efficient strategy to synthesize this class of compounds, along with the synthesis of a core macrocycle (shown in red) and the key fragment AHMOA, is described.

Objective: Thoracentesis with chest tube placement is often needed to decompress a clinically significant pneumothorax or pleural effusion. The risks of such a procedure may be considered too great to perform on a systemically anticoagulated patient supported by extracorporeal membrane oxygenation (ECMO). Results: An 8-year-old child with respiratory failure due to necrotizing pneumonia and autoimmune vasculitis, on veno-venous ECMO, developed a severe tension pneumothorax that required emergent decompression with a chest tube. Post-procedure, the patient developed a hemothorax that was approaching non-sustainability. We developed a strategy based on Virchow’s triad to favor homeostasis in the patient while avoiding thrombosis in the ECMO circuit. We employed selective lung ventilation, passive pleural drainage, high flow ECMO, and aggressive coagulation cascade control, including the use of aminocaproic acid and activated factor VIIa. Following this strategy, the hemorrhage was controlled and, later, the patient was able to successfully come off ECMO. Conclusions: With careful coagulation cascade manipulation, complete lung rest for the affected lung, control of ECMO blood flow, and prudent hemothorax drainage, we were able to facilitate hemostasis that was required for the successful recovery of our patient while avoiding critical ECMO circuit thrombosis. Even with today’s highly advanced medical technologies, centuries-old basic medical principles can still assist in the care of our sickest and most complex patients. Chest tube placement while on ECMO is rare and, although necessary, may be a risky procedure. With precise coagulation control, it can be a successful procedure on ECMO.

Various binuclear metal ion clusters and complexes have been reconstituted in crystalline human arginase I by removing the Mn(2+)(2) cluster of the wild-type enzyme with metal chelators and subsequently soaking the crystalline apoenzyme in buffer solutions containing NiCl(2) or ZnCl(2). X-ray crystal structures of these metal ion variants are correlated with catalytic activity measurements that reveal differences resulting from metal ion substitution. Additionally, treatment of crystalline Mn(2+)(2)-human arginase I with Zn(2+) reveals for the first time the structural basis for inhibition by Zn(2+), which forms a carboxylate-histidine-Zn(2+) triad with H141 and E277. The imidazole side chain of H141 is known to be hyper-reactive, and its chemical modification or mutagenesis is known to similarly compromise catalysis. The reactive substrate analogue 2(S)-amino-6-boronohexanoic acid (ABH) binds as a tetrahedral boronate anion to Mn(2+)(2), Co(2+)(2), Ni(2+)(2), and Zn(2+)(2) clusters in human arginase I, and it can be stabilized by a third inhibitory Zn(2+) ion coordinated by H141. Because ABH binds as an analogue of the tetrahedral intermediate and its flanking transition states in catalysis, this implies that the various metallo-substituted enzymes are capable of some level of catalysis with an actual substrate. Accordingly, we establish the following trend for turnover number (k(cat)) and catalytic efficiency (k(cat)/K(M)): Mn(2+)Ni(2+) ≈ Co(2+) ≫ Zn(2+). Therefore, Mn(2+) is required for optimal catalysis by human arginase I.

Arginase expression and activity have been noted to be heightened in conditions associated with erectile dysfunction, including aging. Previously, arginase inhibition by chronic administration of the arginase inhibitor 2-(S)-amino-6-boronohexanoic acid (ABH) has been shown to improve endothelial dysfunction in aged rats. The objective of this study was to assess whether chronic oral ABH administration affects cavernosal erectile function. Rats were divided into 4 groups: young control, young treated with arginase inhibitor, aged control, and aged treated with arginase inhibitor. Arginase activity was measured and presented as a proportion of young untreated rats. In vivo erectile responses to cavernous nerve stimulation were measured in all cohorts. The cavernous nerve was stimulated with a graded electrical stimulus, and the intracavernosal/mean arterial pressure ratios and total intracavernosal pressure were recorded. Arginase activity was elevated in the aged rats compared with young controls; however, arginase activity was significantly decreased in aged rats treated with ABH. With the addition of ABH, erectile responses improved in the aged rats (P.05). Oral inhibition of arginase with ABH results in improved erectile function in aged rats, resulting in erectile hemodynamics similar to young rats. This represents the first documentation of systemic arginase inhibition positively affecting corporal cavernosal function.

Objective Arginase plays important regulatory roles in polyamine, ornithine, and nitric oxide syntheses. However, its role in the healing process has not been delineated. In this study, we used a highly potent and specific inhibitor of arginase, namely 2(S)-amino-6-boronohexanoic acid NH4 (ABH) to evaluate the role of arginase function in wound healing. Materials and Methods ABH or saline was applied topically to full thickness, dorsal, excisional wounds in C57BL/6 mice every 8 hours for 14 days post surgery and the rate of wound closure was estimated planimetrically. Wound tissue was harvested from mice sacrificed on postoperative days 3 and 7 and examined histologically. The extent of epithelial, connective, and granulation tissue present within the wound area was estimated histomorphometrically. The effect of ABH on wound arginase activity, production of nitric oxide metabolites (NO x ), and presence of smooth muscle actin positive cells (myofibroblasts) was evaluated. Results While arginase activity was inhibited in vivo, the rate of wound closure significantly increased 7 days post-surgery, (21 ± 4%: P t test) in ABH treated animals. This was accompanied by an early increase in wound granulation tissue and accumulation of NO x followed by enhanced re-epithelialization and localization of myofibroblasts beneath the wound epithelium. Conclusion Arginase inhibition improves excisional wound healing and may be used to develop therapeutics for early wound closure.

Macrophages, upon phagocytosing endospores of Bacillus anthracis, up-regulate the expression of the immunological isoform of nitric oxide synthase, NOS 2, concomitant with production of nitric oxide (NO•) from metabolism of l-arginine. We have previously demonstrated that macrophages that secrete NO• kill the bacilli of B. anthracis. To circumvent this microbicidal activity of NO•, B. anthracis has evolved pathways that include the enzyme arginase, which metabolizes l-arginine to ornithine and urea. Compounds that inhibit arginase might, therefore, offer a therapeutic approach to controlling B. anthracis infection. 2(S)-Amino-6-boronohexanoic acid (ABH) has been reported to be an inhibitor of mammalian arginase. In this study, we explore the inhibitory effect of ABH against B. anthracis arginase and its potential for future development, as an effective therapeutic agent against microbial infection. We found that ABH is an inhibitor of bacterial arginase in several different endospore strains of B. anthracis. Further, ABH inhibits neither the phagocytosis of these endospores nor the up-regulation of NOS 2 concomitant with secretion of NO•. These findings set the stage to determine how efficacious ABH will be in promoting NO•-mediating killing of B. anthracis.

The present report describes development of hexamethonium complexes based on fullerene C60. Hexamethonium has a limited penetration into CNS and therefore can antagonize central effects of nicotine only when given at high doses. In the present studies conducted in laboratory rodents, intraperitoneal administration of hexamethonium-fullerene complexes blocked effects of nicotine (convulsions and locomotor stimulation). When compared to equimolar doses of hexamethonium, complexes of hexamethonium with derivatives of fullerene C60 were 40 times more potent indicating an enhanced ability to interact with central nicotine receptors. Thus, fullerene C60 derivatives should be explored further as potential carrier systems for polar drug delivery into CNS.

Airway remodelling, characterised by increased airway smooth muscle (ASM) mass, subepithelial fibrosis, goblet cell hyperplasia and mucus gland hypertrophy, is a feature of chronic asthma. Increased arginase activity could contribute to these features via increased formation of polyamines and L-proline downstream of the arginase product L-ornithine, and via reduced nitric oxide synthesis.Using the specific arginase inhitibor 2(S)-amino-6-boronohexanoic acid (ABH), we studied the role of arginase in airway remodelling using a guinea pig model of chronic asthma. Ovalbumin-sensitised guinea pigs were treated with ABH or PBS via inhalation before each of 12 weekly allergen or saline challenges, and indices of arginase activity, and airway remodelling, inflammation and responsiveness were studied 24 h after the final challenge.Pulmonary arginase activity of repeatedly allergen-challenged guinea pigs was increased. Allergen challenge also increased ASM mass and maximal contraction of denuded tracheal rings, which were prevented by ABH. ABH also attenuated allergen-induced pulmonary hydroxyproline (fibrosis) and putrescine, mucus gland hypertrophy, goblet cell hyperplasia, airway eosinophilia and interleukin-13, whereas an increased L-ornithine/L-citrulline ratio in the lung was normalised. Moreover, allergen-induced hyperresponsiveness of perfused tracheae was fully abrogated by ABH.These findings demonstrate that arginase is prominently involved in allergen-induced airway remodelling, inflammation and hyperresponsiveness in chronic asthma.

After both the intravenous and oral administration of zinc acexamate [ZAC; ion-pairing between zinc and ϵ-acetamidocaproic acid (AACA)] and cimetidine together, the areas under the curve (AUCs) of AACA were significantly greater [by 28.2 and 98.9% after the intravenous and oral administration, respectively, for control rats and 13.5 and 16.9% for indomethacin-induced acute gastric ulcer (IAGU) rats, respectively] than those of ZAC alone due to the significantly slower renal clearance (CL(R)). The significantly greater AUCs of AACA after both the intravenous and oral administration of ZAC and cimetidine together in control and IAGU rats could have been due to the inhibition of active renal tubular secretion of AACA by cimetidine. After the intravenous and oral administration of both drugs together, the AUCs of cimetidine in control and IAGU rats were not different compared with those with cimetidine alone.

The preparation and the antibacterial activity of alaremycin derivatives such as their CF3-derivatives and (R)- and (S)-4-oxo-5-acetylaminohexanoic acid for the porphobilinogen synthase (PBGS), were described. The IC50 values of the antibacterial activity of the prepared materials for the inhibitor of PBGS, were determined using PBGS assay.

Objectives Intraperitoneal (IP) injection of 3.5 g/kg L-arginine (known to induce acute pancreatitis) in rats will result in much greater increases in serum ornithine versus citrulline concentration (Crit Care Med. 2008;36:2117-2127). These data indicate a major role of arginase in the catabolism of L-arginine. Therefore, we tested the effects of the irreversible arginase inhibitor (+)-S-2-amino-6-iodoacetamidohexanoic acid (AIHA) on L-arginine-induced acute pancreatitis. Methods The inhibitory effect of AIHA on arginase activity was tested on rat liver homogenate and purified bovine arginase. Male Wistar rats were administered 15 mg/kg AIHA or its vehicle IP 1 hour before the injection of physiological saline or 3.5 g/kg L-arginine IP. Laboratory and histological parameters of pancreatitis were determined 24 hours after the last injection. Results Sixty micromolars of AIHA (equimolar to an in vivo dose of 15 mg/kg) significantly inhibited arginase activity by about 25%. Pretreatment with AIHA significantly ameliorated pancreatic damage caused by L-arginine administration. It decreased pancreatic weight/body weight ratio, pancreatic glutathione peroxidase and myeloperoxidase activities, and histological damage. Administration of AIHA in itself significantly increased levels of pancreatic heat shock proteins. Conclusions Pretreatment with AIHA reduces the severity of L-arginine-induced pancreatitis most likely by inhibiting arginase activity.

The spirochete Leptospira interrogans is a highly invasive pathogen of worldwide public health importance. Studies from our laboratories and another have demonstrated that L. interrogans can acquire host plasminogen on its surface. Exogenous plasminogen activators can then convert bound plasminogen into the functionally active protease plasmin. In this study, we extend upon those observations and report that leptospiral endostatin-like protein A (LenA) binds human plasminogen in a dose-dependent manner. LenA-plasminogen interactions were significantly inhibited by the lysine analog ξ-aminocaproic acid, suggesting that the lysine-binding sites on the amino-terminal kringle portion of the plasminogen molecule play a role in the binding. Previous studies have shown that LenA also binds complement regulator factor H and the extracellular matrix component laminin. Plasminogen competed with both factor H and laminin for binding to LenA, which suggests overlapping ligand-binding sites on the bacterial receptor. Finally, LenA-bound plasminogen could be converted to plasmin, which in turn degraded fibrinogen, suggesting that acquisition of host-derived plasmin by LenA may aid bacterial dissemination throughout host tissues.

Background: Aprotinin is the only Food and Drug Administration-approved agent to reduce haemorrhage related to cardiac surgery and its safety and efficacy has been extensively studied. Our study sought to compare the efficacy, early and late mortality and major morbidity associated with aprotinin compared with e-aminocaproic acid (EACA) in cardiac surgery operations. Methods: Between January 2002 and December 2006, 2101 patients underwent coronary artery bypass grafting (CABG), valve surgery or CABG and valve surgery in our institution with the use of aprotinin (1898 patients) or EACA (203 patients).Logistic regression and propensity score analysis were usedto adjust for imbalances in the patients’ preoperative characteristics. The propensity score-adjusted sample included 570 patients who received aprotinin and 114 who receivedEACA (1—5matching).Results:Operativemortalitywashigher in theaprotinin groupin univariate(aprotinin4.3% vs EACA1%,p = 0.023) butnotpropensityscore-adjustedmultivariateanalysis(4%vs0.9%,p = 0.16).Inpropensity score-adjustedanalysis,aprotininwasalso associated with a lower rate of blood transfusion (38.8% vs 50%, p = 0.04), a lower rate of haemorrhage-related re-exploration (3.7% vs 7.9%, p = 0.04) and a higher risk of in-hospital cardiac arrest (3.7% vs 0%, p = 0.03) and a marginally but not statistically significantly higher risk of acute renal failure (6.8% vs 2.6%,p = 0.09). In Cox proportional hazardsregression analysis, the riskof late death was higher in the aprotinin comparedto EACA group (hazard ratio = 4.33, 95% confidence interval (CI) = 1.60—11.67, p = 0.004). Conclusion: Aprotinin decreases the rate of postoperative blood transfusion and haemorrhage-related re-exploration, but increases the risk of in-hospital cardiac arrest and late mortality after cardiac surgery when compared to EACA. Cumulative evidence suggests that the risk associated with aprotinin may not be worth the haemostatic benefit. # 2009 European Association for Cardio-Thoracic Surgery. Published by Elsevier B.V. All rights reserved.

There is increasing evidence that upregulation of arginase contributes to impaired endothelial function in aging. In this study, we demonstrate that arginase upregulation leads to endothelial nitric oxide synthase (eNOS) uncoupling and that in vivo chronic inhibition of arginase restores nitroso-redox balance, improves endothelial function, and increases vascular compliance in old rats. Arginase activity in old rats was significantly increased compared with that shown in young rats. Old rats had significantly lower nitric oxide (NO) and higher superoxide (O2−) production than young. Acute inhibition of both NOS, with NG-nitro-l-arginine methyl ester, and arginase, with 2( S)-amino- 6-boronohexanoic acid (ABH), significantly reduced O2− production in old rats but not in young. In addition, the ratio of eNOS dimer to monomer in old rats was significantly decreased compared with that shown in young rats. These results suggest that eNOS was uncoupled in old rats. Although the expression of arginase 1 and eNOS was similar in young and old rats, inducible NOS (iNOS) was significantly upregulated. Furthermore, S-nitrosylation of arginase 1 was significantly elevated in old rats. These findings support our previously published finding that iNOS nitrosylates and activates arginase 1 (Santhanam et al., Circ Res 101: 692–702, 2007). Chronic arginase inhibition in old rats preserved eNOS dimer-to-monomer ratio and significantly reduced O2− production and enhanced endothelial-dependent vasorelaxation to ACh. In addition, ABH significantly reduced vascular stiffness in old rats. These data indicate that iNOS-dependent S-nitrosylation of arginase 1 and the increase in arginase activity lead to eNOS uncoupling, contributing to the nitroso-redox imbalance, endothelial dysfunction, and vascular stiffness observed in vascular aging. We suggest that arginase is a viable target for therapy in age-dependent vascular stiffness.

In a syngeneic tumour-host system the disappearance of circulating tumour cells was studied in mice treated with heparin, a coumarin anticoagulant and an antifibrinolytic. The cells were injected intravenously in two doses and blood was drawn from the right heart at intervals after the cell transfusion and inoculated intraperitoneally into new recipients. Only minor changes in the disappearance rate of tumour cells from the blood were obtained by anticoagulant and antifibrinolytic treatment. Thus thrombus formation, if present, cannot play but a minor role for the retention of tumour cells in organs in this system. The coumarin anticoagulant delayed somewhat the removal from the blood of 2 × 106 injected tumour cells. This was in contrast to heparin which, if anything, seemed to hasten the removal of the cells from the blood.

– The purpose of this study was to investigate the long-term efficacy and tolerability of gamma-vinyl GABA (GVG) in the treatment of epilepsy. 36 patients with severe therapy resistant epilepsies participated, the majority exhibiting complex partial seizures. The mean follow-up period was 9.3 months. GVG was administered as add-on therapy, to keep serum levels of concomitant treatment constant. The mean dose of GVG was 2.6 g's per day. Fifty-six per cent of the patients, including three patients with juvenile myoclonic epilepsy, experienced more than a 50% reduction in seizure frequency. No signs of tolerance development to the antiepileptic effect of GVG was demonstrated. Two patients were withdrawn from GVG treatment due to increased seizure frequency, and two due to side effects in the form of vomiting and nausea. Incidentally, the side effects observed were harmless and transient. Fifty per cent of the patients experienced no side effects at all. GVG seems to be a valuable antiepileptic compound. The results of this long-term study confirm observations from several short controlled trials.

Highly purified elastin from porcine aorta was submitted to elastase digestion. The enzymic products were subjected to gel filtration on Sephadex G 25. The excluded fraction was then submitted to thermolysin digestion and gel filtration. The excluded fraction was submitted to partial acid hydrolysis and gel filtration. Several sub-fractions were obtained. The F3 subfraction containing cross-linking agents (desmosines and lysinonorleucine) was finally subjected to ion exchange chromatography. A highly enriched peptide fraction containing lysinomorleucine was obtained and then purified by preparative electrophoresis. The ratio of enrichment passed from 2 residues of lysinonorleucine (expressed as lysine) from starting material (elastin) to 178 out of 1,000 residues in the final step. In this peptide fraction, if we express in molar ratio and consider the amount of lysinonorleucine is one residue, the following amino-acid composition is Pro:3, Gly:1, Ala:2, LNL:1, Lys:2. No traces of desmosines are detected. The role of lysinonorleucine in bridging functions in elastin is discussed.

Rationale: In a guinea pig model of allergic asthma, using perfused tracheal preparations ex vivo, we demonstrated that L-arginine limitation due to increased arginase activity underlies a deficiency of bronchodilating nitric oxide (NO) and airway hyperresponsiveness (AHR) after the allergen-induced early and late asthmatic reaction.Objectives: Using the same animal model, we investigated the acute effects of the specific arginase inhibitor 2(S)-amino-6-boronohexanoic acid (ABH) and of L-arginine on AHR after the early and late reaction in vivo. In addition, we investigated the protection of allergen-induced asthmatic reactions, AHR, and airway inflammation by pretreatment with the drug.Methods: Airway responsiveness to inhaled histamine was measured in permanently instrumented, freely moving guinea pigs sensitized to ovalbumin at 24 hours before allergen challenge and after the allergen-induced early and late asthmatic reactions by assessing histamine PC(100) (provocative concentration causing a 100% increase of pleural pressure) values.Measurements and Main Results: Inhaled ABH acutely reversed AHR to histamine after the early reaction from 4.77 +/- 0.56-fold to 2.04 +/- 0.34 fold (P Conclusions: Inhalation of ABH or L-arginine acutely reverses allergen-induced AHR after the early and late asthmatic reaction, presumably by attenuating arginase-induced substrate deficiency to NO synthase in the airways. Moreover, ABH considerably reduces the airway sensitivity to inhaled allergen and protects against allergen-induced bronchial obstructive reactions, AHR, and airway inflammation. This is the first in vivo study indicating that arginase inhibitors may have therapeutic potential in allergic asthma.

Increases (> 2.5-fold) in GABA levels in rat brain lead to a large decrease in the level of the 67-kDa form of glutamate decarboxylase (GAD67) through a mechanism involving either a change in GAD67 protein stability or a change in GAD67 mRNA translation. In the present study, brain levels of GABA were manipulated by treating rats with various doses of gamma-vinyl-gamma-aminobutyric acid (GVG), and the dependence of total GAD activity and levels of GAD67 and GAD65 protein on the levels of GABA was analyzed. Initial studies showed that both GABA and GAD67 protein levels reached new steady-state levels after two to four daily injections; GABA increased 1.5- (30 mg of GVG/kg) and fourfold (150 mg of GVG/kg), and GAD67 protein content decreased by 30 and 70%. To assess the sensitivity of GAD67 to GABA, rats were injected with eight different doses of GVG (15-150 mg/kg) for 5 days. With increasing doses of GVG, we observed a gradual increase in both whole-tissue and synaptosomal GABA levels and a gradual decrease in GAD67 protein and GAD activity. The levels of GAD65 remained constant at all GVG doses. GAD67 was remarkably sensitive to GABA. The synaptosomal GAD67 level decreased approximately 12% and the whole-neuron GAD67 level decreased approximately 3% for each 1% increase in nerve terminal GABA content when it was close to its physiological level. Our results clearly demonstrate that GAD67 is tightly controlled by intraneuronal GABA, and we suggest that this regulatory mechanism has important implications for the physiological regulation of GABAergic function in the mammalian brain.

Antifibrinolytic agents are commonly used during cardiac surgery to minimize bleeding and to reduce exposure to blood products. We sought to determine whether aprotinin was superior to either tranexamic acid or aminocaproic acid in decreasing massive postoperative bleeding and other clinically important consequences.In this multicenter, blinded trial, we randomly assigned 2331 high-risk cardiac surgical patients to one of three groups: 781 received aprotinin, 770 received tranexamic acid, and 780 received aminocaproic acid. The primary outcome was massive postoperative bleeding. Secondary outcomes included death from any cause at 30 days.The trial was terminated early because of a higher rate of death in patients receiving aprotinin. A total of 74 patients (9.5%) in the aprotinin group had massive bleeding, as compared with 93 (12.1%) in the tranexamic acid group and 94 (12.1%) in the aminocaproic acid group (relative risk in the aprotinin group for both comparisons, 0.79; 95% confidence interval [CI], 0.59 to 1.05). At 30 days, the rate of death from any cause was 6.0% in the aprotinin group, as compared with 3.9% in the tranexamic acid group (relative risk, 1.55; 95% CI, 0.99 to 2.42) and 4.0% in the aminocaproic acid group (relative risk, 1.52; 95% CI, 0.98 to 2.36). The relative risk of death in the aprotinin group, as compared with that in both groups receiving lysine analogues, was 1.53 (95% CI, 1.06 to 2.22).Despite the possibility of a modest reduction in the risk of massive bleeding, the strong and consistent negative mortality trend associated with aprotinin, as compared with the lysine analogues, precludes its use in high-risk cardiac surgery. (Current Controlled Trials number, ISRCTN15166455 [controlled-trials.com].).