Dual inhibition of alpha/beta-hydrolase domain 6 and fatty acid amide hydrolase increases endocannabinoid levels in neurons

Agonists at cannabinoid receptors, such as the phytocannabinoid Δ(9)-tetrahydrocannabinol, exert a remarkable array of therapeutic effects but are also associated with undesirable psychoactive side effects. Conversely, targeting enzymes that hydrolyze endocannabinoids (eCBs) allows for more precise fine-tuning of cannabinoid receptor signaling, thus providing therapeutic relief with reduced side effects. Here, we report the development and characterization of an inhibitor of eCB hydrolysis, UCM710, which augments both N-arachidonoylethanolamine and 2-arachidonoylglycerol levels in neurons. This compound displays a unique pharmacological profile in that it inhibits fatty acid amide hydrolase and α/β-hydrolase domain 6 but not monoacylglycerol lipase. Thus, UCM710 represents a novel tool to delineate the therapeutic potential of compounds that manipulate a subset of enzymes that control eCB signaling.     

Influence of the ABCG2 gout risk 141?K allele on urate metabolism during a fructose challenge

Introduction

Both genetic variation in ATP-binding cassette sub-family G member 2 (ABCG2) and intake of fructose-containing beverages are major risk factors for hyperuricemia and gout. This study aimed to test the hypothesis that the ABCG2 gout risk allele 141 K promotes the hyperuricaemic response to fructose loading.

Methods

Healthy volunteers (n = 74) provided serum and urine samples immediately before and 30, 60, 120 and 180 minutes after ingesting a 64 g fructose solution. Data were analyzed based on the presence or absence of the ABCG2 141 K gout risk allele.

Results

The 141 K risk allele was present in 23 participants (31%). Overall, serum urate (SU) concentrations during the fructose load were similar in those with and without the 141 K allele (P_SNP = 0.15). However, the 141 K allele was associated with a smaller increase in SU following fructose intake (P_SNP <0.0001). Those with the 141 K allele also had a smaller increase in serum glucose following the fructose load (P_SNP = 0.002). Higher fractional excretion of uric acid (FEUA) at baseline and throughout the fructose load was observed in those with the 141 K risk allele (P_SNP <0.0001). However, the change in FEUA in response to fructose was not different in those with and without the 141 K risk allele (P_SNP = 0.39). The 141 K allele effects on serum urate and glucose were more pronounced in Polynesian participants and in those with a body mass index ≥25 kg/m².

Conclusions

In contrast to the predicted responses for a hyperuricemia/gout risk allele, the 141 K allele is associated with smaller increases in SU and higher FEUA following a fructose load. The results suggest that ABCG2 interacts with extra-renal metabolic pathways in a complex manner to regulate SU and gout risk.

Clinical Trials Registration

The study was registered by the Australian Clinical Trials Registry (ACTRN12610001036000).     

An <Emphasis Type="Italic">Escherichia coli aer</Emphasis> mutant exhibits a reduced ability to colonize the streptomycin-treated mouse large intestine

The oxygen sensor Aer of Escherichia coli affects the expression level of genes that are involved in sugar acid degradation. Phenotypes of Aer mediated gene regulation, namely growth on sugar acids was tested ‘in vitro’ with Phenotype MicroArrays and colonization of the mouse large intestine was tested ‘in vivo’. The aer mutant did not grow on the sugar acids d-gluconate, d-glucuronate, d-galacturonate, as well as the sugar alcohol d-mannitol. Since sugar acids are the predominant carbon source for E. coli in the intestinal mucosa, the ability of the aer mutant to colonize the streptomycin-treated mouse large intestine was tested. The mutant exhibited a decreased ability to colonize the intestine when compared to wild-type cells. This effect was more pronounced under competitive conditions. The colonization phenotype of the aer mutant was complemented with either of two plasmids. One of them expressed the Aer protein, whereas the other one expressed the sugar acid degradation enzymes that are encoded by edd and eda. The data support the interpretation that decreased expression of edd and eda along with the decreased ability to grow on sugar acids may contribute to the reduced capacity of the aer mutant to colonize the mouse intestine. While Aer seems to be important during the initiation phase of colonization, FlhD/FlhC appears to be of disadvantage during maintenance phase. FlhD/FlhC is the master regulator of all flagellar genes and required for Aer expression. Mutants in flhD exhibited an initial competitive disadvantage during the first 3 days of colonization, but recovered lateron.     

Pneumococcal Forssman antigen: enrichment in mesosomal membranes and specific binding to the autolytic enzyme of Streptococcus pneumoniae.

The choline-containing pneumococcal membrane teichoic acid (Forssman antigen) can be isolated with the membrane fractions of the bacteria. The small vesicle (mesosomal) fraction generated during the formation of protoplasts seems to be highly enriched in this material. Forssman antigen was identified in cell fractions on the basis of (i) radioactive choline label, (ii) autolysin-inhibitory activity, and (iii) the sedimentation profile in sucrose density gradients with and without detergent. A membrane teichoic acid could also be isolated from pneumococci grown in medium in which choline was replaced by ethanolamine as the nutritionally required amino alcohol. This material contained radioactive ethanolamine label and behaved similarly to the choline-containing membrane teichoic acid during centrifugation in detergent-containing and detergent-free density gradients. On the other hand, the material had only low autolysin-inhibitory activity. Binding of pure pneumococcal autolysin to micelles of purified Forssman antigen could be demonstrated by mixing these components in vitro and analyzing them by sucrose density gradients and by agarose chromatography. No binding could be observed between the pneumococcal enzyme and the micellar forms of either cardiolipin or polyglycerophosphate-type lipoteichoic acid isolated from Streptococcus lactis.     

THE FINE STRUCTURE AND MODE OF ATTACHMENT OF THE SHEATHED FLAGELLUM OF VIBRIO METCHNIKOVII

The sheathed flagellum of Vibrio metchnikovii was chosen for a study of the attachment of the flagellum to the bacterial cell. Normal and autolysed organisms and isolated flagella were studied by electron microscopy using the techniques of thin sectioning and negative staining. The sheath of the flagellum has the same layered structure as the cell wall of the bacterium, and in favourable thin sections it appears that the sheath is a continuation of the cell wall. After autolysis the sheath is usually absent and the core of the flagellum has a diameter of 120 A. Electron micrographs of autolysed bacteria negatively stained with potassium phosphotungstate show that the core ends in a basal disc just inside the plasma membrane. The basal disc is about 350 A in diameter and is thus considerably smaller than the "basal granules" described previously by other workers.     

A Commentary on the Need for 3D-Biologically Relevant In Vitro Environments to Investigate Astrocytes and Their Role in Central Nervous System Inflammation

Astrocytes execute essential functions in the healthy CNS, whilst also being implicated as a limitation to functional regeneration and repair after injury. They respond to injury to minimize damage to healthy tissue whilst also attempting to seal the broken blood-brain-barrier, however, they impede recovery if they are persistent and form a permanent scar in the injured brain. As such, it is of great importance to understand the mechanism underlying the astrocytic response to injury, and this understanding is currently limited by the in vitro environments available to scientists. Biomaterials such as nanofibres and hydrogels offer great potential for the development of superior, 3D cell culture environments in which to study astrocyte behavior and phenotype. The implementation of such in vitro environments with a particularly interdisciplinary approach can improve the field’s understanding of astrocytes, their role in central nervous system inflammation, and elucidate potential strategies to achieve functional regeneration.     

Histological and immunohistochemical effects of Curcuma longa on activation of rat hepatic stellate cells after cadmium induced hepatotoxicity

The liver is a target for toxic chemicals such as cadmium (Cd). When the liver is damaged, hepatic stellate cells (HSC) are activated and transformed into myofibroblast-like cells, which are responsible for liver fibrosis. Curcuma longa has been reported to exert a hepato-protective effect under various pathological conditions. We investigated the effects of C. longa administration on HSC activation in response to Cd induced hepatotoxicity. Forty adult male albino rats were divided into: group 1 (control), group 2 (Cd treated), group 3 (C. longa treated) and group 4 (Cd and C. longa treated). After 6 weeks, liver specimens were prepared for light and electron microscopy examination of histological changes and immunohistochemical localization of alpha smooth muscle actin (αSMA) as a specific marker for activated HSC. Activated HSC with a positive αSMA immune reaction were not detected in groups 1 and 3. Large numbers of activated HSC with αSMA immune reactions were observed in group 2 in addition to Cd induced hepatotoxic changes including excess collagen deposition in thickened portal triads, interlobular septa with hepatic lobulation, inflammatory cell infiltration, a significant increase in Kupffer cells and degenerated hepatocytes. In group 4, we observed a significant decrease in HSC that expressed αSMA with amelioration of the hepatotoxic changes. C. longa administration decreased HSC activation and ameliorated hepatotoxic changes caused by Cd in adult rats.