Determination of absolute configuration in 4‐aryl‐3,4‐dihydro‐2(1H)‐pyrimidones by high performance liquid chromatography and CD spectroscopy

The absolute configuration of three 4‐aryl‐3,4‐dihydro‐2(1H)‐pyrimidones (Biginelli compounds, DHPMs) was established by comparison of the typical circular dichroism (CD) spectra of individual enantiomers with reference samples of known absolute configuration. The enantiomers were obtained by semipreparative separation of racemic mixtures on a Chiralcel OD‐H chiral stationary phase. The method was used to establish the enantiopreference of various lipases in biocatalytic kinetic resolution experiments employing activated DHPM esters. Chirality 11:659–662, 1999. © 1999 Wiley‐Liss, Inc.     

Alterations in Phosphorylation of Hepatocyte Ribosomal Protein S6 Control Plasmodium Liver Stage Infection

1. Download high-res image (160KB)
2. Download full-size image

     

Cenicriviroc,a Novel CCR5 (R5) and CCR2 Antagonist,Shows In Vitro Activity against R5 Tropic HIV-2 Clinical Isolates

Background

Maraviroc activity against HIV-2, a virus naturally resistant to different HIV-1 antiretroviral drugs, has been recently demonstrated. The aim of this study was to assess HIV-2 susceptibility to cenicriviroc, a novel, once-daily, dual CCR5 and CCR2 antagonist that has completed Phase 2b development in HIV-1 infection.

Methods

Cenicriviroc phenotypic activity has been tested using a PBMC phenotypic susceptibility assay against four R5-, one X4- and one dual-tropic HIV-2 clinical primary isolates. All isolates were obtained by co-cultivation of PHA-activated PBMC from distinct HIV-2-infected CCR5-antagonist-naïve patients included in the French HIV-2 cohort and were previously tested for maraviroc susceptibility using the same protocol. HIV-2 tropism was determined by phenotypic assay using Ghost(3) cell lines.

Results

Regarding the 4 R5 HIV-2 clinical isolates tested, effective concentration 50% EC₅₀ for cenicriviroc were 0.03, 0.33, 0.45 and 0.98 nM, similar to those observed with maraviroc: 1.13, 0.58, 0.48 and 0.68 nM, respectively. Maximum percentages of inhibition (MPI) of cenicriviroc were 94, 94, 93 and 98%, similar to those observed with maraviroc (93, 90, 82, 100%, respectively). The dual- and X4-tropic HIV-2 strains were resistant to cenicriviroc with EC50 >1000 nM and MPI at 33% and 4%, respectively.

Conclusions

In this first study assessing HIV-2 susceptibility to cenicriviroc, we observed an in vitro activity against HIV-2 R5-tropic strains similar to that observed with maraviroc. Thus, cenicriviroc may offer a once-daily treatment opportunity in the limited therapeutic arsenal for HIV-2. Clinical studies are warranted.     

Infrared Thermography and Ultrasonography to Indirectly Monitor the Influence of Liner Type and Overmilking on Teat Tissue Recovery

Eight Danish Holstein cows were milked with a 1-mm thick specially designed soft liner on their right rear teat and a standard liner mounted under extra high tension on their left rear teat. Four of the animals were overmilked for 5 min. Rear teats were subjected to ultrasound examination on the first day and to infrared thermography on the second day. Teats were submersed in ethanol 20 min post-milking on the second day. Ultrasonography measurements showed that teat canal length increased by 30–41% during milking. Twenty minutes after milking, teats milked with modified standard liners still had elongated teat canals while teats milked with the soft liner were normalized. Overmilking tended to increase teat wall thickness. Approximately 80% of variability in teat canal length, from before teat preparation to after milking, could be explained by changes during teat preparation. Thermography indicated a general drop in teat temperature during teat preparation. Teat temperature increased during milking and continued to increase until the ethanol challenge induced a significant drop. Temperatures approached pre-challenge rather than pre-milking temperatures within 10 minutes after challenge. Teat temperatures were dependent on type of liner. Mid-teat temperatures post-challenge relative to pre-teat preparation were dependent on overmilking. Thermography and ultrasound were considered useful methods to indirectly and non invasively evaluate teat tissue integrity.     

L-FABP is a critical host factor for successful malaria liver stage development

The malaria parasite liver stage produces tens of thousands of red cell-infectious forms within its host hepatocyte. It is thought that the vacuole-enclosed parasite completely depends on the host cell for successful development but the molecular parasite-host cell interactions underlying this remarkable growth have remained elusive. Using a yeast two-hybrid screen and a yeast overexpression system we show that UIS3, a parasite protein essential for liver stage development, interacts directly with liver-fatty acid binding protein, L-FABP. Down-regulation of L-FABP expression in hepatocytes severely impairs parasite growth and overexpression of L-FABP promotes growth. This is the first identified direct liver stage-host cell protein interaction, providing a possible explanation for the importance of UIS3 in liver infection.     

Acute toxicity effects of ibuprofen on behaviour and haematological parameters of African catfish Clarias gariepinus (Burchell, 1822)

Indiscriminate discharge of pharmaceutical waste into the aquatic ecosystem may pose serious health challenges to aquatic biota. The effect of acute exposure to ibuprofen was evaluated using changes in behaviour and haematological parameters under static bio-assay method in Clarias gariepinus. Test specimens were exposed to acute concentrations of ibuprofen (0.28, 0.33, 0.38, 0.43 and 0.48 mg l^?1) for 24, 48, 72 and 96 h durations respectively. Behavioural and phenotypic changes were observed in surviving fish. There were significant (p < 0.05) concentration and duration-dependent increases in erythrocyte (RBC), haemoglobin (Hb), pack cell volume (PCV) and leukocytes (WBC) in treated fish compared to the control. Insignificant decreases (p > 0.05) in mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH) and mean corpuscular haemoglobin concentration (MCHC) were observed in treated fish compared to the control. Ibuprofen elicited dose and duration- dependent decrease in neutrophil counts with the decreases being significant (p < 0.05) in the higher doses of 0.43 and 0.48 mg l^?1. Ibuprofen did not elicit any significant changes in monocytes, basophils and eosinophils. Changes observed in this study showed that ibuprofen negatively affected the health of the fish and we recommend that discharge of ibuprofen into the aquatic environment should be monitored and controlled.     

Malaria parasite liver stages render host hepatocytes susceptible to mitochondria-initiated apoptosis

Intracellular eukaryotic parasites and their host cells constitute complex, coevolved cellular interaction systems that frequently cause disease. Among them, Plasmodium parasites cause a significant health burden in humans, killing up to one million people annually. To succeed in the mammalian host after transmission by mosquitoes, Plasmodium parasites must complete intracellular replication within hepatocytes and then release new infectious forms into the blood. Using Plasmodium yoelii rodent malaria parasites, we show that some liver stage (LS)-infected hepatocytes undergo apoptosis without external triggers, but the majority of infected cells do not, and can also resist Fas-mediated apoptosis. In contrast, apoptosis is dramatically increased in hepatocytes infected with attenuated parasites. Furthermore, we find that blocking total or mitochondria-initiated host cell apoptosis increases LS parasite burden in mice, suggesting that an anti-apoptotic host environment fosters parasite survival. Strikingly, although LS infection confers strong resistance to extrinsic host hepatocyte apoptosis, infected hepatocytes lose their ability to resist apoptosis when anti-apoptotic mitochondrial proteins are inhibited. This is demonstrated by our finding that B-cell lymphoma 2 family inhibitors preferentially induce apoptosis in LS-infected hepatocytes and significantly reduce LS parasite burden in mice. Thus, targeting critical points of susceptibility in the LS-infected host cell might provide new avenues for malaria prophylaxis.     

Release of hepatic Plasmodium yoelii merozoites into the pulmonary microvasculature

Plasmodium undergoes one round of multiplication in the liver prior to invading erythrocytes and initiating the symptomatic blood phase of the malaria infection. Productive hepatocyte infection by sporozoites leads to the generation of thousands of merozoites capable of erythrocyte invasion. Merozoites are released from infected hepatocytes as merosomes, packets of hundreds of parasites surrounded by host cell membrane. Intravital microscopy of green fluorescent protein-expressing P. yoelii parasites showed that the majority of merosomes exit the liver intact, adapt a relatively uniform size of 12-18 microm, and contain 100-200 merozoites. Merosomes survived the subsequent passage through the right heart undamaged and accumulated in the lungs. Merosomes were absent from blood harvested from the left ventricle and from tail vein blood, indicating that the lungs effectively cleared the blood from all large parasite aggregates. Accordingly, merosomes were not detectable in major organs such as brain, kidney, and spleen. The failure of annexin V to label merosomes collected from hepatic effluent indicates that phosphatidylserine is not exposed on the surface of the merosome membrane suggesting the infected hepatocyte did not undergo apoptosis prior to merosome release. Merosomal merozoites continued to express green fluorescent protein and did not incorporate propidium iodide or YO-PRO-1 indicating parasite viability and an intact merosome membrane. Evidence of merosomal merozoite infectivity was provided by hepatic effluent containing merosomes being significantly more infective than blood with an identical low-level parasitemia. Ex vivo analysis showed that merosomes eventually disintegrate inside pulmonary capillaries, thus liberating merozoites into the bloodstream. We conclude that merosome packaging protects hepatic merozoites from phagocytic attack by sinusoidal Kupffer cells, and that release into the lung microvasculature enhances the chance of successful erythrocyte invasion. We believe this previously unknown part of the plasmodial life cycle ensures an effective transition from the liver to the blood phase of the malaria infection.