Cutaneous dendritic cells in allergic inflammation.

The skin represents a physical barrier, which is capable of protecting the body from damaging invaders. Moreover, the skin operates as an active immunological organ, harbouring a complex network of dendritic cells (DCs), which serve as a bridge between innate and adaptive immunity. Equipped with specific pattern recognition receptors (PRRs), DCs are able to capture, process and present antigens to naïve T cells in the skin draining lymph nodes, thereby inducing adaptive antigen-specific immunity. However, the outcome of the immune response is shaped by numerous factors including the DC subtype, maturation state of DCs, composition of PRRs expressed by DCs, type of pathogen as well as factors in the microenvironment. Thus, cutaneous DC subtypes are known to contribute to both, peripheral tolerance and the generation of allergic skin inflammation. Identifying the underlying mechanisms is a challenging task in understanding DC biology. Based on their functional diversity, cutaneous DCs might represent promising therapeutic targets, with the potential of down-modulating pro-inflammatory immune responses and inducing tolerogenic pathways, thereby ensuring the maintenance of tissue homeostasis and restoring the balance of dysregulated immune reactions in the context of allergic skin diseases. In this review, we summarize the versatile character of DC subtypes in human skin and highlight their phenotypic characteristics and role in allergic skin inflammation. In addition, we discuss current therapeutic approaches for the management of inflammatory skin diseases such as atopic dermatitis with the main focus on strategies targeting DCs. We point towards potential challenges, benefits, risks and limitations for the treatment of patients.

Human skin and oral mucosal dendritic cells as 'good guys' and 'bad guys' in allergic immune responses.

Recent progress achieved by an impressive number of studies focusing upon the ontogenesis and immunobiology of epidermal Langerhans cells (LCs) and other cutaneous dendritic cell (DC) populations as well as DCs at oral mucosal tissue has profoundly revised our understanding of the role of DCs in different tissues and microenvironments. By sensing their environment for microbial signals or allergens and bridging innate and adaptive immunity in a sophisticated manner, subtypes of DCs play a critical role in the maintenance of the immunological homeostasis in the periphery. Thereby, DCs, located directly at the interface to the environment, fulfil opposing tasks as they are key players in both the control and the generation of allergic inflammation. Furthermore, it is under ongoing debate whether DCs attenuate or aggravate allergic inflammation. As a consequence, accumulated knowledge gained in this field within the last few years has provided an excellent basis for innovative therapeutic opportunities which tend to target specifically the multi-faceted properties of DCs at distinct anatomical sites.

Automated selection of the optimal cardiac phase for single-beat coronary CT angiography reconstruction.

PURPOSE: Reconstructing a low-motion cardiac phase is expected to improve coronary artery visualization in coronary computed tomography angiography (CCTA) exams. This study developed an automated algorithm for selecting the optimal cardiac phase for CCTA reconstruction. The algorithm uses prospectively gated, single-beat, multiphase data made possible by wide cone-beam imaging. The proposed algorithm differs from previous approaches because the optimal phase is identified based on vessel image quality (IQ) directly, compared to previous approaches that included motion estimation and interphase processing. Because there is no processing of interphase information, the algorithm can be applied to any sampling of image phases, making it suited for prospectively gated studies where only a subset of phases are available. METHODS: An automated algorithm was developed to select the optimal phase based on quantitative IQ metrics. For each reconstructed slice at each reconstructed phase, an image quality metric was calculated based on measures of circularity and edge strength of through-plane vessels. The image quality metric was aggregated across slices, while a metric of vessel-location consistency was used to ignore slices that did not contain through-plane vessels. The algorithm performance was evaluated using two observer studies. Fourteen single-beat cardiac CT exams (Revolution CT, GE Healthcare, Chalfont St. Giles, UK) reconstructed at 2% intervals were evaluated for best systolic (1), diastolic (6), or systolic and diastolic phases (7) by three readers and the algorithm. Pairwise inter-reader and reader-algorithm agreement was evaluated using the mean absolute difference (MAD) and concordance correlation coefficient (CCC) between the reader and algorithm-selected phases. A reader-consensus best phase was determined and compared to the algorithm selected phase. In cases where the algorithm and consensus best phases differed by more than 2%, IQ was scored by three readers using a five point Likert scale. RESULTS: There was no statistically significant difference between inter-reader and reader-algorithm agreement for either MAD or CCC metrics (p > 0.1). The algorithm phase was within 2% of the consensus phase in 15/21 of cases. The average absolute difference between consensus and algorithm best phases was 2.29% ± 2.47%, with a maximum difference of 8%. Average image quality scores for the algorithm chosen best phase were 4.01 ± 0.65 overall, 3.33 ± 1.27 for right coronary artery (RCA), 4.50 ± 0.35 for left anterior descending (LAD) artery, and 4.50 ± 0.35 for left circumflex artery (LCX). Average image quality scores for the consensus best phase were 4.11 ± 0.54 overall, 3.44 ± 1.03 for RCA, 4.39 ± 0.39 for LAD, and 4.50 ± 0.18 for LCX. There was no statistically significant difference (p > 0.1) between the image quality scores of the algorithm phase and the consensus phase. CONCLUSIONS: The proposed algorithm was statistically equivalent to a reader in selecting an optimal cardiac phase for CCTA exams. When reader and algorithm phases differed by >2%, image quality as rated by blinded readers was statistically equivalent. By detecting the optimal phase for CCTA reconstruction, the proposed algorithm is expected to improve coronary artery visualization in CCTA exams.

Design, synthesis, and biological evaluation of new growth inhibitors of Trypanosoma cruzi (epimastigotes).

As a continuation of our project aimed at the search for new chemotherapeutic agents against Chagas' disease, several drugs structurally related to the insect growth regulator Fenoxycarb and the naturally occurring juvenile hormone of insects were designed, synthesized, and evaluated as antiproliferative agents against the parasite responsible of this disease. Isoprenoid derivatives (compounds 33, 34, 36, and 37) were potent growth inhibitors of Trypanosoma cruzi epimastigotes. In addition, taking into account the high activity observed for compound 30 and the inhibitory action of related compounds, the allyl ether moiety bonded at the polar extreme of these inhibitors proved to be a promising group for the design of new drugs.

Structure-activity relationship of new growth inhibitors of Trypanosoma cruzi.

Several drugs bearing the 4-phenoxyphenoxy skeleton and other closely related structures were designed, synthesized, and evaluated as antiproliferative agents against Trypanosoma cruzi, the etiologic agent of Chagas' disease. The new class of drugs was envisioned by modifying the nonpolar 4-phenoxyphenoxy moiety replacing selected aromatic protons by different groups via electrophilic aromatic substitution reactions as well as introducing a sulfur atom at the polar extreme. Of the designed compounds, sulfur-containing derivatives were shown to be potent antireplicative agents against T. cruzi. Among these drugs, 4-phenoxyphenoxyethyl thiocyanate (compound 56) proved to be an extremely active growth inhibitor of the epimastigote forms of T. cruzi and displayed an IC50 of 2.2 microM. Under the same assay conditions, this drug was much more active than Nifurtimox, one of the drugs currently in clinical use to control this disease. This thiocyanate derivative was also a very active inhibitor against the intracellular form of the parasite at the nanomolar level. Other sulfur derivatives prepared also exhibited very potent antiproliferative action against T. cruzi. The presence of a sulfur atom at the polar extreme for this family of compounds seems to be very important for biological action because this atom was always associated with high inhibition values. 4-Phenoxyphenoxyethyl thiocyanate presents very good prospective not only as a lead drug but also as a potential chemotherapeutic agent.

Effects of juvenile hormone on mammalian steroidogenesis.

The effects of juvenile hormone-III (JH-III) and the JH analogue 2-(4-phenoxyphenoxy)-ethoxyte-trahydropiran on testicular steroidogenesis were studied. By using cultured MA-10 Leydig tumor cells as a model, these compounds were found to be potent inhibitors of LH/hCG steroidogenic action in a dose-dependent manner. Scatchard plot analysis of the binding data indicated that the JH analogue did not significantly alter the affinity nor the number of hCG binding sites, as well as GTP binding to plasma membranes. JH analogue inhibited the stimulatory action of both cholera toxin and forskolin on cAMP production and the concomitant steroidogenic response. JH analogue inhibited (Bu)2cAMP-stimulated progesterone synthesis, indicating that a process downstream to the adenylyl cyclase in the steroidogenic pathway is also affected.

Sulphur-containing derivatives structurally related to fenoxycarb are potent growth inhibitors against the intracellular form of Trypanosoma cruzi.

Sulphur-containing derivatives structurally related to the insect growth regulator fenoxycarb were shown to be extremely active antiproliferative agents against the amastigote form of Trypanosoma cruzi in in vitro assays. All of these drugs had previously been proved to be remarkably potent growth inhibitors against the epimastigote form of the parasite.

A pregnane glycoside from the roots of Mandevilla pentlandiana.

A pregnane triglycoside containing a new genin 3 beta,14 beta-dihydroxy-21-methoxy-5 beta-pregnan-20-one has been isolated from the dried roots of Mandevilla pentlandiana. Chemical and spectroscopic evidence for the glycoside are consistent with the structure 3 beta, 14 beta-dihydroxy-21-methoxy-5 beta-pregnan-20-one-3-O-beta-D- diginopyranosyl-(1----4)-O-beta-D-cymaropyranosyl-(1----4)-O -beta-D-cymaropyranoside. It is noteworthy that 3 beta,14 beta,21-trihydroxy 5 beta-pregnan-20-one, biosynthetically related to the genin of this glycoside, has been proved to be a precursor of cardenolides, also produced by this plant.

Synthesis and bioactivity of natural and C-3 fluorinated biosynthetic precursors of 28-homobrassinolide.

In this paper we describe the synthesis of two new fluorinated brassinosteroids: (22R,23R)-22,23-dihydroxy-3alpha-fluorostigmastan-6-one and (22R,23R)-22,23-dihydroxy-3beta-fluorostigmastan-6-one. Their bioactivities were evaluated in the rice lamina inclination test and compared with that of 28-homocastasterone, 28-homotyphasterol and 28-homoteasterone, possible biosynthetic precursors of 28-homobrassinolide. Results confirmed expected similarities between the biosynthesis of 24-ethylbrassinosteroids (named as the 28-homo series) and that described for 24-methylbrassinosteroids, and also indicated that these precursors might exhibit per se activities.

Carbon-13 nuclear magnetic resonance spectral data of steroidal vicinal ketols and related compounds.

Carbon-13 nuclear magnetic resonance spectra for 31 3 beta-hydroxy and acetoxy androstane derivatives bearing vicinal oxygenated functions at ring D with and without oxygenated functions at C-6 are reported. Relative substituent effects are discussed.

Acyl migration in the Reformatsky reaction of 21-acyloxy-5-pregnen-20-one derivatives with ethyl bromoacetate.

Reformatsky reaction of 3beta, 21-diacyloxy-and 3beta-methoxy-21-acyloxy-5-pregnen-20-one with ethyl bromoacetate yields, through an intramolecular 1,2-acyl migration, 38, 20 XI-acyloxy-14alpha-card-5-enolides were converted into the respective 20 XI-hydroxy-14alpha-card-5-enolides and the 14 alpha-carda-5,20(22)-dienolides. Experimental support to the proposed intramolecular 1,2-acyl migration is provided by the use of labelled compounds.

Biosynthesis of bufadienolides in toads. VI. Experiments with [1,2-3H]cholesterol, [21-14C]coprostanol, and 5 beta-[21-14 C]pregnanolone in the toad Bufo arenarum.

[1,2-3H]Cholesterol, 5 beta-[21-14C]cholestan-3 beta-ol (coprostanol), and 3 beta-hydroxy-5 beta-[21-14C]pregnan-20-one were injected into intact Bufo arenarum toads. Arenobufagin, the main bufadienolide present in the venom of the mentioned toad, was isolated and purified by means of chromatographic procedures. The first two compounds, having an intact cholesterol side chain, were incorporated, at comparable levels, into the bufadienolide while the labeled pregnane derivative yielded non-radioactive arenobufagin. The above results support the hypothesis that cholesterol and those steroids having an intact cholesterol-type side chain are able to penetrate to the site of bufadienolide biosynthesis and are converted into bufadienolides by a still-unknown mechanism. On the other hand, those steroid derivatives bearing a degraded side chain, e.g., 20-keto-pregnanes, are not converted into bufadienolides because they are not incorporated into the bufadienolide-producing cells.

A new approach to the synthesis of 3 beta, 23-diacetoxy-24-nor-5-cholene from 3 beta, 21-diacetoxy-5-pregnen-20-one.

Synthesis of 3 beta, 23-diacetoxy-24-nor-5-cholene in six steps from 3 beta, 21-diacetoxy-5-pregnen-20-one has been achieved by a new approach. The method would allow to label the side chain of a bile acid at carbon-atom 21.

Cleavage reaction of 5 beta, 6 beta steroidal epoxides carrying different groups at C-3.

The particular behavior of 5 beta, 6 beta steroidal epoxides carrying different groups at C-3 was studied. These epoxides may exhibit different cleavage behavior according to the nature of the solvent, the acid-base state of the medium, and the leaving abilities of the C-3 substituent. Results and alternative mechanisms are presented.

Synthesis and bioactivity evaluation of brassinosteroid analogs.

Four new analogs of 28-homocastasterone have been synthesized and completely characterized for the first time from stigmasterol. (22R, 23R,24S)-3beta-acetoxy-22,23-dihydroxy-5alpha-stigmastan+ ++-6-one (17), (22R,23R,24S)-3beta-bromo-22,23-dihydroxy-5alpha-stigmast an-6-one (18), (22R,23R,24S)-3beta-acetoxy-5,22, 23-trihydroxy-5alpha-stigmastan-6-one (20), and (22R,23R, 24S)-3beta-bromo-5,22,23-trihydroxy-5alpha-stigmastan-6-one (21), were obtained through a synthetic route based on regioselective Delta(5) epoxidation. Compounds 17 and 18, bearing a 5alphaH moiety, were prepared through a reductive opening of the 5beta,6beta epoxy precursor, and compounds 20 and 21, analogs with a 5alphaOH moiety were obtained by hydrolytic opening of a mixture of 5alpha,6alpha and 5beta,6beta epoxy precursors. Known compounds 19 and 22 were also obtained following the described synthetic routes, respectively. The new compounds were tested with the traditional auxin-like bioassay for brassinosteroids with 19 and 22 as standards. All compounds were comparatively evaluated for their inhibitory effect on the replication of DNA (HSV-1) virus.

Biosynthesis of bufadienolides in toads. V. The origin of the cholesterol used by toad parotid glands for biosynthesis of bufadienolides.

Sodium (1-14C)acetate and (5-3H) mevalonate were incubated with Bufo arenarum toad parotid gland and liver tissues. Both labelled compounds were incorporated into cholesterol produced by liver while the incubations with parotid gland produced no labelled cholesterol. Low and high density lipoproteins isolated from toad plasma were iodinated and used for binding studies. Membrane preparations of parotid gland showed high affinity binding sites for 125I-LDL and 125I-HDL. In addition, while colchicine inhibits the in vitro uptake of (3H)cholesteryl linoleate-LDL into parotid gland tissue an opposite effect was seen with (3H)cholesteryl linoleate-HDL. The above mentioned results would support the hypothesis that the cholesterol used by the parotid gland for the biosynthesis of bufadienolides would be produced in the liver, transported by the circulating lipoproteins and incorporated by the glands by a receptor-mediated mechanism.

The sterols of Cucurbita moschata ("calabacita") seed oil.

From the sterol fraction of seed oil from commercial Cucurbita moschata Dutch ("calabacita") delta 5 and delta 7 sterols having saturated and unsaturated side chain were isolated by chromatographic procedures and characterized by spectroscopic (1H and 13C-nuclear magnetic resonance, mass spectrometry) methods. The main components were identified as 24S-ethyl 5 alpha-cholesta-7,22E-dien-3 beta-ol (alpha-spinasterol); 24S-ethyl 5 alpha-cholesta-7,22E,25-trien-3 beta-ol (25-dehydrochondrillasterol); 24S-ethyl 5 alpha-cholesta-7,25-dien-3 beta-ol; 24R-ethyl-cholesta-7-en-3 beta-ol (delta 7-stigmastenol) and 24-ethyl-cholesta-7, 24(28)-dien-3 beta-ol (delta 7,24(28)-stigmastadienol).

Effects of juvenile hormone analogues (JHA) on the development of Trypanosoma cruzi.

Juvenile hormone analogues were tested for their lytic activity on Trypanosoma cruzi Chagas, 1909 blood tripomastigotes cultivated in vitro. The results indicated that the carbamate 4 and the phenoxyphenol derivative 1 are considered good candidates for blood sterilization. The compounds were also assayed for inhibition of development of parasites in infected mice showing a moderated degree of activity.