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The nematode Trichinella pseudospiralis is a cosmopolitan parasite capable of infecting both birds and mammals including humans. T. pseudospiralis has a limited zoonotic importance in comparison to that of the other Trichinella species. However, it has been recognized as the etiological agent of two outbreaks of trichinellosis due to the consumption of wild boar meat. The role played by birds in the epidemiology of T. pseudospiralis is still unclear and needs to be deepened. The aim of our work was to show the results of an extensive wild bird surveillance carried out in the Emilia-Romagna (E-R) region, northern Italy, over the last 16 years. As part of the regional wildlife surveillance program, 14,933 raptors and carrion-eating birds’ carcasses were necropsied from 2006 to 2021 and tested for the presence of Trichinella spp. larvae with only one positive result, an adult female of western marsh harrier. The larvae load (LPG) was evaluated on breast (100 LPG), wings (3.6 LPG), shoulder (2 LPG), head (4.5 LPG), thighs (8 LPG), lower legs (2 LPG) and tongue (0.77 LPG). The results of the present study confirm that also in northern Italy T. pseudospiralis has a low prevalence comparing to that of other Trichinella species. However, this study demonstrates that T. pseudospiralis can reach a high parasitic load in infested birds. The large distribution range, probably facilitated by its ability to infest birds, suggests the need for a continuous monitoring program.

Epidemiologic monitoring of wild animals is always an important step in defining potential zoonoses that can threaten humans. Particular emphasis should be given to those zoonotic agents permanently cycling within wild animal populations and represent a permanent reservoir for other wild or domesticated animals that can be direct sources of disease for humans. In Croatia, there are two European jackal populations: the Dalmatian population (DP) that has been inhabiting the islands and coastal area along the Adriatic Sea since medieval times, and the South East European population (SEEP) that is found in continental Croatia. Research on Trichinella infections in jackal populations in Croatia was conducted from 2008 to 2022. During this 15-year period, we tested 186 jackal samples and confirmed infection in 47 individuals (25.3 %). The dominant species was T. spiralis, identified in 28 samples (60 %), T. britovi was found in 13 samples (28 %), while for six samples (12 %) the PCR test was unsuccessful. In both populations, the Trichinella species of the domestic cycle (T. spiralis) was found, though in varying ratios: in DP the ratio of identified species was 10:6 in favour of T. britovi, as opposed to 22:3 in favour of T. spiralis in SEEP. The frequency of infection with parasites from the genus Trichinella was significantly different in DP (22.9 %) than in SEEP (26.7 %) (p<0.001), while the larval count in analysed tissue did not differ by type of Trichinella species (p=0.1028). Infected jackals were found in nine of ten tested counties. The results were analysed statistically and the origin of tested and positive samples shown on a map of Croatia. Based on these findings, both jackal populations can be considered to represent an exceptionally important indicators of parasites from the genus Trichinella in Croatia, both for the sylvatic and domestic cycles. There is an evident need for epidemiological monitoring for members of both populations., Competing Interests: Declaration of Competing Interest The authors declared that there is no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

In some Southeastern European (SEE) countries, like Serbia, Romania, and Bulgaria, trichinellosis is one of the most important foodborne zoonotic diseases. In those countries, EU regulation and local authorities require the staff of laboratories performing official controls on meat to be properly trained and to check their competence by participating regularly in proficiency testing (PT). PTs are organized by National Reference Laboratories for Trichinella of each country and involve all official laboratories testing meat. In Romania and Bulgaria, the organization of PT for the detection of Trichinella larvae in meat by Magnetic Stirrer Method (MSM) started in 2012. In Croatia and Serbia PT was first organized in 2015 and 2017, respectively. This study presents data on the performance obtained by official laboratories of SEE countries that organize PT at national level and compares the performance obtained by laboratories belonging to different countries. Results suggest that the constant participation in PT leads to an increase in the performance of participating laboratories by positively affecting the staff accuracy in sample testing by MSM. Since the percentage of recovered larvae was in some cases suboptimal (<80%) and occasionally very poor (<40%), there is room for improvement. The regular participation in PT by laboratories involved in official controls on meat intended for human consumption is fundamental to guarantee consumer safety., Competing Interests: Declaration of Competing Interest All authors declare no conflict of interest in this study., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The 1p/19q codeletion is the result of a translocation between chromosome 1 (Chr1p) and chromosome 19 (Chr19q) with the loss of derivative (1;19)(p10;q10) chromosome. The 1p/19q codeletion has predictive and prognostic significance, and it is essential for the classification of gliomas. In routine practice, the fluorescence in situ hybridization (FISH) diagnosis of 1p/19q codeletion is sometimes unexpected. This study aimed to develop a next-generation sequencing panel for the concurrent definition of the 1p/19q codeletion and IDH1/IDH2 mutation status to resolve these equivocal cases. A total of 65 glioma samples were investigated using a 1p/19q-single-nucleotide polymorphism (SNP)-IDH panel. The panel consists of 192 amplicons, including SNPs mapping to Chr1 and Chr19 and amplicons for IDH1/IDH2 analysis. The 1p/19q SNP-IDH panel consistently identified IDH1/IDH2 mutations. In 49 of 60 cases (81.7%), it provided the same 1p/19q results obtained by FISH. In the remaining 11 cases, the 1p/19q SNP-IDH panel uncovered partial chromosome imbalances as a result of interstitial amplification or deletion of the regions where the FISH probes map, leading to a mistaken overdiagnosis of 1p/19q codeletion by FISH. The 1p/19q SNP-IDH next-generation sequencing panel allows reliable analysis of the 1p/19q codeletion and IDH1/IDH2 mutation at the same time. The panel not only allows resolution of difficult cases but also represents a cost-effective alternative to standard molecular diagnostics procedures., (Copyright © 2021 Association for Molecular Pathology and American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Breast cancer (BC) is the most common cancer in women and, among different BC subtypes, triple negative (TN) and human epidermal growth factor receptor 2 (HER2)-positive BCs have the worst prognosis. In this study, we investigated the anticancer activity of the root ethanolic and hexane extracts from Lithospermum erythrorhizon, a traditional Chinese herbal medicine known also as tzu ts'ao or tzu-ken, against in vitro and in vivo models of TNBC and HER2-positive BC. Treatment with L. erythrorhizon root extracts resulted in a dose-dependent inhibition of BC cell viability and in a significant reduction of the growth of TNBC cells transplanted in syngeneic mice. Acetylshikonin, a naphthoquinone, was identified as the main bioactive component in extracts and was responsible for the observed antitumor activity, being able to decrease BC cell viability and to interfere with autochthonous mammary carcinogenesis in Δ16HER2 transgenic mice. Acetylshikonin anticancer effect depends on its ability to act as a potent inhibitor of dihydrofolate reductase (DHFR), to down-regulate key mediators governing cancer growth and progression, such as HER2, Src and STAT3, and to induce apoptosis by caspase-3 activation. The accumulation of acetylshikonin in blood samples as well as in brain, kidney, liver and tumor tissues was also investigated by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) highlighting that L. erythrorhizon treatment is effective in delivering the active compound into the target tissues. These results provide evidence that L. erythrorhizon extract and in particular its main component acetylshikonin are effective against aggressive BC subtypes and reveal new acetylshikonin mechanisms of action., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Adenosine receptor antagonists are generally based on heterocyclic core structures presenting substituents of various volumes and chemical-physical profiles. Adenine and purine-based adenosine receptor antagonists have been reported in literature. In this work we combined various substituents in the 2, 6, and 8-positions of 9-ethylpurine to depict a structure-affinity relationship analysis at the human adenosine receptors. Compounds were rationally designed trough molecular modeling analysis and then synthesized and evaluated at radioligand binding studies at human adenosine receptors. The new compounds showed affinity for the human adenosine receptors, with some derivatives endowed with low nanomolar K i data, in particular at the A 2A AR subtype. The purine core proves to be a versatile core structure for the development of novel adenosine receptor antagonists with nanomolar affinity for these membrane proteins., (Copyright © 2019 Elsevier Inc. All rights reserved.)

In this work, an enlarged series of 1,2,4-triazolo[4,3-a]pyrazin-3-ones was designed to target the human (h) A 2A adenosine receptor (AR) or both hA 1 and hA 2A ARs. The novel 8-amino-1,2,4-triazolopyrazin-3-one derivatives 1-25 featured a phenyl or a benzyl pendant at position 2 while different aryl/heteroaryl substituents were placed at position 6. Two compounds (8 and 10) endowed with high affinity (K i  = 7.2 and 10.6 nM) and a complete selectivity for the hA 2A AR were identified. Moreover, several derivatives possessed nanomolar affinity for both hA 1 and hA 2A ARs (both K i  < 20 nM) and different degrees of selectivity versus the hA 3 AR. Two selected compounds (10 and 25) demonstrated ability in preventing β-amyloid peptide (25-35)-induced neurotoxicity in SH-SY5Y cells. Results of docking studies at the hA 2A and hA 1 AR crystal structures helped us to rationalize the observed affinity data and to highlight that the steric hindrance of the substituents at the 2- and 6-position of the bicyclic core affects the binding mode in the receptor cavity., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Objective: To analyze relationships between CD34 expression and several demographic, clinical, and pathologic features in patients with histopathologic evidence of low-grade epilepsy-associated tumors who underwent epilepsy surgery., Methods: A retrospective study enrolling 187 patients with low-grade epilepsy-associated tumors who underwent surgery between January 2009 and June 2015 at 8 Italian epilepsy surgery centers was conducted. All cases were histologically diagnosed according to the World Health Organization classification of central nervous system tumors. Univariate and multivariate analyses were performed to identify variables associated with CD34 expression., Results: Of 187 patients, 95 (50.8%) were CD34 positive. Tumor type and duration of epilepsy were independently associated with CD34 expression on multivariate analysis. Ganglioglioma and pleomorphic xanthoastrocytoma were the histologic types with the strongest association with CD34 positivity with an odds ratio of 9.2 and 10.4, respectively, compared with dysembryoplastic neuroepithelial tumors. Patients with a duration of epilepsy >10 years had a significantly greater likelihood to show CD34 expression, with an odds ratio of 2.8 compared with patients with a duration of epilepsy <2 years. On univariate analysis, CD34 expression appeared to be significantly related to older age at surgery, higher antiepileptic drug intake, and female sex., Conclusions: CD34 expression holds promise as a useful biomolecular marker for patients with low-grade epilepsy-associated tumors with evidence of a link with clinicopathologic features. This study confirmed the association between CD34 expression and tumor type and demonstrated a significantly higher probability of CD34 expression in patients with longer duration of epilepsy, independent of histology., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Aims: A 3 adenosine receptor (A 3 AR) signalling activation seems to mediate anticancer effect, and it has been targeted for drug development. The identification of potent and selective A 3 AR agonists could be crucial for cancer drug development., Materials and Methods: In the present study was determined the in vitro activity of known 1-3 and newly 4-6 synthesized compounds with high A 3 AR affinity and selectivity (K i in the low nanomolar range) in binding studies. Effect of known and novel A 3 AR agonists on human prostate cancer (PC3), hepatocellular carcinoma (Hep G2), and epithelial colorectal carcinoma (Caco-2) cells were analysed by cytotoxicity assay, dose and time dependent inhibitor assay, migration, apoptosis, autophagy and reactive oxygen species (ROS) assays., Key Findings: Results show that the anticancer effect is not due to A 3 AR activation alone. In fact, the more active and selective agonist versus A 3 AR, compound 1, results inactive on cancer cells such as compounds 2-4. Moreover, results show that the novel compound 5, at micromolar concentration range (IC 50  = 28.0 μM), inhibits the growth of PC3, Hep G2, and Caco-2 cells and their migration in time- and dose- dependent manner. The mechanism involved in cell death is attributable to apoptosis. At the same time compound 5 promotes autophagy, which induce apoptosis producing autophagic cell death. Further investigation revealed that compound 5 elevates the level of ROS in all cancer cells tested, suggesting the involvement of ROS in cell death., Significance: These results show that the new compound 5 exerts inhibitory effect on cancer cells through differential effect and may serve as a potential anticancer agent., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: Diffuse leptomeningeal glioneuronal tumor (DLGNT) was recently added to the World Health Organization classification of central nervous system tumors. DLGNT is a rare entity that occurs more commonly in pediatric patients, but occasional cases have been reported in adults. This tumor has been recognized as a distinct pathologic entity; however, its biologic behavior remains unclear. It is considered an indolent neoplasm, although considerable morbidity has been reported. For this reason, further characterization and collection of evidence are crucial., Methods: In this article, we reported a case of a 36-year-old woman with a DLGNT characterized by rapid, aggressive behavior. We also performed a review of the literature for reported cases of low-grade and high-grade forms involving adults and children., Results: DLGNTs should no longer be considered only as low-grade tumors affecting pediatric patients. The spectrum of presentations also includes aggressive tumors affecting adults., Conclusions: Further clinical and pathologic data supported by cytogenetic and molecular investigations are mandatory to better characterize DLGNTs., (Copyright © 2018 Elsevier Inc. All rights reserved.)

In Europe, Trichinella spiralis, the most dangerous species for humans of the genus Trichinella, has a patchy distribution with important foci in Eastern countries and Spain. This zoonotic pathogen was apparently not circulating among wild and domestic animals of Italy. In 2016, muscle larvae belonging to this nematode species were detected in a red fox (Vulpes vulpes) shot in the Piacenza province (Northern Italy). This parasite may have been introduced into northern Italy from eastern Europe by hunters, by a hunting dog, or by immigrants, who illegally carried infected meat in their personal baggage. In the same year, T. spiralis infected sausages illegally introduced by personal baggage into Italy from Romania, were inadequately disposed of in the garbage of a central Italian town. Even though these two episodes may not be connected in time and space, they represent an increased risk of infection for domestic and wild swine, which are highly susceptible to this pathogen. In these animals, T. spiralis shows a higher larval burden and a longer survival time than other Trichinella species. Since most of the Italian pig production plants are in northern Italy, the circulation of T. spiralis should be strictly monitored in wildlife living in these areas., (Copyright © 2017. Published by Elsevier B.V.)

According to the Commission Implementing Regulation (EU) 2015/1375 (replacing the Commission Regulation (EC) No 2075/2005), all animals, which are potential carriers of Trichinella spp. larvae, should be tested at the slaughterhouse or game-handling establishments according to one of the approved tests. One of the core duties of the European Union Reference Laboratory for Parasites is to organize proficiency testing (PT), as stated in the Commission Regulation (EC) No. 882/2004 of the European Parliament and of the Council. The aim of this work was to evaluate the results of PTs of the digestion method carried out by the National Reference Laboratories for Parasites (NRLPs) over a nine year period (2007-2015). Participating laboratories received a panel of samples consisting in 35g or 100g of minced pork or horse meat spiked with Trichinella spiralis live larvae. The number of spiked samples varied from 2 to 9 over the years. A negative control was also included in the panel, except during the 2015 PT, when only positive samples were used. The percentage of NRLPs, which passed the PT, increased from 83.3% in 2007 to 100% in 2014. Considering the number of recovered larvae, the heterogeneity in participant's results reduced overtime. The values of the overall mean difference between spiked and recovered larvae decreased during the study period, witnessing a general improvement of NRLPs performance and confirming the effectiveness of PT for a good performance of this test., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Trichinella spp. are zoonotic parasites transmitted to humans by the consumption of raw or insufficiently cooked meat of different animal species. Carnivore mammals are important reservoir hosts of these nematodes. The aims of this work were to establish the prevalence of Trichinella spp. and infection intensity in sylvatic carnivore mammals of Latvia, to identify the etiological agents at the species level and their circulation in the Latvian regions. From 2010 to 2014, muscle samples were collected from 1286 hunted animals (2 European badgers, 137 pine martens, 24 stone martens, 4 golden jackals, 394 raccoon dogs, 668 red foxes, 23 grey wolves, and 34 Eurasian lynxes). Trichinella spp. larvae were isolated by muscle digestion. Overall, 633 animals (49.2%; 95% CI 46.5%-52.0%) belonging to all the eight investigated species, tested positive for Trichinella spp. larvae. Trichinella britovi was the most common species (94.0%; 95% CI 91.7%-95.7%). Trichinella nativa was detected in 30 animals as single (6, 1.1%; 95% CI 0.4%-2.3%) or mixed infection (24, 4.4%; 95% CI 2.9%-6.4%) with T. britovi. Trichinella spiralis was detected in only three animals as mixed infection with T. britovi. The high prevalence of Trichinella spp. infection in sylvatic carnivore mammals suggests that they are good indicators for the risk assessment of Trichinella spp. in Latvia., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: The collection of the greatest possible amount of pathologic tissue is of paramount importance in neurosurgery to achieve the most accurate histopathologic diagnosis, to perform all of the necessary biomolecular tests on the pathologic specimen, and to collect biological material for basic or translational science studies. This problem is particularly relevant in pituitary surgery because of the possible small size and soft consistency of tumors, which make them suitable for removal through suction, reducing the amount of available pathologic tissue. To solve this issue, we adopted a filter connected to the suction tube, which allows the surgeon to collect all of the tissue aspirated during surgery., Methods: Our experience of 1734 endoscopic endonasal procedures, performed adopting this device since 1998 to December 2015, has been revised to assess its advantages and limitations., Results: This system is easy-to-use, does not impair the surgical maneuvers, and does not add any relevant cost to the surgery. The tissue collected through the filter proved useful for diagnostic histologic and biomolecular analyses and for research purposes, without any relevant artifacts as a result of this method of collection., Conclusions: The use of a filter has allowed us to obtain the greatest amount possible of pathologic tissue at each surgery. This surgical material has revealed to be helpful both for diagnostic and basic science purposes. The use of the filter has proven to be of particular importance for microadenomas, soft tumors, and supradiaphragmatic or skull base lesions with heterogeneous features, improving the accuracy of histopathologic diagnosis., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Hippocampal sclerosis (HS) is the most common cause of drug-resistant medial temporal lobe epilepsy (MTLE). Structural abnormalities such as HS, granule cell pathology (GCP), and focal cortical dysplasia (FCD) have been classified histopathologically, possibly allowing a more accurate assessment of prognostic seizure and neuropsychologic outcomes. We correlated seizure outcome with comprehensive temporal lobe pathologic findings, identified according to the most recent classification systems of HS, GCP, and FCD., Methods: All the 83 patients who underwent anterior temporal lobectomy (ATL) for drug-resistant MTLE and with a proven diagnosis of HS between April 2001 and May 2014 were collected. Patients were divided in 2 main groups: 1) isolated HS with/without GCP (HS +/- GCP); and 2) HS associated with FCD with/without GCP (HS+FCD +/- GCP). Patients were followed up at least 1 year, and seizure outcome was reported in accordance with Engel classification., Results: Group I: HS +/- GCP: Statistical analysis confirmed a better outcome in HS + GCP patients than in HS-no GCP (P < 0.05). Moreover, a better outcome for the patients affected by GCP type I was observed (P < 0.05). Group II: HS+FCD +/- GCP: Patients with HS variant type I presented a better seizure outcome than the patients with HS type II (Engel class IA HS type I vs. type II: 69% vs. 40%)., Conclusions: A pathology-based approach to epilepsy surgery might improve the interpretation of the results, could predict which cases will enjoy a better seizure outcome, and could help to the comprehension of the causes of failures., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The zoonotic nematode Trichinella britovi has been documented in animals and/or humans of the Mediterranean islands of Corsica and Sardinia since 2004. From 2005 to 2007 in the Sardinia island, several surveys had shown that T. britovi was circulating among backyard and free-ranging pigs reared in the Orgosolo municipality but all attempts had failed to detect this parasite in wild susceptible animals. The aim of the present work was to investigate the circulation of T. britovi in pigs and wildlife of the Orgosolo municipality, and of surrounding municipalities and provinces in the 2010-2014 slaughtering/hunting seasons. The results show that the T. britovi circulation was still restricted to the Orgosolo municipality with a prevalence of 2.6% in free-ranging pigs and 0.2% in backyard pigs but, for the first time, this parasite was detected also in 0.4% of wild boar, and 27.6% of red foxes. No infection was detected in backyard pigs, wild boar, and red foxes of the other municipalities and provinces. Since 1978, African swine fever is endemic in Sardinia and foci of this virus are still active in the investigated areas favoring cannibalism and, consequently, the T. britovi transmission, due to the high mortality rate caused by this virus. This is the first documented report on the transmission of T. britovi between the domestic and the sylvatic cycle. The health authority of the island must provide a service to dispose animal carcasses and offal, stamping out illegal free-ranging pigs, and train hunters and pig owners to manage waste and by-products according to the EU regulations., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Active surveillance of red foxes for Trichinella has been undertaken in mainland Britain since 1999. Post-mortems are carried out, followed by a magnetic stirrer method for sample digestion based on European Commission (EC) Regulation 216/2014 (which amends 2075/2005). Initially samples are tested in batches of 20 foxes and in December 2013, for the first time under the surveillance programme, a batch tested positive for Trichinella at the Animal and Plant Health Agency, York. Further individual tests identified one infected fox, from the Bristol area. The larvae were identified as Trichinella pseudospiralis. This is the first report of T. pseudospiralis in Great Britain and suggests the possibility of a cycle of infection existing in wildlife., (Crown Copyright © 2015. Published by Elsevier B.V. All rights reserved.)

Trichinella spiralis and Trichinella britovi are the two most common species of the genus Trichinella persisting in the European wildlife. To investigate the spatial distribution of these Trichinella spp. and the factors influencing their circulation in Hungary, 3304 red foxes (Vulpes vulpes) and 0.29 million wild boars (Sus scrofa) were tested for Trichinella sp. infection in Hungary from 2006 to 2013. Trichinella spp. larvae from 68 (2.06%) foxes and 44 (0.015%) wild boars were identified by a multiplex PCR as T. britovi or T. spiralis. The locality of origin of foxes and wild boars were recorded in a geographic information system database. There was no correlation between environmental parameters in the home range of foxes and wild boars and the T. spiralis larval counts, but there was a positive correlation between the boundary zone of Hungary and T. spiralis infection (P<0.0001; odds ratio: 24.1). These results indicate that the distribution of T. spiralis in the Hungarian wildlife is determined by the transborder transmission of the parasite from the surrounding endemic countries. Multiple regression analysis was performed with environmental parameter values and T. britovi larval counts. Based on the statistical analysis, non-agricultural areas (forests, scrubs, herbaceous vegetation and pastures) and the mean annual temperature (P<0.0001; odds ratios: 9.53 and 0.61) were the major determinants of the spatial distribution of T. britovi in Hungary. The positive relationship with non-agricultural areas can be explained by the generalist feeding behaviour including scavenging of foxes in these areas. The negative relationship with the mean annual temperature can be attributed to the slower decomposition of wildlife carcasses favouring a longer survival of T. britovi larvae in the host carrion and to the increase of scavenging of foxes., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Trichinella infections in humans and pigs have been documented in Greece since 1945 and a high prevalence of infection in pigs occurred in the 1950s. Up to 1984 only sporadic infections in humans were documented, and this zoonosis was not considered as a public health problem until 2009 when a human outbreak caused by the consumption of pork from an organic pig farm occurred. In the present study, we describe the re-emergence of Trichinella spp. infections in free-ranging pigs from organic farms of 3 counties (Dramas, Evros and Kavala) in Northern-Eastern Greece during the period 2009-2012. Totally 37 out of 12,717 (0.29%) free-ranging pigs which were tested during the period in question, were positive for Trichinella spp. larvae. The etiological agent was identified as Trichinella britovi. The average larval burden was 13.7 in the masseter, 6.2 in the foreleg muscles and 7.5 in the diaphragm. The 37 positive animals originated from seven free range pig farms. The practice of organic pig production systems in Greece has grown in popularity over the last years due to the increasing interest of consumers for products considered as traditional. However, this type of pig production increases the risk for Trichinella spp. infections, since animals can acquire the infection by feeding on carcasses or the offal of hunted or dead wild animals. The awareness and education of hunters and farmers is extremely important to reduce the transmission among free ranging pigs and the risk for humans., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Over the last decades the distribution area of the golden jackal (Canis aureus) has increased significantly in Europe, particularly in the Balkan Peninsula and in Central Europe. Vagrant individuals were described in many European countries. Herein, we report Echinococcus multilocularis (total worm count: 412) and Trichinella spiralis (101 larvae/g for muscles of the lower forelimb) infections in two golden jackals shot in Hungary. It is a new host record of E. multilocularis and T. spiralis in Europe and Hungary, respectively. As jackals migrate for long distances through natural ecological corridors (e.g., river valleys), they may play a significant role in the long distance spread of zoonotic parasites into non-endemic areas of Europe. Therefore, monitoring zoonotic parasites in this host species can be recommended in the European Union., (Copyright © 2013 Elsevier B.V. All rights reserved.)

An antigen detection kit (Trichin-L), based on latex agglutination and developed by the Bio-Rad company was validated at five European laboratories. The validation parameters included specificity, sensitivity, robustness and reproducibility. Specificity was evaluated by testing parasite antigens from five non-Trichinella parasites in addition to the Trichinella genus. To evaluate sensitivity, 10 pork samples spiked with 1, 3, 6 or 15 Trichinella larvae were tested in each laboratory. To evaluate the robustness of the test, the solubilized antigens were maintained at room temperature and tested at different times. Reproducibility was assessed in each laboratory using 40, 100g minced pork samples, each spiked with Trichinella spiralis. The use of larval homogenates obtained from the Trichin-L kit as a template for parasite identification at the species level by a multiplex PCR, was also evaluated. The results showed a high specificity and sensitivity where solubilized antigens maintained their stability and reactivity for up to three days. Reproducibility was high, as similar results were obtained in the five laboratories. The larval homogenates obtained using the Trichin-L kit were successfully used in multiplex PCRs to identify Trichinella species., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Although larvae of the genus Trichinella are the most common parasite species detected in vertebrate muscles using artificial digestion, nematode larvae belonging to other genera are sometimes detected and incorrectly identified as Trichinella. However, it is often very difficult to identify these larvae at the species, genus or family level using microscopy because of the absence of specific morphological characters or cuticle damage, and the only means of identification is PCR and sequencing of specific molecular markers (12S mtDNA; COI; 18S rDNA; and ITS1). From 2008 to 2011, 18 nematode isolates not belonging to the genus Trichinella were collected from different host species. Eleven of these isolates were successfully identified at the species, genus or superfamily level: larvae from two common kestrels, three hooded crows, a hen harrier and a domestic pig were identified as Toxocara cati; larvae from a badger were identified as Toxocara canis; larvae from a domestic pig were identified as a free-living nematode of the genus Panagrolaimus; larvae from a wild boar were identified as belonging to the Metastrongylus genus; and larvae from a rough-legged buzzard were identified as belonging to the superfamily Filarioidea. The recovery of nematodes belonging to genera other than Trichinella during routine meat inspection suggests that the persons performing the analyses need to be informed of the possibility of false positives and that a molecular-based identification system that allows for a rapid and reliable response must be adopted (i.e., a DNA barcoding-like system)., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Objective: The relationship between hippocampal histopathological abnormalities, epileptogenesis and neurogenesis remains rather unclear., Methods: Tissue samples including the subgranular zone of dentate gyrus (DG) were freshly collected for tissue culture for neurospheres generation in 16 patients who underwent surgery for drug-resistant temporal lobe epilepsy. Remaining tissues were histologically examined to assess the presence of mesial temporal sclerosis (MTS) and focal cortical dysplasia., Results: MTS was detected in 8 cases. Neurospheres were formed in 10/16 cases. Only three out of these 10 cases exhibited MTS; on the contrary 5/6 cases lacking neurosphere proliferation presented MTS. There was a significant correlation between presence of MTS and absence of proliferation (p = 0.0389). We also observed a correlation between history of febrile seizures (FS) and presence of MTS (p = 0.0004) and among the 6 cases lacking neurosphere proliferation, 4 cases (66.6%) had experienced prolonged FS. Among "proliferating" cases the percentage of granular cells pathology (GCP) was lower (20% vs 50%) compared to "non proliferating" cases., Conclusion: A decreased potential to generate neurosphere from the SGZ is related to MTS and to alterations of dentate gyrus granule cells, especially in MTS type 1b and GCP type 1. These histological findings may have different prognostic implications, regarding seizure and neuropsychological outcome, compared to patients with other epileptogenic lesions (such as FCD, glioneuronal tumours, vascular lesions)., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Dysembryoplastic neuroepithelial tumors (DNTs) are usually located within the supratentorial cortex, often in the temporal lobe and they are frequently associated with intractable complex partial seizures. DNTs in extracortical sites are rare. Thus far, 21 cases of 36 DNT-lesions occurring in these areas have been reported; only 8 out of them had epilepsy. We report a case of a 39-year-old woman who had pharmacoresistant epilepsy associated to a DNT in the caudate nucleus-periventricular area treated by lesionectomy. During a 4-year follow-up period, the patient was seizure free and the tumor did not recur. We discuss the hypothetical epileptogenic mechanism involved and we review the pertinent literature., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Trichinellosis is a foodborne disease caused by the consumption of raw meat and raw meat-derived products from swine, horse and some game animals infected with nematode worms of the genus Trichinella. Between June 2006 and February 2011, 16 million domestic pigs and 0.22 million wild boars (Sus scrofa) were tested for Trichinella sp. in Hungary. Trichinella infection was not found in any pigs slaughtered for public consumption. Nevertheless, Trichinella spiralis was detected in four backyard pigs when trace back was done following a family outbreak. Trichinella infection was demonstrated in 17 wild boars (0.0077%). Larvae from wild boars were identified as Trichinella britovi (64.7%), T. spiralis (29.4%) and Trichinella pseudospiralis (5.9%). Although the prevalence of Trichinella sp. infection in wild boars and domestic pigs is very low, the spatial analysis reveals that the level of risk differs by region in Hungary. Most of the T. britovi infected wild boars (63.6%) were shot in the north-eastern mountain area of Hungary; whereas domestic pigs and wild boars infected with T. spiralis were detected only in the southern counties bordering Croatia and Romania. In the north-western and central counties, the prevalence of Trichinella infection seems to be negligible., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Gene delivery to eukaryotic cells is the technique to study the regulation of gene expression. Human astrocytoma cell line 1321N1 could be useful to study G-protein-coupled receptors (GPCRs). Different transient transfection methods, namely calcium phosphate, Lipofectamine, FuGENE, Arrest-In, and microporation (Microporator), were investigated. Results were analyzed by fluorescence-activated cell sorting and fluorescence microscope using green fluorescent protein (GFP) as a reporter gene. To verify the transfection efficiency of these techniques, the expression of human GPR17 gene (hgpr17) was analyzed by transcription quantitative polymerase chain reaction. Microporation resulted in the best method to promote enriched hgpr17 delivery into the human astrocytoma cell line., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Trichinella pseudospiralis is a non-encapsulated species infecting both mammals and birds. In Italy, this parasite was reported only in two night-birds of prey of Central Italy. In January 2010, Trichinella larvae were detected in three wild boars (Sus scrofa) of two regions of Northern Italy by enzymatic digestion. The parasites were identified as T. pseudospiralis by multiplex-PCR. The first infected wild boar was hunted in the Emilia Romagna region and the other two infected wild boars were bred outdoors in a small family farm of the Friuli Venezia Giulia region. These new epidemiological data reinforce the role of the wild boar as the main reservoir of T. pseudospiralis in Europe., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Immunoproteasome has been associated to neurodegenerative and autoimmune diseases as a marker and regulator of inflammatory mechanisms. Its expression in the brain may occur upon neuroinflammation in different cell types and affect a variety of homeostatic and inflammatory pathways including the oxidized protein clearance and the self-antigen presentation. In the present study we investigated the immunoproteasome expression in hippocampi and cortex of patients affected by different histopathological forms of pharmaco-resistent mesial temporal lobe epilepsy. We identified a pathology-specific pattern of immunoproteasome expression, which could provide insight into the complex neuroinflammatory pathogenic components of this disease., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Nematodes of the genus Trichinella are zoonotic parasites with a cosmopolitan distribution. In Iran, these parasites have mainly been detected in carnivorous mammals, yet information on the Trichinella taxa circulating in this country date back to a time when biochemical and molecular tests were not available. We describe the first detection of Trichinella larvae in a leopard (Panthera pardus saxicolor) in Asia and its identification at the species level. The larvae recovered from the leopard muscles were identified as Trichinella britovi using multiplex PCR. The detection of Trichinella infection in a leopard confirms literature data on the high prevalence of infection in carnivorous mammals in Iran.

Trichinella infections in horses continue to represent a health problem and, despite the rarity of infection, it is necessary to continue to control properly horse meat. In 2008, a 10-year-old horse imported from Poland to Italy for consumption found to have been positive at the digestion test. Both Trichinella britovi and Trichinella spiralis larvae in a proportion of 4:1 were detected in the horse muscles. This is the first report of a mixed Trichinella species infection in a horse. The epidemiological investigation revealed that the infected horse originated from a small farm about 120km from Warsaw and the horse owner had bought the horse at a horse market. The findings suggest that the horse was fed more than once with infected meat.

Recent discovery of Trichinella zimbabwensis in crocodiles from Zimbabwe, Lake Cahora Basa, Mozambique, and from lake Abaja, Ethiopia, prompted strict control measures to curb the possible spread of the infection to humans and also to prevent its introduction to other countries, which were considered free of this pathogen. In 2006, the Chief Directorate Veterinary Services of Mpumalanga Province of South Africa launched a survey to investigate the status of wild and commercial breeding crocodiles in the province. To evaluate if T. zimbabwensis was circulating in the environments where crocodiles are living in South Africa, 9 fish, 36 reptiles (including 27 Nile crocodiles) and 4 mammals have been investigated to detect Trichinella sp. larvae in their muscles. In January 2008, a Nile crocodile from Komatipoort, sampled by means of a tail biopsy, tested positive for Trichinella larvae. In June-July 2008, Trichinella sp. larvae were also detected in four other Nile crocodiles from the Olifants River Gorge. The prevalence of Trichinella infection in the investigated wild Nile crocodiles from South Africa is 38.5%. The larvae were identified as belonging to T. zimbabwensis by multiplex-PCR. These are the first reports of T. zimbabwensis in South Africa and suggest that the distribution area of this parasite species is wider than that believed in the past.

For 60 years, the islands of the Mediterranean basin were considered to be Trichinella-free. In April 2005, an outbreak of human trichinellosis due to the consumption of infected pork involved 11 persons in the villages of Orgosolo and Lanusei (Nuoro province) on the island of Sardinia (Italy). We conducted an investigation to identify free-range and backyard pigs and other humans with Trichinella infection in the area of the 2005 outbreak. We also tested wild animals from various parts of Sardinia. In December 2005, eight persons were found to have been infected, and in May 2007 there was a single case of infection. The sources of all infections were domestic pigs. Artificial digestion of muscle samples from 681 pigs (325 free-range and 356 backyard pigs) revealed Trichinella sp. larvae in four sows (1.2%). All larvae, including those from the consumed pork products, were identified as Trichinella britovi. All infected pigs originated from the Orgosolo municipality. None of the 6188 wild boars (Sus scrofa) or 13 red foxes (Vulpes vulpes) examined were positive for Trichinella sp., suggesting that this parasite is restricted to free-range pigs. The origin of infected animals on Sardinia remains to be determined, although it could be related to the presence of T. britovi-infected animals on the island of Corsica (France).

The aim of this study was to evaluate the inflammatory response in the muscle tissue against Trichinella larvae of encapsulated (T. spiralis, T. britovi) and non-encapsulated (T. pseudospiralis) species. The inflammatory response was estimated in histological sections of muscle tissues from Trichinella-infected CD1 mice by a newly developed method. Nuclei were stained with one fluorescent probe, which binds nucleic acids with high affinity, and fluorescence was analysed by a software program. Evaluation of the relative fluorescence units was performed in both peri-capsular (close to the nurse cell-parasite complex) and extra-capsular (where the parasite was not visible) areas. The increase in the number of nuclei in the muscle tissues of Trichinella-infected mice was considered an inflammation marker, since uninfected muscles show low nucleus density. In order to evaluate differences in the nitrosylation pattern between encapsulated (T. spiralis, T. britovi) and non-encapsulated (T. pseudospiralis, Trichinella papuae, Trichinella zimbabwensis) species, L(1) larvae were tested by immunoblotting with an anti-nitrotyrosine polyclonal antibody. Inflammation induced by T. spiralis larvae in muscle tissues is statistically higher than that elicited by the other species, both in peri- and extra-capsular areas. Nitrosylation occurs at a higher level in encapsulated than in non-encapsulated species. The method developed in this work allows demonstration of differences in the host inflammatory response against encapsulated and non-encapsulated Trichinella species.

To control Trichinella spp. infection in the European Union, all slaughtered pigs should be tested by one of the approved digestion methods described in EU directive 2075/2005. The aim of the present work was to evaluate, by a ring trial, the sensitivity of the digestion method used at the National Reference Laboratories for Parasites (NRLP). These Laboratories are responsible for the quality of the detection method in their own country. Of the 27 EU countries, only three (Hungary, Luxembourg and Malta) did not participate in the ring trial. Each participating laboratory received 10 samples of 100g of minced pork containing 3-5 larvae (3 samples), 10-20 larvae (3 samples), 30-50 larvae (3 samples), and one negative control. In each positive sample, there were living Trichinella spiralis larvae without the collagen capsule, obtained by partial artificial digestion of muscle tissue from infected mice. No false positive sample was found in any laboratories, whereas nine laboratories (37.5%) failed to detect some positive samples with the percentage of false negatives ranging from 11 to 100%. The variation between expected and reported larval counts observed among the participating laboratories was statistically significant. There was a direct correlation between the consistency of the results and the use of a validated/accredited digestion method. Conversely, there was no correlation between the consistency of the results and the number of digestions performed yearly by the NRLP. These results support the importance of validating the test.

In South Africa, Trichinella sp. was first discovered in 1966 in the wildlife of the Kruger National Park (KNP). Since then, both Trichinella T8 and Trichinella nelsoni have been detected in the KNP, leading to a debate on the existence of a gene flow between the two taxa. In 2006-2008, four lions were killed in the Manyeleti Game Reserve, the Mthethomusha Nature Reserve, Numbi Gate, and Skukuza, which border the KNP. Larvae were isolated from muscles by artificial digestion. The molecular identification of single larva by multiplex PCR, followed by a specific PCR to distinguish between Trichinella T8 and Trichinella britovi, revealed Trichinella T8 in the lions from Manyeleti and Skukuza, a mixed infection with T. nelsoni and Trichinella T8 in the lion from Mthethomusha, and T. nelsoni in the lion from Numbi. No larva with a hybrid pattern between the two taxa was observed. No hybrid offspring resulted when crossing single males and females of T. nelsoni and Trichinella T8 in both directions, whereas hybrid offspring were obtained when crossing T. britovi and Trichinella T8 in both directions. This is the first report of a mixed infection with two Trichinella taxa in a host from the KNP, where both Trichinella T8 and T. nelsoni circulate among wildlife. Despite the sympatry status of these two taxa, field and laboratory data seem to exclude the possibility of gene flow, confirming their evolutive separation.

The red fox (Vulpes vulpes) is considered one of the main reservoir of Trichinella spp. in Europe. As limited information on Trichinella infection in wildlife of Hungary is available, 2116 red foxes, representing more than 3% of the estimated fox population of the country, were screened to detect Trichinella larvae by a digestion method. Trichinella larvae from the 35 positive foxes were identified by a multiplex PCR as Trichinella britovi (30 isolates, 85.7%), Trichinella spiralis (4 isolates, 11.4%), and Trichinella pseudospiralis (1 isolate, 2.9%). The true mean intensity of T. britovi, T. spiralis and T. pseudospiralis larvae in lower forelimb muscles was 23.6, 3.5 and 13.5larvae/g, respectively. T. spiralis was detected only in the southern and eastern regions. The non-encapsulated T. pseudospiralis was recorded for the first time in Hungary. Although the overall true prevalence of Trichinella infection in foxes was only 1.8% (95% confidence interval, CI=1.5-2.1%), the spatial analysis reveals different risk regions. In the north-eastern counties bordering Slovakia and Ukraine (21% of the Hungarian territory), the true prevalence of Trichinella infection is significantly higher than that observed in other regions (6.0%, CI=4.8-7.1%). In the southern counties bordering Croatia, Serbia and Romania (41% of the Hungarian territory), the true prevalence of Trichinella infection is moderate (1.4%, CI=1.0-1.8%). In the north-western and central counties (38% of Hungarian territory), the prevalence of Trichinella infection is significantly lower (0.2%, CI=0.1-0.4%) than that of the other regions. Based on the statistical analysis and the evaluation of epidemiological data, none of the counties can be considered free of Trichinella infection. In the past decade, Trichinella infection has been detected only in few backyard pigs, and only few wild boar-related autochthonous infections in humans were described. Nevertheless, these results highlight the need of the maintenance of a strict monitoring and control programmes on Trichinella infection in farmed and hunted animals of Hungary.

In 1995, a new species of Trichinella (Trichinella zimbabwensis) was discovered in farmed Nile crocodiles (Crocodylus niloticus) in Zimbabwe, where the mode of transmission was the consumption of the meat of slaughtered crocodiles, used as feed. To determine whether T. zimbabwensis affects poikilotherm vertebrates in the wild, monitor lizards (Varanus niloticus) and Nile crocodiles were collected in Zimbabwe and Mozambique. In 5 (17.6%) of the 28 monitor lizards from Zimbabwe, T. zimbabwensis larvae were identified. For the wild Nile crocodiles from Mozambique, species-level identification was not possible, yet immunohistochemical analysis revealed that 8 (20%) of the 40 animals harboured non-encapsulated Trichinella sp. larvae, which probably belonged to T. zimbabwensis. This is the first report of T. zimbabwensis in wild reptiles, and the findings are consistent with reports that vertebrates with scavenger and cannibalistic behaviour are the most important hosts of Trichinella spp. The wide distribution of monitor lizards and crocodiles in Africa and the development of national crocodile breeding programs in many African countries should be taken into consideration when evaluating the risk of transmission of this parasite to mammals, including humans.

Trichinella sp. infection has been documented in both humans and animals in most Mediterranean countries, yet in the past 60 years no infections have been reported on the Mediterranean islands. We describe the first outbreak of Trichinella sp. infection to have been reported on the island of Sardinia. The outbreak occurred in two villages in 2005 and involved 11 persons who had eaten raw sausages made from the same free-ranging sow. All 11 persons developed signs and symptoms of trichinellosis and seroconverted within 48 days of consuming the infected meat. The etiological agent was Trichinella britovi. Sardinia, like all Mediterranean islands, had been considered to be Trichinella-free. The present report, together with a recent report of T. britovi infection in animals on the nearby island of Corsica (France), raises questions as to the validity of the concept of Trichinella-free areas or regions.

The recent discovery of Trichinella zimbabwensis in farmed crocodiles (Crocodilus niloticus) of Zimbabwe and its ability to infect mammals, and the development of both T. zimbabwensis and Trichinella papuae in experimentally infected reptiles led to an investigation of Trichinella infection in saltwater crocodiles (Crocodylus porosus) and in wild pigs (Sus scrofa) of Papua New Guinea, to see if T. papuae also, is present in both cold- and warm-blooded animals. Of 222 crocodiles examined, 47 animals (21.2%), all from Kikori, Gulf Province, were positive for non-encapsulated larvae in the muscles. The greatest number of larvae was found usually in the biceps, with an average of 7 larvae/g. One isolate from a crocodile infected successfully both laboratory rats and mice. Of 81 wild pigs examined, 9 from Bensbach river area (Western Province) and 1 from Kikori area (Gulf Province) were positive for non-encapsulated larvae in the muscles. Trichinella larvae from both saltwater crocodiles and wild pigs have been identified by multiplex-PCR analysis as T. papuae. The sequence analysis of the region within the large subunit ribosomal DNA, known as the expansion segment V, has shown the presence of a molecular marker distinguishing T. papuae isolates of Bensbach river area from those of Kikori area. This marker could be useful to trace back the geographical origin of the infected animal. The epidemiological investigation carried out in the Kikori area has shown that local people catch young crocodiles in the wild and keep them in holding pens for several months, before sending them to the crocodile farm in Lae (Morobe Province). They feed the crocodiles primarily with wild pig meat bought at the local market and also with fish. These results stress the importance of using artificial digestion for routinely screening of swine and crocodiles, and of adopting measures for preventing the spread of infection, such as the proper disposal of carcasses and the adequate freezing of meat.

The synthesis and preliminary pharmacological profile of a new series of muscarinic antagonists, derived from previously studied 2,2-diphenyl-2-ethylthio-acetic acid esters, are reported. The parent molecules were decorated with linkers of different length, carrying an amino group to catch a putative anionic function outside the recognition site of the receptor. It was hoped that the interception of this function would give molecules with higher potency and selectivity. The attempt has not been successful, but a new series of compounds with a peculiar pharmacological profile has been identified.

In Sweden, the prevalence of Trichinella infection in domestic pigs has greatly decreased since the 1970s, with no reports in the past 4 years. However, infected wild animals continue to be found. The objective of the present study was to identify the species of Trichinella present in animals of Sweden, so as to contribute to the knowledge on the distribution area and hosts useful for the prevention and control of this zoonosis. In the period 1985-2003, Trichinella larvae were detected in the muscles of 81/1800 (4.5%) red foxes (Vulpes vulpes), 1/6 (16.7%) arctic fox (Alopex lagopus), 1/7 (14.3%) wolf (Canis lupus), 10/200 (5.0%) lynxes (Lynx lynx), 4/8000 (0.05%) wild boars (Sus scrofa), and 27/66 x 10(6) (0.000041%) domestic pigs. All four Trichinella species previously found in Europe were detected (Trichinella spiralis, T. nativa, T. britovi and T. pseudospiralis). The non-encapsulated species T. pseudospiralis was detected in three wild boars from Holo (Stockholm area) and in one lynx from Froso (Ostersund area), suggesting that this species is widespread in Sweden. These findings are consistent with those of a study from Finland, both for the unexpected presence of T. pseudospiralis infection and the presence of the same four Trichinella species, suggesting that this epidemiological situation is present in the entire Scandinavian region. The widespread diffusion of T. pseudospiralis in the Scandinavian region is also important in terms of it potential impact on public health, given that human infection can occur and the difficulties to detect it by the trichinelloscopic examination.

A series of new derivatives, related to diphenidol and to its 2-carbonyl analogue, were designed as antimuscarinic agents. The synthesized compounds were evaluated both as hydrochlorides and as methiodides by functional tests at guinea-pig heart (M(2)), guinea-pig ileum (M(3)) and rabbit vas deferens (putative M(4)). Two derivatives (3a and 5a) showed an M(3)-selective profile similar to that of the reference compounds, though they resulted less potent.

A survey was carried out to investigate the prevalence and worm burden of extraintestinal nematodes in 100 red foxes (Vulpes vulpes) of Hungary. The overall prevalence of nematode infections of the respiratory tract was 76%. Eucoleus aerophilus (Capillaria aerophila) was the predominant species (66%), followed by Crenosoma vulpis (24%), Eucoleus (Capillaria) böhmi (8%) and Angiostrongylus vasorum (5%). Pearsonema (Capillaria) plica was found in 52% of the urinary bladders. In 3% of the foxes, Trichinella britovi was present in muscle samples. The high prevalence of lungworms and P. plica and the fox colonisation in urban areas may enhance the prevalence of these nematode infections in domestic dogs and cats, and the flow of T. britovi from the sylvatic cycle to the domestic cycle, enhancing the risk of infections in humans.

A series of quinazoline derivatives, 2-20, structurally related to the racemic alpha(1)-adrenoceptor antagonist cyclazosin (1), were synthesized and evaluated for their functional antagonism at alpha(1)- and alpha(2)-adrenoceptors and for their binding affinity at human cloned alpha(1a)-, alpha(1b)- and alpha(1d)-adrenoceptor subtypes. They displayed, like 1, preferential antagonism and selectivity for alpha(1) versus alpha(2)-adrenoceptors. Compounds 10, 13, and 18 showed high potency at alpha(1)-adrenoceptors similar to that of 1 (pK(B) values 8.47-8.89 versus 8.67), whereas 13 and 15 were endowed with the highest alpha(1)-adrenoceptor selectivity, only 3- to 4-fold lower than that of 1. In binding experiments, all of the compounds displayed an affinity practically similar to that found for 1, with the exception of 19 and 20 that were definitely less potent. The s-triazine analogue 18 was the most potent of the series with pK(i) values of 10.15 (alpha(1a)), 10.22 (alpha(1b)) and 10.40 (alpha(1d)), resulting 77-fold more potent than 1 at alpha(1a)-adrenoceptors. In addition, the majority of compounds, like prototype 1, showed the same trend of preferential affinity for alpha(1d)- and alpha(1b)-adrenoceptors that alpha(1a)-subtype. In conclusion, we identified compounds 2-5, 10, 12 and 13, bearing either an aliphatic- or an arylalkyl- or aryloxyalkyl-acyl function, with an interesting subtype-selectivity profile, which makes them suitable candidates for their resolution as enantiomers structurally related to (+)-cyclazosin.

A series of muscarinic agonists, straight chained, branched, cyclic alkyl and aromatic derivatives of the oxime 1 (demox) was designed with the aim of investigating their activity on muscarinic receptor subtypes. Effects on M1 receptor were assessed functionally by a microphysiometer apparatus, while M2, M3, and M4 receptor potency and affinity were studied on isolated preparations of guinea pig heart, ileum, and lung, respectively. The results suggest that the substitution of a hydrogen with a long side-chain or bulky group generally induces a decrease in potency at M1 and M3 subtypes, while a general increase in this parameter is obtained at M2 subtype. Among the agonists 2-18, compound 4 behaves as a full agonist with a preference for M3 subtype. Moreover, compound 12 is inactive at M1 and M4 receptors while it displays a full agonist activity at M2 and M3 subtypes. Since demox displays a variable response on cardiac M2 receptors regulating heart force, an in-depth inquiry of the functional behaviour of this compound was carried out at M2 receptors. In presence of 10(-11) and 10(-10) M demox, the binding of [3H]-NMS was increased by approximately 30% as a consequence of an increase of the association of [3H]-NMS to membranes; this effect was not observed in presence of a higher concentration of [3H]-NMS. Higher concentrations of demox decreased the binding of [3H]-NMS to heart atrial membranes but significantly retarded the dissociation of this radioligand. Our results suggest that demox may interact with orthosteric and allosteric sites of atrial M2 muscarinic receptor.