In silico, NMR and pharmacological evaluation of an hydroxyoxindole cholinesterase inhibitor.

From a screening study of various potential inhibitors for cholinesterases (ChEs), compound (rac)-1 (4-((3-hydroxy-2-oxo-3-phenylindolin-1-yl) methyl) piperidin-1-ium chloride) showed an IC50 of 18 µM for butyrylcholinesterase (BuChE). Herein we present a toxicological and pharmacological evaluation of (rac)-1 to determine its potential for use as an alternative ChE inhibitor for the treatment of Alzheimer's disease. The strategy adopted included in vivo and ex vivo studies with mouse models, Molecular Modelling and Saturation Transfer Difference (STD) NMR studies. Preliminary molecular docking studies were conducted with both (R) and (S)-1 with acetylcholinesterase (AChE) and BuChE, prior to advancing to the mouse model, and indeed favorable interactions were observed, with (R)-1 showing the best binding with AChE and (S)-1 with BuChE. STD-NMR studies were used to successfully validate these results. Toxicological studies were also conducted using the Artemia salina model, with donepezil as reference. It was found that in the in vivo mouse studies that (rac)-1 presented a slightly better inhibition of AChE (0.096 µmol.min-1.mg-1) than donepezil (0.112 µmol.min-1.mg-1) and the same level of inhibition for BuChE as donepezil (0.014 µmol.min-1.mg-1).

An Integrated Soft Computing Approach to Hughes Syndrome Risk Assessment.

The AntiPhospholipid Syndrome (APS) is an acquired autoimmune disorder induced by high levels of antiphospholipid antibodies that cause arterial and veins thrombosis, as well as pregnancy-related complications and morbidity, as clinical manifestations. This autoimmune hypercoagulable state, usually known as Hughes syndrome, has severe consequences for the patients, being one of the main causes of thrombotic disorders and death. Therefore, it is required to be preventive; being aware of how probable is to have that kind of syndrome. Despite the updated of antiphospholipid syndrome classification, the diagnosis remains difficult to establish. Additional research on clinically relevant antibodies and standardization of their quantification are required in order to improve the antiphospholipid syndrome risk assessment. Thus, this work will focus on the development of a diagnosis decision support system in terms of a formal agenda built on a Logic Programming approach to knowledge representation and reasoning, complemented with a computational framework based on Artificial Neural Networks. The proposed model allows for improving the diagnosis, classifying properly the patients that really presented this pathology (sensitivity higher than 85%), as well as classifying the absence of APS (specificity close to 95%).

Molecular approach to characterize ectomycorrhizae fungi from Mediterranean pine stands in Portugal

Stone pine (Pinus pinea L.), like other conifers, forms ectomycorrhizas (ECM), which have beneficial impact on plant growth in natural environments and forest ecosystems. An in vitro co-culture of stone pine microshoots with pure mycelia of isolated ECM sporocarps was used to overcome the root growth cessation not only in vitro but also to improve root development during acclimation phase. Pisolithus arhizus (Scop.) Rauschert and Lactarius deliciosus (L. ex Fr.) S.F. Gray fungi, were collected, pure cultured and used in in vitro co-culture with stone pine microshoots. Samples of P. arhizus and L. deliciosus for the in vitro co-cultures were collected from the pine stands southwest Portugal. The in situ characterization was based on their morphotypes. To confirm the identity of the collected material, ITS amplification was applied using the pure cultures derived from the sporocarps. Additionally, a molecular profile using PCR based genomic fingerprinting comparison was executed with other genera of Basidiomycetes and Ascomycetes. Our results showed the effectiveness of the techniques used to amplify DNA polymorphic sequences, which enhances the ­characte­rization of the genetic profile of ECM fungi and also provides an option to verify the fungus identity at any stage of plant mycorrhization.

Molecular approach to characterize ectomycorrhizae fungi from Mediterranean pine stands in Portugal

Stone pine (Pinus pinea L.), like other conifers, forms ectomycorrhizas (ECM), which have beneficial impact on plant growth in natural environments and forest ecosystems. An in vitro co-culture of stone pine microshoots with pure mycelia of isolated ECM sporocarps was used to overcome the root growth cessation not only in vitro but also to improve root development during acclimation phase. Pisolithus arhizus (Scop.) Rauschert and Lactarius deliciosus (L. ex Fr.) S.F. Gray fungi, were collected, pure cultured and used in in vitro co-culture with stone pine microshoots. Samples of P. arhizus and L. deliciosus for the in vitro co-cultures were collected from the pine stands southwest Portugal. The in situ characterization was based on their morphotypes. To confirm the identity of the collected material, ITS amplification was applied using the pure cultures derived from the sporocarps. Additionally, a molecular profile using PCR based genomic fingerprinting comparison was executed with other genera of Basidiomycetes and Ascomycetes. Our results showed the effectiveness of the techniques used to amplify DNA polymorphic sequences, which enhances the ­characte­rization of the genetic profile of ECM fungi and also provides an option to verify the fungus identity at any stage of plant mycorrhization.