In silico and in vitro analysis of genetic variants of the equine CYP3A94, CYP3A95 and CYP3A97 isoenzymes.

Cytochrome P450 enzymes (CYPs) of the equine CYP3A subfamily are predominantly involved in drug metabolism. In this study, genetic variants of the equine CYP3A94, CYP3A95, and CYP3A97 were identified and characterized using in silico modeling and in vitro enzyme kinetics. The genomes of 81 horses were sequenced to obtain the genetic variants. Structural CYP modifications of the most frequent variants were analyzed in silico using the 3D-structures predicted by homology modeling. Enzyme kinetic analyses were performed using testosterone as substrate. Twenty genetic variants were found including five missense variants (CYP3A94:p.Asp217Asn, CYP3A95:p.Asp214His, CYP3A95:p.Ser392Thr, CYP3A97:p.Ile119Thr, CYP3A97:p.Met500Val) with a higher percentage of minor allele frequency (MAF) (range 0.2-0.4). A splice-site variant (c.798 + 1G > A) in CYP3A94, likely to generate a truncated protein, was found in 50% of the horses. CYP3A94:p.Asp217Asn and CYP3A95:p.Asp214His were localized on the CYP F-α-helix, an important region for the substrate interactions in the human CYP3A4. Testosterone 2ß-hydroxylation was diminished in CYP3A94217Asn and CYP3A95392Thr. Ketoconazole inhibited 2ß-hydroxylation differently in the five variants with the most pronounced inhibition obtained for CYP3A95392Thr. In vitro and in silico analyses of genetic variants allow unraveling structural features in equine CYPs that correlate with changes in the CYP activity.

Testosterone metabolism of equine single CYPs of the 3A subfamily compared to the human CYP3A4.

Cytochrome P450 enzymes (CYPs) are responsible for the phase I metabolism of drugs, xenobiotics and endogenous substances. Knowledge of single CYPs and their substrates is important for drug metabolism, helps to predict adverse effects and may prevent reduced drug efficacy in polypharmacy. In this study, three equine isoenzymes of the 3A subfamily, the equine flavoprotein NADPH-P450 oxidoreductase (POR), and the cytochrome b5 (CYB5) were cloned, sequenced and heterologously expressed in a baculovirus expression system. Testosterone, the standard compound for characterization of the human CYP3A4, was used to characterize the newly expressed equine CYPs. The metabolite pattern was similar in equine and the human CYPs, but the amounts of metabolites were isoform-dependent. All equine CYPs produced 2-hydroxytestosterone (2-OH-TES), a metabolite never described in equines. The main metabolite of CYP3A4 6ß-hydroxytestosterone (6ß-OH-TES) was measured in CYPs 3A95 and 3A97 with levels close to the detection limit. Ketoconazole inhibited 2-OH-TES in the human CYP3A4 and the equine CYP3A94 and CYP3A97 completely, whereas a 70% inhibition was found in CYP3A95. Testosterone 6ß- and 2-hydroxylation was significantly different in the equine CYPs compared to CYP3A4. The expression of single equine CYPs allows characterizing drug metabolism and may allow prevention of drug-drug interactions.

Clumsiness in fine motor tasks: evidence from the quantitative drawing evaluation of children with Down Syndrome.

INTRODUCTION: Drawing tests are commonly used for the clinical evaluation of cognitive capabilities in children with learning disabilities. We analysed quantitatively the drawings of children with Down Syndrome (DS) and of healthy, mental age-matched controls to characterise the features of fine motor skills in DS during a drawing task, with particular attention to clumsiness, a well-known feature of DS gross movements. METHODS: Twenty-three children with DS and 13 controls hand-copied the figures of a circle, a cross and a square on a sheet. An optoelectronic system allowed the acquisition of the three-dimensional track of the drawing. The participants' posture and upper limb movements were analysed as well. RESULTS: Results showed that the participants with DS tended to draw faster but with less accuracy than controls. DISCUSSION: While clumsiness in gross movements manifests mainly as slow, less efficient movements, it manifests as high velocity and inaccurate movements in fine motor tasks such as drawing.

Mobile ECMO team for inter-hospital transportation of patients with ARDS: a retrospective case series.

INTRODUCTION: Transport of patients undergoing extracorporeal membrane oxygenation is currently available in 5 referral centers in our country. METHODS: Retrospective case series of patients managed by our mobile extracorporeal membrane oxygenation team and transferred to San Gerardo University Hospital from December 2004 to December 2012. RESULTS: 42 patients were transported. The mean age was 42.11 (standard deviation ±18.11) years, with a range between 2 years and 70. 14 patients were females (33%) and 28 males (67%). The average transport distance was 121.69 km (±183.08) with a range between 9 km and 1044 Km. The mission's mean time was equal to 508 minutes (±185) with range of 120-960 minutes. 29 patients (69%) were transported with extracorporeal membrane oxygenation support, while 13 patients (31%) were transported with conventional ventilation. In 28 patients (97%) a veno-venous bypass was utilized, while in one case (3%) a Veno-Arterial cannulation was performed. 32 patients survived (76%) and have been discharged alive from hospital. No major clinical or technical issues were observed during the transport. CONCLUSIONS: According to our data, we conclude that a dedicated mobile team allowed safe ground transportation of patients with severe acute lung injury to our tertiary care institution.

The relation between Parkinson's disease and ageing. Comparison of the gait patterns of young Parkinson's disease subjects with healthy elderly subjects.

BACKGROUND: The gait of healthy elderly and of subjects with Parkinson's disease (PD) displays some common features, suggesting that PD may be a model of ageing. AIM: The aim of the study was to quantify highlight the differences and similarities between the gait patterns of young PD and healthy elderly, to uncover if PD could be assumed as a model of ageing. DESIGN: An optoelectronic system was used for 3D gait analysis evaluation. POPULATION AND METHODS: We compared the gait parameters of 15 young PD (YPD) with the gait of 32 healthy elderly subjects (ES) and 21 healthy subjects age-matched with the PD subjects. RESULTS. Common features between YPD and ES were majorly found in the parameters that reflect the presence of an unstable, uncertain gait, and of corrective strategies employed to reduce instability. On the other side, typical features were present in the gait patterns of PD subjects. CONCLUSION. Our study helped identifying some typical characteristics of the onset disease, and to unravel the symptoms of ageing from those of PD by comparing young PD subjects to elderly healthy subjects. CLINICAL REHABILITATION IMPACT: This allows a deeper understanding of the mechanisms underlying the gait in ageing and PD.