Amyloid-ß deposits in human astrocytes contain truncated and highly resistant proteoforms.

Alzheimer's disease (AD) is a neurodegenerative disorder that develops over decades. Glial cells, including astrocytes are tightly connected to the AD pathogenesis, but their impact on disease progression is still unclear. Our previous data show that astrocytes take up large amounts of aggregated amyloid-beta (Aß) but are unable to successfully degrade the material, which is instead stored intracellularly. The aim of the present study was to analyze the astrocytic Aß deposits composition in detail in order to understand their role in AD propagation. For this purpose, human induced pluripotent cell (hiPSC)-derived astrocytes were exposed to sonicated Aß42 fibrils and magnetic beads. Live cell imaging and immunocytochemistry confirmed that the ingested Aß aggregates and beads were transported to the same lysosomal compartments in the perinuclear region, which allowed us to successfully isolate the Aß deposits from the astrocytes. Using a battery of experimental techniques, including mass spectrometry, western blot, ELISA and electron microscopy we demonstrate that human astrocytes truncate and pack the Aß aggregates in a way that makes them highly resistant. Moreover, the astrocytes release specifically truncated forms of Aß via different routes and thereby expose neighboring cells to pathogenic proteins. Taken together, our study establishes a role for astrocytes in mediating Aß pathology, which could be of relevance for identifying novel treatment targets for AD.

Quantitative assessment of training effects using EksoGT® exoskeleton in Parkinson's disease patients: A randomized single blind clinical trial.

Background: Gait alterations are among the most disabling motor-symptoms associated with Parkinson's Disease (PD): reduced stride length, stride velocity and lower limb joint range of motion are hallmarks of parkinsonian gait. Research focusing on optimal functional rehabilitation methods has been directed towards powered lower-limb exoskeletons which combines the advantages delivered from the grounded robotic devices with the ability to train the patient in a real-world environment. As gait involves both central (CNS) and peripheral nervous systems (PNS), targeted rehabilitation must restore not only mechanics but also neurophysiological gait patterns. Methods: Two cohorts of subjects will be enrolled and equally distributed between one group (n = 25) who will undergo a functional kinematic therapy, and one group (n = 25) who will undergo an overground wearable-exoskeleton training. Participants are evaluated at three time points: before the therapy (T0), after the therapy (T1), 4-weeks after T1 (T2). Comprehensive gait analysis and surface electromyography will be combined into neuromusculoskeletal modelling to determine modifications at the PNS level. Functional magnetic resonance imaging coupled with electroencephalography will be used to determine modifications at the CNS level. Conclusion: The findings of the proposed trial will likely give substantial solutions for the management of gait and postural disorders in PD where valid interventions are lacking. The coupling of movement evaluation, which assesses neuromuscular and biomechanical features, with neurological data, will better define the impact of the therapy on the relationship between PD motor alterations and brain activity. This will provide an active treatment that is personalized and shared to large populations.

Assessment of eosinophilic airway inflammation as a contribution to the diagnosis of occupational asthma.

BACKGROUND: Ascertaining the presence of asthma through the assessment of nonspecific bronchial hyperresponsiveness (NSBH) is a key step in the diagnosis of occupational asthma (OA). We aimed at investigating whether indices of airway inflammation including fractional exhaled nitric oxide (FeNO) and sputum eosinophils would be useful adjuncts to the measurement of NSBH in diagnosing OA defined as a positive specific inhalation challenge (SIC). METHODS: The study included 240 consecutive subjects with a suspicion of OA who completed a SIC, of whom 133 showed a positive response. The sensitivity, specificity, and predictive values of NSBH, and FeNO, as well as sputum eosinophil counts assessed at baseline of the SIC were determined. RESULTS: A concentration of histamine inducing a 20% decline in FEV1 (PC20 ) ≤16 mg/mL showed a sensitivity of 87% and a specificity of 36%. A FeNO level ≥25 ppb and a sputum eosinophil count ≥2% provided lower sensitivity rates (47% and 39%, respectively) than the PC20 value. Eight of the 17 subjects without baseline NSBH despite a positive SIC showed a sputum eosinophil count ≥2%, a FeNO level ≥25 ppb, or both outcomes. Combining either a PC20 value ≤16 mg/mL or a FeNO ≥25 ppb increased the sensitivity to 91%. Using either a PC20 ≤16 mg/mL or a sputum eosinophil count ≥1% increased the sensitivity to 94%. CONCLUSION: Adding the assessment of FeNO level and sputum eosinophils to NSBH improves the identification of subjects who may have OA and require further objective testing before excluding the possibility of OA.

Reproductive efficiency of adult and prepubertal goats subjected to repeated follicular aspiration / Eficiência reprodutiva em cabras adultas e pré-púberes após repetidas sessões de aspiração folicular

The objective of this study was to assess the reproductive response of adult and prepubertal goats subjected to repeated laparoscopic ovum pick-up (LOPU). The study animals were divided into two groups, specifically, adult nanny goats (GA, n=10) and prepubertal nanny goats (GP, n=10), which were subjected to estrous synchronization and ovarian stimulation for LOPU. Both groups underwent six LOPU procedures at seven-day intervals and were subsequently subjected to controlled mating and pregnancy diagnosis to evaluate their future fertility. The study showed a reduction in the number of follicles visualized and in the amount and quality of the oocytes that were recovered and exposed to in vitro maturation. As indicated by the fertility test, however, no complications were found during the laparoscopic procedures that would impair the reproductive future of the animals. Therefore, a viable number of oocytes were obtained even with the decreased reproductive efficiency, proving that repeated LOPUs do not interfere with the reproductive of adult and prepubertal nanny goats. These results indicate a positive aspect of this procedure, allowing for increasing reproductive performance of this kind, when used for the production in vitro.

O objetivo deste estudo foi avaliar a resposta reprodutiva de cabras adultas e pré-púberes submetidas a repetidas aspirações foliculares por videolaparoscopia (LOPU). Os animais do estudo foram divididos em dois grupos, especificamente cabras adultas (GA, n = 10) e pré-púberes (GP, n = 10), que foram submetidas a sincronização de estro e estimulação ovariana para LOPU. Ambos os grupos foram submetidos a seis procedimentos LOPU em intervalos de sete dias e foram posteriormente submetidos à monta controlada e posterior diagnóstico de gestação para avaliar sua fertilidade. O estudo mostrou uma diminuição do número de folículos visualizados, na quantidade e na qualidade dos oócitos que foram recuperados e expostos à maturação in vitro. Tal como indicado pelo teste de fertilidade, no entanto, não foram observadas grandes complicações durante os procedimentos laparoscópicos que pudessem prejudicar o futuro reprodutivo dos animais. Portanto, foi obtido um número de oócitos viáveis, mesmo com a diminuição da eficiência reprodutiva, provando que repetidas LOPU não interferem na reprodutividade de animais adultos e pré-púberes. Estes resultados indicam um aspecto positivo desse procedimento, possibilitando um aumento da eficiência reprodutiva dessa espécie quando utilizado para a produção in vitro.

Long-term outcome of living donors older than 60 years.

Long-term outcomes of renal transplantation using kidneys from donors >60 years old are generally considered to be poor. This retrospective study included 265 living donor (LD) transplants in adult recipients with a mean follow-up of 13.1 +/- 6.1 years (range, 1.3-25.8), all of them under CNI. They were grouped according to the donor age at least (n = 49) or less (n = 216) than 60 years. Graft and patient survivals were compared using the Kaplan-Meier method and Cox multiple regression. At 1, 3, and 10 years, postoperatively patient survivals in the group of older LD recipients were 97%, 96%, and 93%, versus 98%, 97% and 92% among the younger LD recipients. At 1, 3 and 10 years, postoperatively graft survivals uncensored for death were 94%, 92%, and 81% among the older LD recipients versus 93%, 89%, 75% among the control group, respectively, despite a slightly increased creatininemia observed at 10 years among the older LD recipients. Deaths censored graft survivals were 96%, 96%, and 87% among the older versus 94%, 91% and 78% among the younger LD recipients, respectively. Therefore, significantly better noncensored death-censored graft survivals, were observed among the recipients of older LD compared with recipients of the younger donor group.

Malignancies in 2,753 kidney recipients transplanted during a 39-year experience.

An increased development of malignancies has been related to modern potent antirejection drugs. The purpose of this retrospective study was to assess the incidence and risk factors for invasive malignancies among 2753 kidney recipients (KRs), who were transplanted in two periods within our 39-year experience; before (group I) versus after (group II) the introduction of calcineurin inhibitors (CNIs). In group I, formed by 703 KRs under conventional therapy, 45 (6.4%) patients developed a malignancy, while in group II, treated with CNIs, of over 2050 KRs, 182 (8.9%) developed a malignancy different from noninvasive skin cancer. The incidence of malignancies was higher in the group of patients treated with CNIs (8.9% vs 6.4%), despite the shorter follow-up period. Moreover, the malignancy was more precocious in the CNI group, namely a mean time of onset of 75 versus 154 months in the conventionally treated group. The older mean age of recipients in group II affected by malignancies (43.6 years vs 34.6 years of the group I) played a significant (P < .001) role when associated with the more powerful immunosuppressive effect of CNIs, while recipient gender, dialysis period, donor source, and retransplants seemed to have few effects on malignancy development. Recipients over 60 under CNIs showed a 21% incidence of malignancies.

Pediatric kidney transplantation: a snapshot 10 years later.

Herein we report the outcomes of pediatric kidney recipients who underwent transplantation at least 10 years prior. A cohort of 36 patients (mean age, 26.4+/-6 years) with a mean follow-up time of 14.2+/-4 years was selected for the study. Immunosuppression consisted of cyclosporine and steroids. Actuarial patient and graft survivals 15 years after the transplantation were 97% and 86%, respectively. Only 1 patient died due to a complicated sclerosant peritonitis. Graft function was good with a mean serum creatinine of this selected cohort of 1.5+/-0.6 mg/dL. Eighteen percent were class 1, 33% class 2, and 49% chronic kidney disease. Hypertension was treated in almost 80% of the patients. The majority of patients were smaller than the average population with a final height (between 0 and -2) standard deviation score (HSDS) but only 27% had a severe growth impairment (HSDS>-2). Regarding nutritional status, fewer than 30% were overweight and only 1 patient was obese with a body mass index (BMI) >30. The majority of patients, except 2 mentally retarded individuals, are or have been attending normal school and achieved full-time employment. In conclusion, long-term survivors of a kidney transplant received during childhood reached a high degree of rehabilitation despite a long period of immunosuppression.

Value of intraoperative resistive index in kidney transplant.

The value of the resistive index (RI) obtained by echo color doppler evaluation of the transplanted kidney is still not well established. Many authors consider the RI to be nonspecific sign of rejection, acute tubular necrosis, or urinary tract obstruction, but its specificity remains low. In this paper, we report our experience with RI determinations in 34 consecutive kidney transplants at different times namely: perioperatively, at 24 hours, at 3 days, at 6 and at 9 days posttransplant. In all patients intraoperative RI was normal. RI increased significantly after transplantation in 10 patients who eventually developed a complication: delayed function, acute rejection, and spontaneous kidney ruptures. This increment from the baseline value was already significant at 24 hours after the kidney transplant, indicating a possible posttransplant complication (0.62 +/- 0.07 vs 0.76 +/- 0.04; P = .0004). We conclude that the value of RI in the early posttransplant phase should be considered an important aid for the early diagnosis of posttransplant complications.

COX-1 and COX-2 inhibition in horse blood by phenylbutazone, flunixin, carprofen and meloxicam: an in vitro analysis.

We report on the inhibitory activity of the NSAIDs meloxicam, carprofen, phenylbutazone and flunixin, on blood cyclooxygenases in the horse using in vitro enzyme-linked assays. As expected, comparison of IC50 indicated that meloxicam and carprofen are more selective inhibitors of COX-2 than phenylbutazone and flunixin; meloxicam was the most advantageous for horses of four NSAIDs examined. However at IC80, phenylbutazone (+134.4%) and flunixin (+29.7%) had greater COX-2 selectivity than at IC50, and meloxicam (-41.2%) and carprofen (-12.9%) had lower COX-2 selectivity than at IC50. We therefore propose that the selectivity of NSAIDs should be assessed at the 80% as well as 50% inhibition level.

A sheer pharmacologic approach to compare the contractile effects of PGF2alpha, DL-cloprostenol and D-cloprostenol on isolated uterine, tracheal, ileal and arterial smooth muscle preparations.

The contractile effects of PGF2alpha and its cloprostenol analogs (D-enantiomer and racemate) were examined on isolated smooth muscle preparations (uterine, tracheal, ileal and arterial) from rat, guinea-pig and horse. DL- and D-cloprostenol were potent contractors of myometrium, but had negligible secondary effects on other types of smooth muscle, except artery whose response to the D-enantiomer may give rise to some concern.

Different contractile effects of alpha1- and alpha2-adrenergic agonists on horse isolated common digital artery smooth muscle ring preparations in vitro.

Despite assays on ring preparations in vitro confirmed that the vasoconstrictor sympathetic control in the horse common digital artery mainly depends on alpha(1)-adrenoceptors stimulation, selective alpha(2)-adrenoceptor agonists were investigated under the same experimental conditions. Both detomidine (DET) and UK 14304 differed from noradrenaline (NA) and phenylephrine (PHE) in provoking contractile effects which were slowly onsetting, concentrations-unrelated and unremovable by repeated washings. While prazosin (PRA) clearly antagonized the effects of NA and PHE, neither pre- nor post-treatments of the preparations with alpha(2)-antagonists succeeded in antagonizing or removing the effects of the two alpha(2)-agonists tested, which moreover were unaffectable either by lowering the organ bath temperature or by depriving the nutritive medium of Ca(2+). To explain this unusual behavior of alpha(2)-adrenoceptors stimulation it has been hypothesized that a Ca(2+) mobilization from the endoplasmic reticulum of the smooth muscle cell occurs which is followed by a hindered reuptake of them.

Early and long-term results using older kidneys from cadaver or living donors.

The persisting shortage of organs for transplantation justifies the use of all possible donors. We compared results for 110 patients, who received solitary kidneys from cadaver donors over age 60 years (OCD) with those for 976 patients, who were transplanted with kidneys from 11-49-year-old cadaver donors, whom we defined as "ideal" age. Although the 4% incidence of primary nonfunction and the 24% rate of delayed graft function were significantly higher (p < 0.001) in the OCD group compared with the ideal group (0.8% and 8.0%, respectively), OCD kidneys can offer good results when low-risk recipients are carefully selected and the cold ischemia time is sharply reduced. Moreover, graft survival rates for 14 OCD grafts, implanted in patients under age 45 were 93% at one, 3 and 10 years compared with 79%, 74% and 42% at the same time points for 96 OCD recipients over age 45. The rate of delayed graft function was higher among 19 OCD grafts preserved for more than 20 hours, and these grafts yielded significantly lower survival rates than 91 OCD grafts preserved for less than 20 hours; with rates of 67%, 58% and 44% and 85%, 81% and 51%, respectively, after one, 3 and 10 years. Thirty-five kidneys from living donors over age 60 had comparable overall graft survival rates to living donor kidneys from donors under age 60 (92%, 92% and 92% vs. 92%, 88% and 80% at one, 3 and 10 years, respectively). An original point scoring system, based only on macroscopic evaluation of the graft, avoids the need for a biopsy and does not prolong cold ischemia time. Microvascular bench reconstructions of the renal artery, damaged by atherosclerosis, expand the possibility for safe transplantation of older kidneys without performing a double renal transplant.

Affinity of isoxsuprine for adrenoreceptors in equine digital artery and implications for vasodilatory action.

We used isolated equine digital arteries to study the vasodilatory mechanism of isoxsuprine, and fowl caecum preparations to investigate the affinity of the drug for beta-adrenoceptors. Isoxsuprine is a potent vasodilator of arterial smooth muscle that has been precontracted by an alpha-adrenoceptor agonist such as noradrenaline (log EC50 = -6.33 [-5.98; -6.68]). The present study indicates that its effect is due to alpha-adrenoceptor blockade since: (1) after a long lasting exposure to cumulative doses of isoxsuprine the vasoconstricting action of noradrenaline cannot be restored; (2) isoxsuprine does not promote relaxation on preparations precontracted by PGF2alpha; (3) isoxsuprine shifts the dose-response curve of noradrenaline to the right; and (4) its affinity (pK(B) = 6.90 [6.60; 7.20]) in this experiment is comparable to that in noradrenaline-precontracted preparations and is 14 times lower than that of the selective alpha1-adrenergic antagonist prazosin [pK(B) = 8.04 (7.40; 8.68]). The affinity of isoxsuprine for beta-adrenoceptors was 100 times lower than that of isoprenaline when tested on fowl caecum. This preparation has a large beta-adrenoceptor and negligible alpha-adrenoceptor population concerned with the control of smooth muscle motility. Our data suggest that the alpha-mediated effect of isoxsuprine on horse arterial smooth muscle is due to higher affinity of the drug for alpha- than beta-adrenoceptors rather than low concentration or functionality of beta-sites at this site. According to these data, pure beta2-agonists seem to be more profitable tools to determine vasodilation of the arterial bed in horses legs.

[Seveso: biological monitoring of dioxin pollution. An example of environmental medicine. Multicentric and interdisciplinary applied research]. / Seveso--Monitoraggio biologico di inquinamento da Diossina: Esempio di Medicina Ambientale. Ricerca Applicata Multicentrica e Interdisciplinare.

In 1976 the population of an area including Seveso (about 30,000) affected by the fallout of a toxic cloud containing 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) started a health monitoring plan which lasted until 1985. Smaller groups were monitored until 1997. The large number of people and the different toxic effects on organs have gathered different discipline experts including informatic. This work has permitted the not easy observation both of minor and bigger effects during two decades.

Protection from renal ischemia-reperfusion injury by the 2-methylaminochroman U83836E.

BACKGROUND: In a prior study the 21-aminosteroid (lazaroid) U74389F provided in vivo protection from oxidative stress when used as a preventive therapy in ischemia-reperfusion injury in the kidney. As the cell membrane is the principal site for lipoperoxidation, in the current study the very lipophilic 2-methylaminochroman U83836E, a recently developed lazaroid, was administered to rats at 3 mg/kg before renal ischemia-reperfusion. In addition to the biochemical parameters, the renal function and the histological appearance were carefully evaluated. METHODS: Glutathione, adenine nucleotides and lipid peroxidation products were determined in kidneys reperfused for 2 and 24 hours after 90 minutes of ischemia. Renal function was assessed by plasma creatinine, and renal injury by histological examination. RESULTS: Reperfusion-induced glutathione oxidation, expressed as an oxidized-to-total glutathione ratio, was significantly attenuated both after 2 and 24 hours of reperfusion by treatment with U83836E. Adenosine triphosphate (ATP) was still significantly depleted after 24 hours in the control group, while at the same time treated animals had already recovered to baseline values. Lipid peroxidation products were significantly lower in lazaroid-groups both after 2 and 24 hours of reperfusion. Renal function after 24 hours of reperfusion was notably better in the treated rats. Histological examination confirmed the protective action of the drug. After 24 hours the control group showed large areas of parenchymal hemorrhage and necrosis with dilated tubules and blood vessel thrombosis, while treated animals showed small necrotic areas with a background of mild interstitial inflammatory cells. CONCLUSIONS: Our results suggest that there is a protective effect of U83836E in ischemia-reperfusion injury, in that tissue damage due to oxidative stress is reduced, thus ameliorating renal function impairment.

Suitability of the old fowl rectal caecum preparation for investigating the selectivity of beta-adrenergic drugs.

We tested alpha- and beta-adrenergic drugs on isolated strips of fowl rectal caecum from 14- to 16-week-old Warren hens. Basal tone and spontaneous motility were dose-dependently reduced by isoprenaline and all the selective beta-agonists tested (except xamoterol) with the following order of potency: isoprenaline=fenoterol=procaterol=clenbuterol>dobutamine> SR58611A. The results indicate that this tissue preparation consists almost entirely of beta2-adrenoceptors. This preparation may, therefore, be considered a suitable assay for discriminating beta1- from beta2-agonists according to their selectivity.

Differences between longitudinal and circular smooth muscle in beta-adrenergic control of motility of isolated equine ileum.

OBJECTIVE: To identify beta-adrenoceptor subtypes involved in motility inhibition of circular and longitudinal smooth muscle layers of equine ileum. SAMPLE POPULATION: Isolated strips of equine ileum circular smooth muscle and membrane preparations from circular and longitudinal muscle layers. PROCEDURE: Functional assays of circular muscle preparations and radioligand binding assays and measurements of cAMP production in smooth muscle membranes from circular and longitudinal layers. RESULTS: Selective beta-adrenergic agonists exerted inhibitory effects on circular muscle preparations. Binding studies of cell membranes indicated that the density and distribution of 3 beta-adrenoceptor subtypes did not differ between longitudinal and circular muscle layers. Measurement of cAMP production in membrane preparations of longitudinal and circular muscle after selective beta-stimulation confirmed presence of the 3 adenylate cyclase-coupled beta-adrenoceptor subtypes; however, preparations from the 2 layers had differing cAMP production efficacy. CONCLUSIONS: The data may partly explain the differing functional responses between circular and longitudinal muscle preparations. CLINICAL RELEVANCE: Findings support the important role of beta-atypical adrenoceptors in the inhibitory regulation of equine ileum motility.