Substantia nigra hyperechogenicity in essential tremor and Parkinson's disease: a longitudinal study.

BACKGROUND AND PURPOSE: Essential tremor (ET) and Parkinson's disease (PD) sometimes overlap in their clinical expression with ET preceding PD onset, often leading to misdiagnosis. Transcranial sonography (TCS) has been shown to be a valid and non-invasive diagnostic tool to identify early idiopathic PD and to differentiate it from ET. The purpose of this study was to investigate the relevance of substantia nigra hyperechogenicity in patients with ET. METHODS: A total of 138 patients (79 with PD, 59 with ET) and 50 matched controls underwent TCS examination at baseline. All patients were followed in a 3-year longitudinal assessment. RESULTS: A total of 10 subjects were excluded from the analysis due to the bilateral absence of a temporal acoustic window. During the follow-up period, 11 of the patients with ET developed new-onset parkinsonian features, without fulfilling criteria for PD diagnosis (ET+). Nine patients developed clinical features meeting diagnostic criteria for probable PD (ET-PD). Patients with ET- did not develop parkinsonian features. For each group, the maximum size of the substantia nigra hyperechogenicity was as follows: 5.62 ± 5.40 mm2 in the control group, 19.02 ± 14.27 mm2 in patients with PD, 9.15 ± 11.26 mm2 in patients with ET-, 20.05 ± 13.78 mm2 in patients with ET+ and 20.13 ± 13.51 mm2 in patients with ET-PD. ET-PD maximum values were significantly different from controls. Maximum values in patients with ET+ were different from both controls and patients with ET-. CONCLUSION: Substantia nigra hyperechogenicity in ET seems to represent a risk marker for developing early parkinsonian symptoms or signs in the 3 years following TCS assessment.

Diet and physical activity "defeated" Tuberil® in treatment of childhood obesity.

AIM: Childhood obesity is remarkably spreading worldwide, involving both industrialized and low-income countries. Its prevalence, outcome and socioeconomic impact call for the attention of medical community. We conducted a monocentric, open, two-arm, parallel-group study to evaluate the efficacy at reducing appetite and increasing dietary compliance of obese children of Tuberil®, a weight-loss supplement derived from potato and devoid of side effects. METHODS: We recruited participants, children with BMI ≥ 85th, through direct referrals in pediatrician's surgeries. Children were randomized to receive Tuberil® (group A) or nothing (group B), following a chronological order (A-B-A-B). Every child received a nutritionally balanced diet and had to record their appetite and to describe their meals in a diary. RESULTS: Even if we found a significant reduction in BMI, weight and waist circumference in both groups, no statistically significant differences between groups were noted. We did not found any significant differences in appetite between group A and B. CONCLUSION: Our data show that Tuberil® has no efficacy neither in reducing appetite in children nor in increasing dietary compliance. We believe that only a nutritionally balanced diet and our attention in verifying their compliance led to the reduction in BMI, weight and waist circumferences noted in our series.

Association between Neuropsychological Performance and CSF Profile in Subjective Cognitive Decline: Towards the Diagnosis of Preclinical AD.

BACKGROUND: In the perspective of novel treatments with disease-modifying drugs, a timely diagnosis of Alzheimer's' disease (AD) at preclinical phase represents a major issue. To this purpose, in clinical setting, there is the need to detect the earliest cognitive symptoms not yet fulfilling Mild Cognitive Impairment criteria, in order to proceed to biomarker assessment for diagnostic definition. In terms of cognitive performance, Subjective Cognitive Decline (SCD) is still a controversial entity, due to the difficulty of reliably measuring subtle deficits. OBJECTIVE: To evaluate the possibility to predict the presence of AD-like CSF pattern in SCD individuals, according to their neuropsychological performance assessed by means of both traditional and computerized measures. DESIGN: Retrospective study. SETTING: Clinical setting (Centre for Memory Disturbances, Section of Neurology, University Hospital of Perugia, Italy). PARTICIPANTS: 74 consecutive SCD subjects who underwent an in-depth (paper-pencil and computerized) neuropsychological assessment and CSF analysis for AD biomarkers (Aß42/Aß40 ratio, phospho-tau, total tau). MEASUREMENTS: Neuropsychological assessment was composed of traditional tests assessing five cognitive domains (verbal memory, attention, executive functions, language, visuo-spatial abilities) and computerized tasks from CAmbridge Neuropsychological Test Automated Battery (CANTAB) (Pattern Recognition Memory, Paired Associates Learning and Spatial Working Memory). According to their performance at traditional tests, SCD individuals were categorized into cognitively normal (CN) and subtle impaired (SI); with respect to CANTAB, they were defined as CANTAB- in presence of normal performance, and CANTAB+ in presence of at least one pathological score. The subgroup with completely normal performance was defined as CN/CANTAB-, and the subgroup with impairment in both measures as SI/CANTAB+. Differences in prevalence of A/T/N profile according to cognitive profiles were assessed by Fisher's exact text for count data. RESULTS: None of CN/CANTAB- subjects showed A+/T+ status. SI/CANTAB+ subjects showed a significantly high prevalence of A+/T+ profile (14/35, 40%, p=0.03 vs CN/CANTAB-). CONCLUSION: The neuropsychological profile may be of help in identifying SCD subjects requiring biomarker assessment. If confirmed in larger cohorts, the combination of traditional and computerized tests (namely, CANTAB) might represent a feasible strategy in clinical setting for carrying out biomarker assessment in individuals before the MCI stage. Detection of AD in these subjects would give them the highest chances to halt disease progression by means of disease modifying treatments.

The new low-toxic histone deacetylase inhibitor S-(2) induces apoptosis in various acute myeloid leukaemia cells.

Histone deacetylase inhibitors (HDACi) induce tumour cell cycle arrest and/or apoptosis, and some of them are currently used in cancer therapy. Recently, we described a series of powerful HDACi characterized by a 1,4-benzodiazepine (BDZ) ring hybridized with a linear alkyl chain bearing a hydroxamate function as Zn(++)--chelating group. Here, we explored the anti-leukaemic properties of three novel hybrids, namely the chiral compounds (S)-2 and (R)-2, and their non-chiral analogue 4, which were first comparatively tested in promyelocytic NB4 cells. (S)-2 and partially 4--but not (R)-2--caused G0/G1 cell-cycle arrest by up-regulating cyclin G2 and p21 expression and down-regulating cyclin D2 expression, and also apoptosis as assessed by cell morphology and cytofluorimetric assay, histone H2AX phosphorylation and PARP cleavage. Notably, these events were partly prevented by an anti-oxidant. Moreover, novel HDACi prompted p53 and α-tubulin acetylation and, consistently, inhibited HDAC1 and 6 activity. The rank order of potency was (S)-2 > 4 > (R)-2, reflecting that of other biological assays and addressing (S)-2 as the most effective compound capable of triggering apoptosis in various acute myeloid leukaemia (AML) cell lines and blasts from patients with different AML subtypes. Importantly, (S)-2 was safe in mice (up to 150 mg/kg/week) as determined by liver, spleen, kidney and bone marrow histopathology; and displayed negligible affinity for peripheral/central BDZ-receptors. Overall, the BDZ-hydroxamate (S)-2 showed to be a low-toxic HDACi with powerful anti-proliferative and pro-apototic activities towards different cultured and primary AML cells, and therefore of clinical interest to support conventional anti-leukaemic therapy.

Automated source of squeezed vacuum states driven by finite state machine based software.

In the last few decades, much effort has been made for the production of squeezed vacuum states in order to reduce quantum noise in the audio-frequency band. This technique has been implemented in all running gravitational-wave interferometric detectors and helped to improve their sensitivity. While the detectors are acquiring data for astrophysical observations, they must be kept in the operating condition, also called "science mode," that is, a state that requires the highest possible duty-cycle for all the instrumental parts and controls. We report the development of a highly automated setup for the generation of optical squeezed states, where all the required control loops are supervised by a software based on finite state machines; we took special care to grant ease of use, stability of operation, and possibility of auto-recovery. Moreover, the setup has been designed to be compatible with the existing software and hardware infrastructure of the Virgo detector. In this paper, we discuss the optical properties of this squeezing setup, the locking techniques, and the automation algorithms.

Precision-based exercise as a new therapeutic option for children and adolescents with haematological malignancies.

Children and adolescents with haematological malignancies (PedHM) are characterized by a severe loss of exercise ability during cancer treatment, lasting throughout their lives once healed and impacting their social inclusion prospects. The investigation of the effect of a precision-based exercise program on the connections between systems of the body in PedHM patients is the new frontier in clinical exercise physiology. This study is aimed at evaluating the effects of 11 weeks (3 times weekly) of combined training (cardiorespiratory, resistance, balance and flexibility) on the exercise intolerance in PedHM patients. Two-hundred twenty-six PedHM patients were recruited (47% F). High or medium frequency participation (HAd and MAd) was considered when a participant joined; > 65% or between 30% and < 64% of training sessions, respectively. The "up and down stairs'' test (TUDS), "6 min walking" test (6MWT), the "5 Repetition Maximum strength" leg extension and arm lateral raise test (5RM-LE and 5RM-ALR), flexibility (stand and reach), and balance (stabilometry), were performed and evaluated before and after training. The TUDS, the 5RM-LE and 5RM-ALR, and the flexibility exercises showed an increase in HAd and MAd groups (P < 0.05), while the 6MWT and balance tests showed improvement only in HAd group (P < 0.0001). These results support the ever-growing theory that, in the case of the treatment of PedHM, 'exercise is medicine' and it has the potential to increase the patient's chances of social inclusion.

Protein metabolism in tumor cells at various stages of growth in vivo.

Protein metabolism of Yoshida ascites hepatoma cells was studied in the early phase of logarithmic proliferation and in the following stage in which cell mass remains constant (resting phase). The rate of protein synthesis was measured by a short-time incorporation of [(8)H]lysine, while degradation was concurrently assessed by following the decrease of specific activity of [(14)C]lysine-labeled proteins. Most of the labeled amino acid injected intraperitoneally into the animal was immediately available for the tumor cells, with only a minor loss towards the extra-ascitic compartment. It was thus possible to calculate the dilution of the isotope in the ascitic pool of the lysine, which increased concurrently with the ascitic plasma volume. Amino acid transport capacity did not change in the log vs. the resting cells. This fact permitted the correction of the specific activity of the proteins synthesized by tumors in the two phases, taking into account the dilution effect. Protein synthesis was found to proceed at a constant rate throughout each of the two phases, although it was 30% lower during the resting as compared to the log phase. When cell mass attained the steady-state, protein degradation occurred at such a level as to balance the synthesis. Throughout the resting phase the amount of lysine taken up by the cells and renewed from the blood remained unchanged. Protein turnover, as studied in subcellular fractions, exhibited a similar rate in nuclei and microsomes, where it proceeded at a higher level than in mitochondria. On the whole, the results encourage the use of the Yoshida ascites hepatoma as a suitable model for studying protein turnover in relation to cell growth in vivo.

Design, synthesis and preliminary biological evaluation of new hydroxamate histone deacetylase inhibitors as potential antileukemic agents.

This study concerns the synthesis of new histone deacetylase inhibitors (HDACi) characterized by a 1,4-benzodiazepine ring used as the cap, joined through an amide function or a triple bond as connection units, to a linear alkyl chain bearing the hydroxamate function as Zn2+-chelating group. Biological tests performed in human acute promyelocytic leukemia NB4 cells showed that new hybrids can induce histone H3/H4 acetylation, growth arrest, and also apoptosis. Notably, chiral compounds exhibit stereoselective activity.

Magnetic coupling to the advanced Virgo payloads and its impact on the low frequency sensitivity.

We study the electromagnetic coupling of the Advanced Virgo (AdV) input mirror payload in response to a slowly time-varying magnetic field. As the problem is not amenable to analytical solution, we employ and validate a finite element (FE) analysis approach. The FE model is built to represent as faithfully as possible the real object, and it has been validated by comparison with experimental measurements. The intent is to estimate the induced currents and the magnetic field in the neighbourhood of the payload. The procedure found 21 equivalent electrical configurations that are compatible with the measurements. These have been used to compute the magnetic noise contribution to the total AdV strain noise. At the current stage of development, AdV seems to be unaffected by magnetic noise, but we foresee a non-negligible coupling once AdV reaches the design sensitivity.

WEB-2086 and WEB-2170 trigger apoptosis in both ATRA-sensitive and -resistant promyelocytic leukemia cells and greatly enhance ATRA differentiation potential.

PAF-receptor antagonists WEB-2086 and WEB-2170 (WEBs) have been previously shown to induce differentiation in murine and human leukemia cells. The present study describes the apoptotic-differentiative effect of WEBs in all-trans-retinoic acid (ATRA)-sensitive (NB4) and -resistant (NB4-007-6 and NB4-MR4) acute promyelocytic leukemia (APL) cell lines as well as blasts from patients with t(15;17) APL. NB4 cells exposed to 0.5-1 mM WEBs underwent striking growth arrest and massive apoptosis without appreciable differentiation; IC50 values after 3-day treatment of NB4 were 0.4 and 0.25 mM for WEB-2086 and WEB-2170, respectively. WEBs induced apoptosis also in the two ATRA-resistant NB4-007-6 and NB4-MR4 cell lines and in blasts from patients with t(15;17) APL. Moreover, subapoptotic WEBs acted synergistically with low-dose (0.025-0.05 microM) ATRA; this allowed to increase ATRA differentiation potential up to 40-fold and to improve both number and intensity of NBT-positive NB4 cells at definitely higher levels than with 1 muM ATRA alone. The powerful antiproliferative-apoptotic activities of WEBs in vitro on ATRA-sensitive, ATRA-resistant APL cells and blasts from patients with APL as well as drug capabilities to enhance ATRA differentiation potential suggested that these agents also due to their recognized tolerability in vivo might improve, alone or in combination, clinical treatment of APL.

Changes in CuZn-superoxide dismutase during induced differentiation of murine erythroleukemia cells.

Superoxide dismutase (SOD) activity in murine erythroleukemia cells (MELC) was determined during differentiation induced by hexamethylene bisacetamide. SOD levels in hexamethylene bisacetamide-treated MELC were about twice as high as those of controls. Dose response and kinetic experiments have shown that SOD activity variations are closely related to the amount of inducer and the duration of treatment in culture. Moreover, phorbol 12-myristate 13-acetate, which inhibits hexamethylene bisacetamide-induced MELC maturation, was also effective in reducing the extent of the SOD increase. SOD changes mainly involved the CuZn form of the enzyme, having a molecular weight of about 32,000 and a striking sensitivity to cyanide inhibition. In addition, the isoelectrophoretic analysis of CuZn-SOD from both treated and untreated cells yielded an identical pattern. This suggests that quantitative rather than qualitative enzyme changes occurred during MELC terminal division. SOD levels are directly related to the degree of differentiation and particularly to the amount of cytosolic hemoglobin, whose synthesis in committed cells is paralleled by a rise in enzyme activity. The SOD increase represents a distinctive marker of erythroleukemia maturation and might tentatively be interpreted as a cellular response to oxidative stress from hemoglobin autoxidation.

Megakaryocyte-like increase in ploidy of Friend's erythroleukemia cells induced to endoreplication by colcemid.

Previous work from this laboratory has shown that Friend's murine erythroleukemia cells (MELCs) express some bio-chemical traits of the megakaryocytic lineage. The supposed mixed erythroid/megakaryocytic nature of these cells has been investigated further by challenging MELCs with the antimicrotubule agent colcemid. This compound, at the concentration of 40 nM, was found to induce a striking arrest of cell growth without significant effects on viability. At the same time, the bulk of treated MELCs underwent a large increase in size to contain, after 3 days, as much as 4 times more proteins and 5 times more DNA than controls. As shown by high rates of 3H-thymidine incorporation, increase in DNA content was the result of active synthesis without completion of intervening mitosis according to a process that closely resembled endoreplication. Eventually, colcemidinduced MELCs presented multilobed nuclei and were arranged into discrete ploidy groups containing up to 16 N levels of DNA. Moreover, upon colcemid addition, MELCs initiated a polyploid response that was shown to continue, even in the absence of the inducer, to yield cells that became strongly positive for acetylcholinesterase (AChE) in the late stages of culture. These effects were compatible with a colcemid-induced commitment of MELCs to megakaryocyte differentiation, for which these cells seemed to be definitely programmed. The expression of megakaryocyte features in MELCs provided further evidence for the bipotentiality (erythroid/megakaryocytic) of this model.

Constitutive expression of a megakaryocytic functional property by murine erythroleukemia (Friend) cells: the incorporation of serotonin.

Murine Friend-derived erythroleukemia cells (MEL) are generally believed to be unipotential progenitors inducible to terminal erythroid differentiation. However, we found that MEL can constitutively incorporate significant amounts of radiolabeled serotonin ([3H]5-HT). Because this process is typical of cells belonging to the megakaryocytic lineage, we investigated the significance and mechanisms of 5-HT incorporation in the MEL system. We observed that: 1) normal murine erythroid cells and erythroid progenitors do not incorporate [3H]5-HT, as well as normal murine myeloid cells and the human myeloid cell line HL-60; on the other hand, the human erythroleukemia cell lines K562 and HEL, which have been shown to constitutively express megakaryocytic features, were able to incorporate [3H]5-HT; 2) MEL incorporated 5-HT by an active and saturable mechanism, dependent on temperature and sodium concentration in the medium; and 3) 5-HT uptake was very rapid. Moreover, because about 65% of cell-associated radioactivity was no longer displaced by the cold substrate, we assumed it to represent "true" cytoplasmic internalization. Finally, 5-HT incorporation by MEL was inhibited by clomipramine, ouabain, and reserpine, which are known inhibitors of 5-HT uptake in platelets. The commitment of MEL to terminal erythroid differentiation by hexamethylene bisacetamide or dimethyl sulfoxide greatly reduced the capacity to incorporate [3H]5-HT. These results seem to suggest that the MEL system, although mainly erythroid as regards its differentiation capability, constitutively expresses features of the megakaryocytic lineage, possibly disclosed by the ability to incorporate 5-HT. This hypothesis was further supported by the findings that 30%-40% of uninduced MEL were labeled by a polyclonal antibody raised against murine platelets that selectively recognized megakaryocytes in murine bone marrow smears.

Results of a national multicentric study on compliance to treatment with various disphosphonate formulations in patients with postmenopausal osteoporosis.

AIM: Compliance to pharmacological treatment for osteoporosis is crucial if the risk of fracture is to be reduced. Case series show that treatment with traditional bisphosphonates in the form of tablets has a compliance of between approximately 30% and 70%. The aims of this paper were to assess compliance to treatment with various formulations of bisphonates and to identify those at highest risk of discontinuation. METHODS: In this multicentre retrospective observational study, a population of 387 women diagnosed with postmenopausal osteoporosis under treatment with bisphosphonates (risedronate, ibandronate, alendronate in tablet form, alendronate in a fluid solution per os) was observed for at least a year. Demographic data and information pertaining to the type of drug taken, compliance to treatment, side effects, reasons for discontinuation, the basal examination and follow-up at 18 months and later were recorded. RESULTS AND CONCLUSION: Analysis of patient compliance to a prescribed treatment plan showed a significantly higher persistence (P<0.001) in the group taking alendronate in soluble solution form (83.3%) compared to the group taking any bisphosphonate in tablet form (66.7%). At the same time, patientspresenting comorbidity, receiving more than one therapy, not taking vitamin D, and in surgical menopause, risked discontinuation.

Italian Intersociety Recommendations on pain management in the emergency setting (SIAARTI, SIMEU, SIS 118, AISD, SIARED, SICUT, IRC).

BACKGRAUND: Pain is the primary reason for admission to the Emergency Department (ED). However, the management of pain in this setting is often inadequate because of opiophagia, fear of excessive sedation, and fear of compromising an adequate clinical assessment. METHODS: An intersociety consensus conference was held in 2010 on the assessment and treatment of pain in the emergency setting. This report is the Italian Intersociety recommendations on pain management in the emergency department setting. RESULTS: The list of level A recommendations includes: 1) use of IV acetaminophen for opioid sparing properties and reduction of opioid related adverse events; 2) ketamine-midazolam combination preferred over fentanyl-midazolam fentanyl-propofol in pediatric patients; 3) boluses of ketamine IV (particularly in the population under the age of 2 years and over the age of 13) can lead to impairment of the upper airways, including the onset of laryngospasm, requiring specific expertise and skills for administration; 4) the use of ketamine increases the potential risk of psychomotor agitation, which can happen in up to 30% of adult patients (this peculiar side effect can be significantly reduced by concomitant systemic use of benzodiazepines); 5) for shoulder dislocations and fractures of the upper limbs, the performance of brachial plexus block reduces the time spent in ED compared to sedation; 6) pain relief and the use of opioids in patients with acute abdominal pain do not increase the risk of error in the diagnostic and therapeutic pathway in adults; 7) in newborns, the administration of sucrose reduces behavioural responses to blood sampling from a heel puncture; 8) in newborns, breastfeeding or formula feeding during the procedure reduces the measures of distress; 9) in pediatric patients, non-pharmacological techniques such as distraction, hypnosis and cognitive-behavioural interventions reduce procedural pain caused by the use of needles; 10) in pediatric patients, preventive application of eutectic mixtures of prilocaine and lidocaine allows arterial and venous samples to be taken in optimum conditions; 11) in pediatric patients, the combination of hypnotics (midazolam) and N2O is effective for procedural pain, but may be accompanied by loss of consciousness. CONCLUSION: The diagnostic-therapeutic pathway of pain management in emergency should be implemented, through further interdisciplinary trials, in order to improve the EBM level of specific guidelines.

Premature induction of aging in sublethally H2O2-treated young MRC5 fibroblasts correlates with increased glutathione peroxidase levels and resistance to DNA breakage.

Human MRC5 fibroblasts, at different passages in cultures, were used as an in vitro model to assess variations and/or induction of aging parameters under basal conditions or following sublethal oxidative stress by H2O2. DNA sensitivities to oxidatively-induced breakage, rather than basal levels of damaged DNA, were significantly different between cultures at low and high population doubling level (PDL): old cells maintained most of their DNA integrity even at high concentrations of H2O2, while young cells showed more extensive DNA damage which developed in a dose-dependent fashion. However, young cells pretreated with low doses of H2O2 exhibited increased resistance against further oxidative damage to DNA thus reproducing a senescent-like profile of sensitivity. In turn, DNA from old cultures incubated in a NAD precursor-free medium was more prone to H2O2-induced strand breaks mimicking DNA sensitivity of young cells. The extent of oxidatively-induced DNA damage in MRC5 populations correlated inversely with the levels of glutathione peroxidase (GPx) activity that almost doubled when cells passed from the young to the senescent stage. In addition, H2O2-pretreatment of young cells induced an increase in GPx expression approaching old cell values and promoted also the premature appearance of neutral beta-galactosidase activity and decreased c-fos expression upon serum stimulation, both of which were assumed to be characteristic traits of the senescent phenotype.

Buprenorphine vs. morphine via the epidural route: a controlled comparative clinical study of respiratory effects and analgesic activity.

Twelve patients with intense or very intense pain of the non-incident type, secondary to neoplasia, were divided at random into two groups and treated with an epidural dose of 3 mg of morphine in 10 ml of glucose solution (6 patients = group M) or with 0.3 mg of buprenorphine in the same vehicle (6 patients = group B). None of the patients had previously been treated with opioids by any route. After first determining basal values, the following assessments were carried out: (1) evaluation of the analgesic effect of the drugs with checks at 30 min and at 1, 2, 3, 4, 6 and 18 h after administration, using a visual analogue scale, a numerical rating scale and a simple descriptive scale; and (2) evaluation of effects on respiration by means of checks at 30 and 90 min and at 6 and 18 h, on control of breathing indices (P0.1; VE; VA; Ti/Ttot; VT/Ti; RR), gas exchange indices (delta(A-a)O2; VD/VT; pAO2; R) and blood gas and acid-base indices (paO2; paCO2; pH; HCO3-). The data obtained were analyzed statistically using analysis of variance and Student's t test. The study results showed very similar analgesic efficacy for both treatments at a single dosage level of morphine (3 mg) compared to buprenorphine (0.3 mg), which was approximately 3 times greater than an equivalent parenteral dose of morphine (10 mg). Analysis of the results revealed statistically, though not clinically, significant changes in respiratory function indices, only in the buprenorphine-treated group. The effects of buprenorphine on respiratory function, when administered epidurally at the above dosage, are less favourable than those of morphine in the early measurements, probably because of its greater systemic absorption; nevertheless, the risk of delayed respiratory depression appears to be less after buprenorphine than after morphine.

Increased plasma levels of soluble CD40, together with the decrease of TGF beta 1, as possible differential markers of Alzheimer disease.

Alzheimer's disease (AD) is a progressive neurodegenerative illness and the most frequent cause of dementia in the elderly. The identification of activated microglia within neuritic plaques, coupled with the presence of numerous inflammatory proteins, suggests that inflammation is an integral part of the pathogenetic process in AD. In the present paper we have investigated the levels of circulating inflammatory mediators as potential AD biomarkers concentrating essentially on (a) soluble CD40 (sCD40), a member of the tumor necrosis factor receptor superfamily lacking the membrane-associated endodomain by alternative splicing, and (b) transforming growth factor (TGF)-beta 1, a cytokine deeply involved in AD and playing a protective role on CNS. Decrease of TGF-beta1 in AD patients could enhance the effects of pro-inflammatory cytokines produced by activated microglia as well as the expression of factors, such as the CD40/CD40 ligand complex, by microglia and astrocytes. Total venous blood samples were obtained from 33 patients with clinical diagnosis of possible late-onset AD, 40 healthy age-matched and 11 healthy young individuals. A significant increase of sCD40 levels plasma of AD patients versus healthy controls was measured, concomitantly with a decrease in TGF-beta1 concentration. These variations, however, showed no correlation with the expression of ApoE epsilon 4 allele, which was determined in order to assess the different frequency of this risk factor between AD and control groups. Since no comparable modifications were detected in patients affected by Parkinson's disease or non-AD-based dementia, we propose that sCD40 and TGF-beta1 plasma levels might represent possible differential biomarkers of AD, and be useful pre-mortem to support the clinical diagnosis of late-onset AD.

Enhanced proteolytic activities in cultured fibroblasts of Alzheimer patients are revealed by peculiar transketolase alterations.

Characteristic alterations of transketolase (TK) in extracts from cultured Alzheimer fibroblasts have previously been reported (Paoletti et al. (1990) Biochem. Biophys. Res. Commun., 172: 396-401). These abnormalities, encountered in 9 out of 13 Alzheimer patients, were revealed following isoelectric focusing and consisted of enzyme forms having unusually high alkaline pI values (alkaline bands). The present work has shown that immunologically detected alkaline bands were progressively expressed when Alzheimer fibroblasts were incubated for three weeks without medium changes. Full expression of the altered enzyme pattern was not linked to relative cell density in the petri dish; rather, it appeared to be dependent directly on the time elapsed since cell confluence was reached. Alkaline bands could artificially be induced also in both crude and pure TK preparations from normal cells by a treatment with commercial proteases, particularly chymotrypsin. Moreover, specific inhibitors of endogenous cysteine-proteases were capable of abolishing TK alkaline bands in Alzheimer fibroblasts thus turning a pathological into a normal enzyme pattern. Results obtained suggest that Alzheimer fibroblasts contain enhanced Ca(2+)-independent cysteine-proteolytic activities as compared to normal and other pathological cells. These enzymes, exhibiting chymotrypsin-like activity, might exert their degradative effects at the time of cell extraction using TK and probably other cell components as potential substrates. However, peculiar TK abnormalities represent so far an useful biochemical marker detectable in fibroblasts of living Alzheimer patients and closely associated to this neurological disorder.

A spectroscopic investigation of cobalt(II) substituted alkaline phosphatase.

The electronic and 1H NMR spectra are reported for the cobalt(II) alkaline phosphatase (EC 3.1.3.1.) system at pH around 6 in the range 0-2 mol of cobalt per mol protein. It is shown that under the present experimental conditions cobalt(II) selectively populates the A sites. Three isotropically shifted NH signals have been detected in the A site that indicate the presence of three histidines in the coordination sphere of cobalt(II). The electronic spectra and the nuclear relaxation properties are consistent with pentacoordination of cobalt(II) in the A site. The finding of reproducible preparation routes for the derivatives, and of appropriate experimental conditions for the observation of their 1H NMR spectra, open new possibilities for the spectroscopic investigation of alkaline phosphatase.