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Nerve growth factor and its receptors, TrkA and p75NTR, are involved in inflammation and airways diseases, but their role in chronic obstructive pulmonary disease is still unclear and not well investigated. our data indicate the stage dependent variation of nerve growth factor and its receptors in chronic obstructive pulmonary disease progression. In fact, for the first time, this study evaluates the presence of nerve growth factor and its receptors in serum and in peripheral blood mononuclear cells of patients with different stages of chronic obstructive pulmonary disease compared to healthy subjects, non-smoker and current smoker. Serum monocyte chemoattractant protein-1, tumor necrosis factor-α, interleukin-10 and forced expiratory volume in 1 s were also analyzed. Compared to healthy subjects, chronic obstructive pulmonary disease patients presented a staging-dependent increase in serum nerve growth factor, negatively correlated to forced expiratory volume in 1 s and positively to monocyte chemoattractant Protein-1. The percentage of p75NTR+ peripheral blood mononuclear cells increased in early stages of chronic obstructive pulmonary disease (I–II), while TrKA+ peripheral blood mononuclear cells increased in late stages (III–IV). Our data demonstrate the involvement and modulation of nerve growth factor and its receptors in chronic obstructive pulmonary disease and in its staging.

The association of lysosomal dysfunction and neurodegeneration has been documented in several neurodegenerative diseases, including Alzheimer's Disease (AD). Herein, we investigate the association of lysosomal enzymes with AD at different stages of progression of the disease (mild and severe) or with mild cognitive impairment (MCI). We conducted a screening of two classes of lysosomal enzymes: glycohydrolases (β-Hexosaminidase, β-Galctosidase, β-Galactosylcerebrosidase, β-Glucuronidase) and proteases (Cathepsins S, D, B, L) in peripheral blood samples (blood plasma and PBMCs) from mild AD, severe AD, MCI and healthy control subjects. We confirmed the lysosomal dysfunction in severe AD patients and added new findings enhancing the association of abnormal levels of specific lysosomal enzymes with the mild AD or severe AD, and highlighting the difference of AD from MCI. Herein, we showed for the first time the specific alteration of β-Galctosidase (Gal), β-Galactosylcerebrosidase (GALC) in MCI patients. It is notable that in above peripheral biological samples the lysosomes are more sensitive to AD cellular metabolic alteration when compared to levels of Aβ-peptide or Tau proteins, similar in both AD groups analyzed. Collectively, our findings support the role of lysosomal enzymes as potential peripheral molecules that vary with the progression of AD, and make them useful for monitoring regenerative medicine approaches for AD.

Herbal medicines have been recently employed in research and clinical studies for the potential treatment of behavioral and psychological symptoms associated with Alzheimer's Disease (AD) and other types of dementia. The present study investigates the effect of trans-crocetin, an active constituent of Crocus sativus L., to restore in vitro the reduced ability of AD patients' monocytes to degrade amyloid-β(1-42) (Aβ42). CD14+ monocytes from 22 sporadic AD patients with moderate cognitive impairment were isolated; then, the role of trans-crocetin, purified from saffron extracts, was evaluated in terms of Aβ42 degradation rate through flow cytometry, as well as expression of cathepsin B by Western blotting. We observed that low micromolar doses of trans-crocetin enhanced Aβ42 degradation in AD monocytes through the upregulation of the lysosomal protease cathepsin B. CA074Me, a potent and selective cathepsin B inhibitor, counteracted such trans-crocetin-induced effect. These data suggest that the carotenoid trans-crocetin improves in vitro the clearance of Aβ42 through the involvement of cathepsin B, and this could be of value in developing a new anti-amyloid strategy in AD.

Osteoarthritis (OA) of the knee is the most common form of non-traumatic joint disease. Previous studies have shown the involvement of β-NGF and its receptors TrKA and p75NTR in OA-related pain, but their role in its pathogenesis is still unclear. The aim of our study was to investigate the amount of β-NGF and the expression levels of its receptors on cells isolated from synovial fluid and blood from OA patients who had undergone total knee arthroplasty, in order to check any possible correlation with the disease staging. Our results show a progressive stage-related increase of β-NGF and its receptors both in serum and synovial fluid. Furthermore, with respect to control subjects, OA patients show an increased amount of inflammatory monocytes along with an increased expression of β-NGF, TrKA and p75NTR. In conclusion, our study suggests a stage-related modulation of β-NGF and its receptors in the inflammatory process of OA.

This study investigated how the design of surface topography may stimulate stem cell differentiation towards a neural lineage. To this end, hydrogenated amorphous carbon (a-C:H) groove topographies with width/spacing ridges ranging from 80/40μm, 40/30μm and 30/20μm and depth of 24 nm were used as a single mechanotransducer stimulus to generate neural cells from human bone marrow mesenchymal stem cells (hBM-MSCs) in vitro. As comparative experiments, soluble brain-derived neurotrophic factor (BDNF) was used as additional biochemical inducer agent. Despite simultaneous presence of a-C:H micropatterned nanoridges and soluble BDNF resulted in the highest percentage of neuronal-like differentiated cells our findings demonstrate that the surface topography with micropatterned nanoridge width/spacing of 40/30μm (single stimulus) induced hBM-MSCs to acquire neuronal characteristics in the absence of differentiating agents. On the other hand, the alternative a-C:H ridge dimensions tested failed to induce stem cell differentiation towards neuronal properties, thereby suggesting the occurrence of a mechanotransducer effect exerted by optimal nano/microstructure dimensions on the hBM-MSCs responses.

Uterine leiomyosarcoma is a severe neoplasia resistant to conventional therapies. In previous studies, we have shown that human SK-UT-1 (ATCC HTB114) uterine leiomyosarcoma cell line secretes nerve growth factor (NGF) and expresses its receptors tyrosine kinase A receptor (TrKA) and low affinity nerve growth factor receptor (p75NTR). Furthermore, we have demonstrated that direct chemical inhibition or IgG neutralization of TrKA receptor induce apoptosis through p75NTR. In the present study, HTB114 cells were exposed to the phosphoinositide 3-kinase (PI3K) inhibitor LY294002 with and without β-NGF: apoptosis, cell cycle, activation of caspase-3 and protein kinase B (AKT) and TrKA/p75NTR phenotypic expression were evaluated. According to the type of exposure, LY294002 not only induced a relevant increase in apoptosis, but also produced a novel and unexpected phenotypic modulation of the NGF receptors with a downregulation of TrKA and an upregulation of p75NTR. This latter increase enhanced HTB114 apoptosis. Our study confirms that the interference on NGF transduction is a promising therapeutical approach in uterine leiomyosarcoma.

Background: Alzheimer’s disease (AD) is characterized by cognitive decline and hallmark neuropathologies, including b-amyloid (Ab). Therapeutic strategies for AD focus on reducing the production or deposition of Ab. Canines develop Ab neuropathology and cognitive decline with age similar to AD patients, making them a useful model for testing potential therapies. Immunization with Ab1-42 (IMM)in aged canines significantly decreases brain Ab and maintains executive function. However, behavioral enrichment (BEH) improves cognition without reducing brain Ab. We hypothesized that IMM combined with BEH would provide larger cognitive benefits and further reduce neuropathology, as compared to controls or individual IMM and BEH treatments alone. Methods: Aged beagles (10.513.6 y) were placed into groups: control (Alum adjuvant only), fibrillar Ab1-42 + Alum vaccine, BEH with Alum, and combination treatment (IMM+BEH). Animals were treated for 18 months. Cognition was measured throughout the study using various learning and memory tasks. Serum IgG antibody titers, cerebral spinal fluid (CSF) Ab levels, and insoluble brain Ab (formic acid extracted) were measured by ELISA. Ab plaque load (6E10, anti-Ab 1-42, Pyro Glu3), CAA (congo red), and microhemorrhages (prussian blue) were measured by immunohistochemistry. Results: Anti-Ab1-42 IgG responses in IMM animals increased significantly and were maintained. BEH significantly increased CSF Ab1-40, while no systematic effects were seen on Ab1-42 or total Ab. IMM significantly reduced both insoluble Ab1-40 and 1-42, as well as all forms of measured plaque load. An overall reduction in 6E10, and Ab 1-42 plaque load due to BEH was also seen. A significant additive affect from BEH and IMM was seen in clearance of Ab 1-42 plaque load. Conclusions: IMM successfully induced an immune response in treated animals and was maintained. IMM significantly reduced all forms of Ab pathology. Additionally, BEH increased CSFAb 1-40 and reduced 6E10 and Ab 1-42 plaque loads. A significant additive effect from BEH and IMM was seen in clearance of Ab 142 plaque load. While changes in cognition, CAA and microhemorrhages remain to be analyzed, it is expected that levels of CAA and microhemorrhages will be lower in IMM animals, and IMM+BEH will show greatest cognitive benefits.

Alzheimer's disease (AD), the most common form of dementia in elderly individuals, is characterized by neurofibrillary tangles, extracellular amyloid-β (Aβ) plaques and neuroinflammation. New evidence has shown that the lysosomal system might be a crossroad in which etiological factors in AD pathogenesis converge. This study shows that several lysosomal enzymes, including Cathepsin B, D, S, β-Galactosidase, α-Mannosidase, and β-Hexosaminidase, were less expressed in monocytes and lymphocytes from patients with a clinical diagnosis of AD dementia compared with cells from healthy controls. In vitro experiments of gain and loss of function suggest that down-regulation is a direct consequence of miR-128 up-regulation found in AD-related cells. The present study also demonstrates that miR-128 inhibition in monocytes from AD patients improves Aβ(1-42) degradation. These results could contribute to clarify the molecular mechanisms that affect the imbalanced Aβ production/clearance involved in the pathogenesis of AD.

Background NO is a key molecule involved in the regulation of cell survival, proliferation and differentiation in many cell types. In this study we investigated the contribution of NO during the differentiation of human peripheral blood hemopoietic stem cells (CD34 + HSCs) toward immunogenic dendritic cells (i-DCs). Methods We depleted autocrine NO production, using N G -monomethyl- l -arginine monoacetate ( l -NMMA) and paracrine NO , using oxy-hemoglobin (HbO 2 ) as a NO scavenger during in vitro differentiation of CD34 + HSCs to i-DCs. We monitored the NO level, cell proliferation, phenotype and differentiation potential. Results We found that the depletion of paracrine or autocrine NO correlated with (I) an active proliferation state at the end of differentiation, when control cells were not proliferating; (II) a significant reduction in the expression levels of differentiative markers (CD1a and HLA-DR) with a parallel high expression of the CD34 marker (III) with a retrieved clonogenic ability compared to control cells. Conclusions On the whole, our data indicate that the depletion of NO during the commitment stage blocks CD34 + HSC differentiation into i-DCs and maintains an undifferentiated, highly proliferating cell population, indicating/revealing a novel role for NO in the commitment of CD34 + HSCs into i-DCs. General significance The essential finding of the present study is that NO , produced in HSCs by NOS enzymes, may act as autocrine and paracrine effectors regulating the in vitro differentiation process of CD34 + -HSCs toward i-DCs.

Multiple epidemiological studies have shown that individuals affected by type-2 diabetes mellitus (T2DM) carry a 2-to-5-fold higher risk of developing Alzheimer's disease (AD) when compared to non-diabetic subjects. Thus, biochemical parameters that can be easily and routinely assessed for high-confidence evaluation of diabetic conditions leading to AD (AD-T2DM) are regarded as efficient tools aimed at early diagnosis and, in turn, timely AD treatment. In this regard, the activity of lysosomal glycohydrolases may of use, in light of the implication of these enzymes in early events that underlie AD pathology and an overt correlation, in diabetes, between altered metabolic homeostasis, abnormal glycohydrolase secretion in body fluids, and occurrence of diabetic complications. Based on marked up-regulation previously shown in a peripheral, cell-based model of AD, we selected β-Galactosidase, β-Hexosaminidase, and α-Mannosidase to discriminate T2DM from AD-T2DM subjects. A screen of 109, 114, and 116 patients with T2DM, AD and AD-T2DM, respectively, was performed by testing enzyme activities in both blood plasma and peripheral blood mononuclear cells. Compared to age-matched, healthy controls (n = 122), β-Galactosidase and β-Hexosaminidase activities markedly diverged across the three groups, whereas virtually unchanged values were observed for α-Mannosidase. In particular, plasma β-Galactosidase and β-Hexosaminidase levels were higher in patients with AD-T2DM compared to those with T2DM, suggesting different mechanisms leading to enzyme secretion. Statistical analyses based on ROC curves showed that both β-Galactosidase and β-Hexosaminidase activities, either intracellular or plasma-secreted, may be used to discriminate AD patients from controls and AD-T2DM from T2DM patients.

The effectiveness of therapeutic treatment based on regenerative medicine for degenerative diseases (i.e., neurodegenerative or cardiac diseases) requires tools allowing the visualization and analysis of the three-dimensional (3D) distribution of target drugs within the tissue. Here, we present a new computational procedure able to overcome the limitations of visual analysis emerging by the examination of a molecular signal within images of serial tissue/organ sections by using the conventional techniques. Together with the 3D anatomical reconstitution of the tissue/organ, our framework allows the detection of signals of different origins (e.g., marked generic molecules, colorimetric, or fluorimetric substrates for enzymes; microRNA; recombinant protein). Remarkably, the application does not require the employment of specific tracking reagents for the imaging analysis. We report two different representative applications: the first shows the reconstruction of a 3D model of mouse brain with the analysis of the distribution of theβ-Galactosidase, the second shows the reconstruction of a 3D mouse heart with the measurement of the cardiac volume.