Growth control mechanisms in neuronal regeneration

AbstractNeurons grow during development and extend long axons to make contact with their targets with the help of an intrinsic program of axonal growth as well as a range of extrinsic cues and a permissive milieu. Injury events in adulthood induce some neuron types to revert to a regenerative state in the peripheral nervous system (PNS). Neurons from the central nervous system (CNS), however, reveal a much lower capacity for regenerative growth. A number of intrinsic regeneration-promoting mechanisms have been described, including priming by calcium waves, epigenetic modifications, local mRNA translation, and dynein-driven retrograde transport of transcription factors (TFs) or signaling complexes that lead to TF activation and nuclear translocation. Differences in the availability or recruitment of these mechanisms may partially explain the limited response of CNS neurons to injury

Activated carbons : In vitro affinity for ochratoxin A and deoxynivalenol and relation of adsorption ability to physicochemical parameters

In vitro affinity tests were conducted to test the effectiveness of 19 activated carbons (ACs), hydrates sodium calcium aluminosilicate (HSCAS) and sepiolite (S) in binding ochratoxin A (OA) and deoxynivalenol (DON) from solution. Relationships between adsorption ability and physicochemical parameters of ACs (surface area, iodine number, methylene blue index) were tested. When 5 ml of a 4-micrograms/ml aqueous solution of OA was treated with 2 mg of AC, the ACs adsorbed 0.80 to 99.86% of the OA. HSCAS and S were not effective in binding OA. In two saturation tests carried out with increased amounts of OA (5 ml of 10-and 50-micrograms/ml aqueous solutions of OA, respectively) three ACs also showed high adsorption ability (adsorbing 92.23 to 96.57% of the OA). When 5 ml of a 4-micrograms/ml aqueous solution of DON was treated with 10 mg of AC, ACs adsored 1.83 to 98.93% of the DON. HSCAS and S were not effective in binding DON. An overall relation of adsorption ability to the physicochemical parameters of ACs was observed. The methylene blue index was more reliable than iodine number and surface area in predicting ability of ACs to adsorb OA and DON. Based on the data observed on the xxxxx eh present study as well as on aflatoxin B1 and fumonisin B1 from previous studies, it is concluded that ACs have high in vitro affinity for chemically different mycotoxins, and can be considered as potential multi-mycotoxin-sequestering agents. However, the ability to bind the main mycotoxins singly or in combination should be confirmed by in vivo investigations. Moreover, information on the amounts of AC to be added to feeds, and on the possible long-term effect on absorption of essential nutrients are needed

Activated Carbons: In Vitro Affinity for Aflatoxin B1 and Relation of Adsorption Ability to Physicochemical Parameters

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Reduction of Carryover of Aflatoxin from Cow Feed to Milk by Addition of Activated Carbons.

According to a double-reversal experimental design on 12 late-lactation Friesian cows the effect of two activated carbons (ACs) (CAC1 and CAC2) and a hydrated sodium calcium aluminosilicate (HSCAS) on carryover of aflatoxin B1 (AFB1) from feed to aflatoxin M1 (AFM1) in milk was determined. Cows were fed a basal diet containing AFB1 naturally contaminated corn meal and copra, During week 1 cows were fed diets containing AFB1 alone (11.28 μg of AFB1/kg of feed); in week 2 the diets contained AFB1 plus 2.0% sorbent; and in week 3 the diets again contained AFB1 alone (13.43 μg of AFB1/kg of feed). ACs reduced the analytical content of AFB1 in the pelleted feed by from 40.6% to 73.6%, whereas reduction by HSCAS was 59.2%, The AFM1 concentrations in milk in weeks 1 and 3 were higher than that in week 2, Decreases in the AFM1 excreted in the milk by addition to feed of 2% of the sorbents ranged from 22% to 45%. CAC1 and HSCAS were significantly different from each other in reducing the AFM1concentration in milk (45.3% versus 32.5%); these reductions were significantly higher than that of CAC2 (22.0%). Carryover reduction by addition of CAC1 (50%) was significantly higher than that of HSCAS (36%). Addition of 2% CAC2 did not allow pelleting of feed because of the caking action of this carbon, The lower performance of CAC2 could be related to the unsuccessful pelleting. The addition of ACs did not influence feed intake, milk production, milk composition, or body weight. Our results suggest that ACs, high-affinity sorbents for AFB1 in vitro, are efficacious in reducing AFB1 carryover from cow feed to milk. Further in vivo investigations should establish lower amounts of ACs which can be efficacious

Development of 3D culture scaffolds for directional neuronal growth using 2-photon lithography

Conventional applications of transplant technology, applied to severe traumatic injuries of the nervous system, have met limited success in the clinics due to the complexity of restoring function to the damaged tissue. Neural tissue engineering aims to deploy scaffolds mimicking the physiological properties of the extracellular matrix to facilitate the elongation of axons and the repair of damaged nerves. However, the fabrication of ideal scaffolds with precisely controlled thickness, texture, porosity, alignment, and with the required mechanical strength, features needed for effective clinical applications, remains technically challenging. We took advantage of state-of-the-art 2-photon photolithography to fabricate highly ordered and biocompatible 3D nanogrid structures to enhance neuronal directional growth. First, we characterized the physical and chemical properties and proved the biocompatibility of said scaffolds by successfully culturing primary sensory and motor neurons on their surface. Interestingly, axons extended along the fibers with a high degree of alignment to the pattern of the nanogrid, as opposed to the lack of directionality observed on flat glass or polymeric surfaces, and could grow in 3D between different layers of the scaffold. The axonal growth pattern observed is highly desirable for the treatment of traumatic nerve damage occurring during peripheral and spinal cord injuries. Thus, our findings provide a proof of concept and explore the possibility of deploying aligned fibrous 3D scaffold/implants for the directed growth of axons, and could be used in the design of scaffolds targeted towards the restoration and repair of lost neuronal connections

Mycotoxin occurrence in maize produced in Northern Italy over the years 2009–2011: focus on the role of crop related factors

The occurrence of mycotoxins associated with Fusarium spp. and Aspergillus flavus in Northern Italy, and the role of cropping systems, were investigated on 140 field samples collected over the years 2009–2011. Samples were analysed for fumonisins B1 and B2 (FBs), aflatoxins B1, B2, G1 and G2 (AFs), deoxynivalenol (DON) and zearalenone (ZEN) using validated analytical methods. Information on: maize hybrid, preceding crop, tillage applied, mineral nutrition, pest and disease control, severity of European Corn Borer (ECB) attack, sowing and harvesting dates, kernel moisture at harvesting and longitude of the sampled province, were also collected. During this period there were distinct differences in FBs and AFs concentrations between years and geographic origins, and very low contamination with DON and ZEN was always found. The incidence of AFs exceeded 75% across all samples, and was almost 100% for FBs. The meteorological trends were quite different in the 3 years surveyed. 2009 was the coldest in June and the warmest in August, 2010 the most humid, and in 2011 cold weather occurred during flowering and dry conditions during ripening. The run of a logistic equation with the backward stepwise approach selected three parameters, (seeding week, ECB severity and longitude of sampling province) to predict AFB1 contamination and four parameters (year, sowing week, ECB severity and longitude of sampling province) to predict FB contamination. The internal validation gave good results, with 76% correct predictions. The probability of harvesting maize with more than 5 µg kg-1 of AFB1 varied between 86 and 5%, and the probability of harvesting maize with more than 4,000 µg kg-1 of FBs varied between 81 and 2%, respectively, for conducive and non-conducive environments. Therefore, considerable variability was found even if a limited area and only 3 years were considered

Anterograde Axonal Transport in Neuronal Homeostasis and Disease

Neurons are highly polarized cells with an elongated axon that extends far away from the cell body. To maintain their homeostasis, neurons rely extensively on axonal transport of membranous organelles and other molecular complexes. Axonal transport allows for spatio-temporal activation and modulation of numerous molecular cascades, thus playing a central role in the establishment of neuronal polarity, axonal growth and stabilization, and synapses formation. Anterograde and retrograde axonal transport are supported by various molecular motors, such as kinesins and dynein, and a complex microtubule network. In this review article, we will primarily discuss the molecular mechanisms underlying anterograde axonal transport and its role in neuronal development and maturation,including the establishment of functional synaptic connections.We will then provide an overview of the molecular and cellular perturbations that affect axonal transport and are often associated with axonal degeneration. Lastly, we will relate our current understanding of the role of axonal trafficking concerning anterograde trafficking of mRNA and its involvement in the maintenance of the axonal compartment and disease

Detection of Merkel Cell Polyomavirus (MCPyV) DNA and Transcripts in Merkel Cell Carcinoma (MCC)

Merkel cell polyomavirus (MCPyV) is the etiological agent of the majority of Merkel cell carcinoma (MCC): a rare skin tumor. To improve our understanding of the role of MCPyV in MCCs, the detection and analysis of MCPyV DNA and transcripts were performed on primary tumors and regional lymph nodes from two MCC patients: one metastatic and one non-metastatic. MCPyV-DNA was searched by a quantitative polymerase chain reaction (qPCR), followed by the amplification of a Large T Antigen (LTAg), Viral Protein 1 (VP1) and Non-Coding Control Region (NCCR). LTAg and VP1 transcripts were investigated by reverse-transcription PCR (RT-PCR). Viral integration was also studied, and full-length LTAg sequencing was performed. qPCR revealed that the primary tumor of both patients and the lymph node of one patient was positive for the small t-antigen, with an average value of 7.0 × 102 copies/µg. The same samples harbored LTAg, NCCR and VP1 DNA. Sequencing results showed truncated LTAg with the conserved retinoblastoma (Rb) protein binding motif and VP1 and NCCR sequences identical to the MCC350 strain. RT-PCR detected LTAg but not VP1 transcripts. The MCPyV genome was integrated into the primary tumor of both patients. The results confirmed the connection between MCPyV and MCC, assuming integration, LTAg truncation and Rb sequestration as key players in MCPyV-mediated oncogenesis

Ultrastructural localization of tyrosine hydroxylase in human peripheral blood mononuclear cells: effect of stimulation with phytohaemagglutinin

Using immunocytochemistry coupled to fluorescence and electron microscopy, we investigated the expression and ultrastructural localization of tyrosine hydroxylase (TH, EC 1.14.16.2), the rate-limiting enzyme in the biosynthesis of catecholamines, in human peripheral blood mononuclear cells (PBMCs), with PC12 cells as positive controls. In unstimulated PBMCs, TH-specific immunoreactivity was localized to the plasma membrane. However, after stimulation with the polyclonal mitogen phytohaemagglutinin (PHA), TH immunoreactivity was almost completely localized to electron-dense cytoplasmic granules, which resembled those found in PC12. TH-positive granules, however, were larger (300-500 nm) than in PC12 cells (100-200 nm). Flow cytometry analysis of TH expression showed about 46-50% positive cells in unstimulated PBMCs and in PHA-stimulated PBMCs in the G0/G1 phase of the cell cycle, but more than 80% positive cells in PHA-stimulated PBMCs in the S+G2/M phase. In agreement with previous observations, PHA stimulation also induced de novo expression of TH mRNA as well as increased intracellular catecholamine content, suggesting the occurrence of TH upregulation at the level of both gene expression and enzyme activity. The ultrastructural localization of TH in human PBMCs seems therefore regulated by cell stimulation and related to the functional activity of the enzyme