Colocalization Analysis of Peripheral Myelin Protein-22 and Lamin-B1 in the Schwann Cell Nuclei of Wt and TrJ Mice

Myelination of the peripheral nervous system requires Schwann cells (SC) differentiation into the myelinating phenotype. The peripheral myelin protein-22 (PMP22) is an integral membrane glycoprotein, expressed in SC. It was initially described as a growth arrest-specific (gas3) gene product, up-regulated by serum starvation. PMP22 mutations were pathognomonic for human hereditary peripheral neuropathies, including the Charcot-Marie-Tooth disease (CMT). Trembler-J (TrJ) is a heterozygous mouse model carrying the same pmp22 point mutation as a CMT1E variant. Mutations in lamina genes have been related to a type of peripheral (CMT2B1) or central (autosomal dominant leukodystrophy) neuropathy. We explore the presence of PMP22 and Lamin B1 in Wt and TrJ SC nuclei of sciatic nerves and the colocalization of PMP22 concerning the silent heterochromatin (HC: DAPI-dark counterstaining), the transcriptionally active euchromatin (EC), and the nuclear lamina (H3K4m3 and Lamin B1 immunostaining, respectively). The results revealed that the number of TrJ SC nuclei in sciatic nerves was greater, and the SC volumes were smaller than those of Wt. The myelin protein PMP22 and Lamin B1 were detected in Wt and TrJ SC nuclei and predominantly in peripheral nuclear regions. The level of PMP22 was higher, and those of Lamin B1 lower in TrJ than in Wt mice. The level of PMP22 was higher, and those of Lamin B1 lower in TrJ than in Wt mice. PMP22 colocalized more with Lamin B1 and with the transcriptionally competent EC, than the silent HC with differences between Wt and TrJ genotypes. The results are discussed regarding the probable nuclear role of PMP22 and the relationship with TrJ neuropathy.This research was funded by the Comisión Sectorial de Investigación Científica de la Universidad de la República (CSIC I+D, 2013), Agencia Nacional de Investigación e Innovación (ANII, FCE_1_2019_1_155539) and Programa de Desarrollo de Ciencias Básicas (PEDECIBA, annual fund allocation for academics).S

Identificación de un mecanismo de segregación y eliminación de un cromosoma supernumerario en núcleos de células trisómicas humanas

Tribunal: Dr. José Badano, Dra. Rossana Bonomi, Dra. Cristina Mazzella

Acute Genetic Damage Induced by Ethanol and Corticosterone Seems to Modulate Hippocampal Astrocyte Signaling

Astrocytes maintain CNS homeostasis but also critically contribute to neurological and psychiatric disorders. Such functional diversity implies an extensive signaling repertoire including extracellular vesicles (EVs) and nanotubes (NTs) that could be involved in protection or damage, as widely shown in various experimental paradigms. However, there is no information associating primary damage to the astrocyte genome, the DNA damage response (DDR), and the EV and NT repertoire. Furthermore, similar studies were not performed on hippocampal astrocytes despite their involvement in memory and learning processes, as well as in the development and maintenance of alcohol addiction. By exposing murine hippocampal astrocytes to 400 mM ethanol (EtOH) and/or 1 μM corticosterone (CTS) for 1 h, we tested whether the induced DNA damage and DDR could elicit significant changes in NTs and surface-attached EVs. Genetic damage and initial DDR were assessed by immunolabeling against the phosphorylated histone variant H2AX (γH2AX), DDR-dependent apoptosis by BAX immunoreactivity, and astrocyte activation by the glial acidic fibrillary protein (GFAP) and phalloidin staining. Surface-attached EVs and NTs were examined via scanning electron microscopy, and labeled proteins were analyzed via confocal microscopy. Relative to controls, astrocytes exposed to EtOH, CTS, or EtOH+CTS showed significant increases in nuclear γlH2AX foci, nuclear and cytoplasmic BAX signals, and EV frequency at the expense of the NT amount, mainly upon EtOH, without detectable signs of morphological reactivity. Furthermore, the largest and most complex EVs originated only in DNA-damaged astrocytes. Obtained results revealed that astrocytes exposed to acute EtOH and/or CTS preserved their typical morphology but presented severe DNA damage, triggered canonical DDR pathways, and early changes in the cell signaling mediated by EVs and NTs. Further deepening of this initial morphological and quantitative analysis is necessary to identify the mechanistic links between genetic damage, DDR, cell-cell communication, and their possible impact on hippocampal neural cells

Biological and dosimetric evaluation of [11C]S-adenosyl methionine as a potential agent for prostate cancer diagnosis

Introduction: [11C]Choline ([11C]COL) has been widely used for prostate cancer diagnosis; however, this radiopharmaceutical is not recommended for patients with a low absolute PSA value (< 1 ng/mL) due to its limited sensitivity and specificity. The enzyme glycine N-methyltransferase is overexpressed during prostate cancer progression. It catalyses the methylation of glycine using S-adenosyl methionine (SAM or AdoMet) as a substrate. The authors have previously reported the automated radiosynthesis of [11C]SAM as a potential agent in the diagnosis of aggressive prostate cancer. In this study, a biological and dosimetric evaluation of [11C]SAM was performed. Results: The evaluation of [11C]SAM in a control group of healthy mouse model showed a relatively high tracer uptake in the kidneys and a rapid blood clearance. Most activity was eliminated in the urine. In a PC3 prostate cancer xenograft tumour model, [11C]SAM tumour uptake was significantly higher in relation to [11C]COL.The human dosimetry of [11C]SAM was estimated by extrapolating the preclinical results. The mean effective dose was 8.17 x 10-3 mSv/MBq and 2.49 x 10-3 mSv/MBq without and with bladder voiding, respectively. The results for kidneys in humans were comparable to those previously described for [11C]COL. Conclusions: The PET/CT studies showed a statistically higher in vivo tumour uptake of [11C]SAM compared to [11C]COL for the cancer xenograft model. The absorbed dose estimations of major organs and the effective dose were determined. The results suggested that [11C]SAM may be a potential PET tracer for prostate cancer diagnosis.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Ciencias Atómicas Nucleares y Moleculares (CICANUM