Sensorimotor cortex ablation induces time-dependent response of ACTH cells in adult rats: Behavioral, immunohistomorphometric and hormonal study

Traumatic brain injury (TBI) represents a serious event with far reaching complications, including pituitary dysfunction. Pars distalis corticotropes (ACTH cells), that represent the active module of hypothalamo-pituitary-adrenocortical axis, seem to be affected as well. Since pituitary failure after TBI has been associated with neurobehavioral impairments the aim of this study was to evaluate the effects of TBI on recovery of motor functions, morphology and secretory activity of ACTH cells in the pituitary of adult rats. Wistar male rats, initially exposed to sensorimotor cortex ablation (SCA), were sacrificed at the 2nd, 7th, 14th and 30th days post-surgery (dps). A beam walking test was used to evaluate the recovery of motor functions. Pituitary glands and blood were collected for morphological and hormonal analyses. During the first two weeks post-injury increased recovery of locomotor function was detected, reaching almost the control value at day 30. SCA induces significant increase of pituitary weights compared to their time-matched controls. The volume of ACTH-immunopositive cells was reduced at the 7th dps, while at the 14th dps their volume was enlarged, in comparison to corresponding sham controls. Volume density of ACTH cells was increased only at 14th dps, while at day 30 this increase was insignificant. The plasma level of ACTH transiently increased after the injury. The most pronounced changes were observed at the 7th and 14th dps, and were followed by decrease toward control levels at the 30th dps. Thus, temporal changes in the hypothalamic-pituitary-adrenal axis after traumatic brain injury appear to correlate with the recovery process. (C) 2013 Elsevier Inc. All rights reserved.Ministry of Education, Science and Technological Development of the Republic of Serbia {[}III 41014, ON 173009

Effect of stab injury in the rat cerebral cortex on temporal pattern of expression of neuronal cytoskeletal proteins: An immunohistochemical study

Compelling evidence now points to the critical role of the cytoskeleton in neurodegeneration. In the present study, using an immunohistochemical approach, we have shown that cortical stab injury (CSI) in adult Wistar rats significantly affects temporal pattern of expression of neurofilament proteins (NFs). a major cytoskeleton components of neurons, and microtubule-associated proteins (MAP2). At 3 days post-injury (dpi) most of the NFs immunoreactivity was found in pyknotic neurons and in fragmentized axonal processes in the perilesioned cortex. These cytoskeletal alterations became more pronounced by 10 dpi. At the subcellular level CSI also showed significant impact on NFs and MAP-2 expression. Thus, at 3 dpi most of the dendrites disappeared, while large neuronal somata appeared like open circles pointing to membrane disintegration. Conversely, at 10 dpi neuronal perikarya and a few new apical dendrites were strongly labeled. Since aberrant NF phosphorylation is a pathological hallmark of many human neurodegenerative disorders, as well as is found after stressor stimuli, the present results shed light into the expression of neurofilaments after the stab brain injury. (C) 2014 Elsevier GmbH. All rights reserved.Ministry of Education, Science and Technological Development of the Republic of Serbia {[}III41014

Cortical Ablation Induces Time-Dependent Changes in Rat Pituitary Somatotrophs and Upregulates Growth Hormone Receptor Expression in the Injured Cortex

The pituitary appears to be vulnerable to brain trauma, and its dysfunction is a common feature after traumatic brain injury. The role of pituitary growth hormone (GH) in brain repair after injury has been envisaged, but more studies must be performed to understand completely the importance of GH in these processes. Because some of the neuroprotective effects of GH are mediated directly through the GH receptor (GHR), we examined GHR expression in the rat cerebral cortex after sensorimotor cortex ablation. RT-PCR, immunohistochemistry, and double immunofluorescence had been performed to analyze the correlation between GHR expression in the injured cortex and activity of GH cells in the pituitary. Our results showed that the volume of GH-immunopositive cells was reduced at days 2 and 7 postsurgery (dps), and volume density of GH cells was significantly decreased at 14 dps, all compared with appropriate sham controls. At 30 dps all investigated parameters had returned to control level. In the injured cortex, GHR expression was transiently upregulated. Increased GHR immunoreactivity was observed in reactive astrocytes at 7 and particularly at 14 dps. In neuronal cells, an increase of GHR immunoreactivity was seen in neuronal cell bodies and well-defined primary dendrites at 14 and especially at 30 dps. The results presented here suggest that, during recovery from brain injury, changes in activity of pituitary GH cells result in upregulation of GHR that may have a role in neuronal arborization and glial proliferation in the injured cortex. (C) 2014 Wiley Periodicals, Inc.Ministry of Education, Science, and Technological Development of the Republic of Serbia {[}III 41014, ON 173009

Tiazofurin modulates lipopolysaccharide-activated microglia in vitro

Tiazofurin is a purine nucleoside analogue, with a broad spectrum of antitumoral and anti-inflammatory properties. In the present study, we have investigated the effect of tiazofurin on microglial inflammatory response to lipopolysaccharide in vitro. The cytotoxic effect of the drug was examined by sulforhodamine B assay. The Griess method was used to quantify nitrite production. Microglial morphology was assessed by measuring cell body size. Release of the pro-inflammatory cytokines, tumor necrosis factor-alpha, interleukin-1 beta, interleukin-6, and the anti-inflammatory cytokine interleukin-10, were evaluated by enzyme-linked immunosorbent assay. Our data showed that tiazofurin decreased the number of activated microglia, lowered nitric oxide production and reduced the average cell surface of these cells. Tiazofurin reduced tumor necrosis factor-alpha, interleukin-6 and increased interleukin-10 secretion. Conversely, this drug promoted the release of interleukin-1 beta. Results obtained in this study indicate that TR displayed both anti-and pro-inflammatory modulation of activated microglia that could be relevant for its antitumor action within the central nervous system.Ministry of Education, Science and Technological Development, Republic of Serbia {[}III41014

Ribavirin shows immunomodulatory effects on activated microglia

Ribavirin (RBV) is synthetic purine nucleoside analogue, licensed as anti-viral drug that displays immunomodulatory actions on various immune cells. Our previous ex vivo studies have demonstrated immunosuppressive effects of RBV on reactive T-lymphocytes in experimental autoimmune encephalomyelitis. Here, we examined the effects of RBV on inflammatory response of microglia. RBV potency to down-regulate microglia inflammatory response was assessed by measuring microglia cell body size, and the production of nitric oxide (NO) and pro-and anti-inflammatory cytokines. RBV exerted cytotoxic effects on LPS-stimulated microglia, leaving non-stimulated microglia unaffected. The exposure of activated microglia to RBV led to: decrease in the level of NO as a result of decreased cell number, lower average cell surface, the reduction of membrane ruffling, the suppression of interleukin-6 release and promoted interleukin-10 production. On the other hand, RBV promoted LPS-induced interleukin-1 beta release. Our results imply that RBV is a complex immunomodulator showing both anti-and pro-inflammatory effects on activated microglia.Ministry of Education, Science and Technological Development of the Republic of Serbia {[}III41014

Repetitive Hyperbaric Oxygenation Attenuates Reactive Astrogliosis and Suppresses Expression of Inflammatory Mediators in the Rat Model of Brain Injury

The exact mechanisms by which treatment with hyperbaric oxygen (HBOT) exerts its beneficial effects on recovery after brain injury are still unrevealed. Therefore, in this study we investigated the influence of repetitive HBOT on the reactive astrogliosis and expression of mediators of inflammation after cortical stab injury (CSI). CSI was performed on male Wistar rats, divided into control, sham, and lesioned groups with appropriate HBO. The HBOT protocol was as follows: 10 minutes of slow compression, 2.5 atmospheres absolute (ATA) for 60 minutes, and 10 minutes of slow decompression, once a day for 10 consecutive days. Data obtained using real-time polymerase chain reaction, Western blot, and immunohistochemical and immunofluorescence analyses revealed that repetitive HBOT applied after the CSI attenuates reactive astrogliosis and glial scarring, and reduces expression of GFAP (glial fibrillary acidic protein), vimentin, and ICAM-1 (intercellular adhesion molecule-1) both at gene and tissue levels. In addition, HBOT prevents expression of CD40 and its ligand CD40L on microglia, neutrophils, cortical neurons, and reactive astrocytes. Accordingly, repetitive HBOT, by prevention of glial scarring and limiting of expression of inflammatory mediators, supports formation of more permissive environment for repair and regeneration

Characteristics of novel myeloid precursor cell line, PC-MDS, established from a bone marrow of the patient with therapy-related myelodysplastic syndrome

We report on characteristics of the first human cell line, PC-MDS, derived from a bone marrow of a patient with therapy-related myelodysplastic syndrome (t-MDS) who had no overt post-MDS leukemia. Classic cytology analyses, immunophenotyping, cytogenetic and molecular genetic procedures were used for characterization of the cell line. PC-MDS cells are positive for the expression of CD13, CD15, CD30, CD33, and CD45 antigen. Positive cytochemical staining and immunophenotype analyses indicated that PC-MDS cells have some characteristics of the early myeloid precursor cell. The karyotype analysis of PC-MDS cell line revealed various numerical and structural changes including those typically associated with t-MDS: del(5)(q13)[7], der(5)t(5;11)(p11;q11)[13], -7[6], del(7)(q31)[2], +20[3], -20[4]. Evaluation of methylation status in a promoter region of p 15, p 16 and MGMT genes showed biallelic hypermethylation pattern of 5 promoter region only in MGMT gene. PC-MDS is the first t-MDS derived cell line, and based on its immunological, cytogenetic and molecular characterization could be a new tool in evaluation of complex biology of MDS and a model for methylation studies. (c) 2007 Elsevier Ltd. All rights reserved