AP124F Sigma-AldrichGoat Anti-Mouse IgG Antibody, (H+L) FITC Conjugated

Predegeneration is a standard technique to obtain mitotically activated and enriched cultures of Schwann cells (SCs). This study, for the first time, evaluated the impact of various duration of predegeneration on cell yield and enrichment of SCs from dog peripheral nerve.Dog sural nerves were subjected to 5, 10, 15 day-long in vitro predegeneration. The total cell yield and the purity of SCs were evaluated in each group on the first and seventh day after plating.The maximum and minimum numbers of cells were counted in 15 day-long predegene-ration and control groups which underwent no predegeneration. The 10 day-long in vitro predegeneration group with 80±0.5% SCs enrichment had the best purity after plating day and could maintain its purity with elapsing on cultures.10 day-long predegeneration results in the higher cell number and the better and prolonged purity of SCs in culture.

Neuropathic pain is a very troublesome and difficult pain to treat. Although opioids are the best analgesics for cancer and surgical pain in clinic, only oxycodone among opioids shows better efficacy to alleviate neuropathic pain. However, many side effects associated with the use of oxycodone render the continued use of it in neuropathic pain treatment undesirable. Hence, we explored whether dextromethorphan (DM, a known N-methyl-D-aspartate receptor antagonist with neuroprotective properties) could potentiate the anti-allodynic effect of oxycodone and underlying mechanisms regarding to glial cells (astrocytes and microglia) activation and proinflammatory cytokines release in a spinal nerve injury (SNL) mice model.Oxycodone produced a dose-dependent anti-allodynic effect. Co-administration of DM at a dose of 10 mg/kg (i.p.) (DM10) which had no anti-allodynic effect by itself enhanced the acute oxycodone (1 mg/kg, s.c.) effect. When the chronic anti-allodynic effects were examined, co-administration of DM10 also significantly enhanced the oxycodone effect at 3 mg/kg. Furthermore, oxycodone decreased SNL-induced activation of glial cells (astrocytes and microglia) and plasma levels of proinflammatory cytokines (IL-6, IL-1β and TNF-α). Co-administration of DM10 potentiated these effects of oxycodone.The combined use of DM with oxycodone may have therapeutic potential for decreasing the effective dose of oxycodone on the treatment of neuropathic pain. Attenuation of the glial activation and proinflammatory cytokines in the spinal cord may be important mechanisms for these effects of DM.

Previous anti-inflammatory strategies against sepsis, a leading cause of death in hospitals, had limited efficacy in clinical trials, in part because they targeted single cytokines and the experimental models failed to mimic clinical settings. Neuronal networks represent physiological mechanisms, selected by evolution to control inflammation, that can be exploited for the treatment of inflammatory and infectious disorders. Here, we report that sciatic nerve activation with electroacupuncture controls systemic inflammation and rescues mice from polymicrobial peritonitis. Electroacupuncture at the sciatic nerve controls systemic inflammation by inducing vagal activation of aromatic L-amino acid decarboxylase, leading to the production of dopamine in the adrenal medulla. Experimental models with adrenolectomized mice mimic clinical adrenal insufficiency, increase the susceptibility to sepsis and prevent the anti-inflammatory effects of electroacupuncture. Dopamine inhibits cytokine production via dopamine type 1 (D1) receptors. D1 receptor agonists suppress systemic inflammation and rescue mice with adrenal insufficiency from polymicrobial peritonitis. Our results suggest a new anti-inflammatory mechanism mediated by the sciatic and vagus nerves that modulates the production of catecholamines in the adrenal glands. From a pharmacological perspective, the effects of selective dopamine agonists mimic the anti-inflammatory effects of electroacupuncture and can provide therapeutic advantages to control inflammation in infectious and inflammatory disorders.

Cellular behavior is dependent on a variety of physical cues required for normal tissue function. In order to mimic native tissue environments, human alveolar bone-derived mesenchymal stem cells (hABMSCs) were exposed to orbital shear stress (OSS) in a low-speed orbital shaker. The synergistic effects of OSS on proliferation and differentiation of hABMSCs were investigated. In particular, we induced the osteoblastic differentiation of hABMSCs cultured in the absence of OM by exposing hABMSCs to OSS (0.86-1.51 dyne/cm(2)). Activation of Cx43 was associated with exposure of hABMSCs to OSS. The viability of cells stimulated for 10, 30, 60, 120, and 180 min/day increased by approximately 10% compared with that of control. The OSS groups with stimulation of 10, 30, and 60 min/day had more intense mineralized nodules compared with the control group. In quantification of vascular endothelial growth factor (VEGF) and bone morphogenetic protein-2 (BMP-2) protein, VEGF protein levels under stimulation for 10, 60, and 180 min/day and BMP-2 levels under stimulation for 60, 120, and 180 min/day were significantly different compared with those of the control. In conclusion, the results indicated that exposing hABMSCs to OSS enhanced their differentiation and maturation.

Articular chondrocytes undergo an obvious phenotypic change when cultured in monolayers. During this change, or dedifferentiation, the expression of type I and type III procollagen is induced where normal chondrocytes express little type I and type III procollagen. In this study, we attempted to determine the mechanism(s) for the induction of such procollagen expression in dedifferentiating chondrocytes.All experiments were performed using primary-cultured human articular chondrocytes under approval of institutional review boards. Integrin(s) responsible for the induction of type I and type III procollagen expression were specified by RNAi experiments. The signal pathway(s) involved in the induction were determined by specific inhibitors and RNAi experiments. Adenovirus-mediated experiments were performed to identify a small GTPase regulating the activity of integrins in dedifferentiating chondrocytes. The effect of inhibition of integrins on dedifferentiation was investigated by experiments using echistatin, a potent disintegrin. The effect of echistatin was investigated first with monolayer-cultured chondrocytes, and then with pellet-cultured chondrocytes.In dedifferentiating chondrocytes, α5β1 integrin was found to be involved in the induction of type I and type III procollagen expression. The induction was known to be mediated by v-akt murine thymoma viral oncogene homolog (AKT) signaling. Among the three AKT isoforms, AKT1 seemed to be most involved in the signaling. Elated RAS viral (r-ras) oncogene homolog (RRAS) was considered to regulate the progression of dedifferentiation by modulating the affinity and avidity of α5β1 integrin to ligands. Echistatin inhibited dedifferentiation of monolayer-cultured chondrocytes. Furthermore, the matrix formed by pellet-cultured chondrocytes more closely resembled that of normal cartilage compared with the controls.The result of this study has shown, for the first time, that α5β1 integrin may be responsible for the induction of non-cartilaginous collagen expression in chondrocytes undergoing dedifferentiation. Again, this study has shown that the inhibition of ligand ligation to integrins may be an effective strategy to inhibit phenotypic change of cultured chondrocytes, and to improve the quality of matrix synthesized by primary cultured chondrocytes.

Based on knowledge from traditional herbal medicine, Ferula assa-foetida (asafoetida) has several therapeutic applications but there is less knowledge about its effect on neurons.In order to evaluate neuronal differentiation, neuronal like cells were stained against neuronal specific markers β-Tubulin III and MAP2. After establishment of neuronal differentiation in cultured cells, aqueous extract of gum-resin of asafoetida were applied on culture medium of neurons with different concentrations then survival rate of neurons were evaluated by cell counting and methyl tetrazolium bromide (MTT) tests.The results showed that asafoetida gum resin particularly with 0.01 and 1 µg/ml concentrations could improve survival rate of neurons, while10 µgr/ml treated group was toxic.RESULTS of this study indicated that gum resin of asafoetida in low doses has neuroprotective effect on neurons and improves survival rate of them, however in higher concentrations it is toxic for neurons.

Based on knowledge from traditional herbal medicine, Ferula assa-foetida (asafoetida) has several therapeutic applications but there is less knowledge about its effect on neurons.In order to evaluate neuronal differentiation, neuronal like cells were stained against neuronal specific markers β-Tubulin III and MAP2. After establishment of neuronal differentiation in cultured cells, aqueous extract of gum-resin of asafoetida were applied on culture medium of neurons with different concentrations then survival rate of neurons were evaluated by cell counting and methyl tetrazolium bromide (MTT) tests.The results showed that asafoetida gum resin particularly with 0.01 and 1 µg/ml concentrations could improve survival rate of neurons, while10 µgr/ml treated group was toxic.RESULTS of this study indicated that gum resin of asafoetida in low doses has neuroprotective effect on neurons and improves survival rate of them, however in higher concentrations it is toxic for neurons.

Retinal ischemia-associated ocular disorders, such as retinal occlusive disorders, neovascular age-related macular degeneration, proliferative diabetic retinopathy, and glaucoma are vision-threatening. In this study, we examined whether and by what mechanisms resveratrol, a polyphenol found in red wine, is able to protect against retinal ischemia/reperfusion injury.In vivo rat retinal ischemia was induced by high intraocular pressure (HIOP), namely, 120 mmHg for 60 min. The mechanism and management was evaluated by electroretinogram (ERG) b-wave amplitudes measurement, immunohistochemistry, and real-time polymerase chain reaction.The HIOP-induced retinal ischemic changes were characterized by a decrease in ERG b-wave amplitudes, a loss of choline acetyltransferase immunolabeling of amacrine cell bodies/neuronal processes, and increased vimentin immunoreactivity, which is a marker of Müller cells, together with upregulation of matrix metalloproteinase-9 (MMP-9), heme oxygenase-1 (HO-1), and inducible nitric oxide (iNOS), and downregulation of Thy-1, both at the mRNA level. The detrimental effects due to the ischemia were concentration-dependent (weaker effect at 0.05 nmole) and/or significantly (at 0.5 nmole) altered when resveratrol was applied 15 min before or after retina ischemia.This study supports the hypothesis that resveratrol may be able to protect the retina against ischemia by downregulation of MMP-9 and iNOS, and upregulation of HO-1.

This study was performed to investigate the effects of extremely low frequency pulsed electromagnetic fields (ELF-PEMFs) on the proliferation and differentiation of human alveolar bone-derived mesenchymal stem cells (hABMSCs). Osteogenesis is a complex series of events involving the differentiation of mesenchymal stem cells to generate new bone. In this study, we examined not merely the effect of ELF-PEMFs on cell proliferation, alkaline phosphatase (ALP) activity, and mineralization of the extracellular matrix but vinculin, vimentin, and calmodulin (CaM) expressions in hABMSCs during osteogenic differentiation. Exposure of hABMSCs to ELF-PEMFs increased proliferation by 15% compared to untreated cells at day 5. In addition, exposure to ELF-PEMFs significantly increased ALP expression during the early stages of osteogenesis and substantially enhanced mineralization near the midpoint of osteogenesis within 2 weeks. ELF-PEMFs also increased vinculin, vimentin, and CaM expressions, compared to control. In particular, CaM indicated that ELF-PEMFs significantly altered the expression of osteogenesis-related genes. The results indicated that ELF-PEMFs could enhance early cell proliferation in hABMSCs-mediated osteogenesis and accelerate the osteogenesis.

Diurnal temperature fluctuations can fundamentally alter mosquito biology and mosquito-virus interactions in ways that impact pathogen transmission. We investigated the effect of two daily fluctuating temperature profiles on Aedes aegypti vector competence for dengue virus (DENV) serotype-1. A large diurnal temperature range of 18.6°C around a 26°C mean, corresponding with the low DENV transmission season in northwestern Thailand, reduced midgut infection rates and tended to extend the virus extrinsic incubation period. Dissemination was first observed at day 7 under small fluctuations (7.6°C; corresponding with high DENV transmission) and constant control temperature, but not until Day 11 for the large diurnal temperature range. Results indicate that female Ae. aegypti in northwest Thailand are less likely to transmit DENV during the low than high transmission season because of reduced DENV susceptibility and extended virus extrinsic incubation period. Better understanding of DENV transmission dynamics will come with improved knowledge of temperature effects on mosquito-virus interactions.