BACKGROUND AND OBJECTIVES: The aim of this study was to determine the effect of ethanol on the regulation of vascular tone. MATERIAL AND METHODS: Using rat aorta ring, isometric contraction and 45Ca uptake were measured. Phorbol 12,13-dibutyrate (PDBu), phenylephrine, KCl were used for the regulation of smooth muscle tone. RESULTS: Ethanol induced transient contraction in rat aorta ring by dose-dependent manner. Ethanol suppressed the dose dependent contractile responses of vascular strip by phenylephrine, KCl and PDBu. Endothelium-dependent relaxation by acetylcholine was inhibited by ethanol. Ethanol depressed 45Ca uptake by high KCl but not by phenylephrine or PDBu in rat aorta. n-butanol selectively suppressed tonic contraction by high KCl, but t-butanol did not at the same concentration of butanol in rat aorta. PDBu-induced contraction was selectively suppressed by n-butanol but not by t-butanol. CONCLUSIONS: These findings suggest that the action of ethanol on phospholipase D is involved in the decreased response of rat aorta strip by vasoconstrictors.
1-Butanol ; Acetylcholine ; Animals ; Aorta* ; Ethanol* ; Isometric Contraction ; Muscle, Smooth* ; Phenylephrine ; Phorbol 12,13-Dibutyrate ; Phospholipase D ; Protein Kinase C ; Rats* ; Relaxation ; tert-Butyl Alcohol ; Vasoconstrictor Agents

Aquaporins (AQPs) are expressed in myocardium and the implication of AQPs in myocardial water balance has been suggested. We investigated the expression patterns of AQP subtypes in normal myocardium and their changes in the process of edema formation and cardiac dysfunction following myocardial infarction (MI). Immunostaining demonstrated abundant expression of AQP1, AQP4, and AQP6 in normal mouse heart; AQP1 in blood vessels and cardiac myocytes, AQP4 exclusively on the intercalated discs between cardiac myocytes and AQP6 inside the myocytes. However, neither AQP7 nor AQP9 proteins were expressed in CD1 mouse myocardium. Echocardiography revealed that cardiac function was reduced at 1 week and recovered at 4 weeks after MI, whereas myocardial water content determined by wet-to-dry weight ratio increased at 1 week and rather reduced below the normal at 4 weeks. The expression of cardiac AQPs was up-regulated in MI-induced groups compared with sham-operated control group, but their time-dependent patterns were different. The time course of AQP4 expression coincided with that of myocardial edema and cardiac dysfunction following MI. However, expression of both AQP1 and AQP6 increased persistently up to 4 weeks. Our findings suggest a different role for cardiac AQPs in the formation and reabsorption of myocardial edema after MI.
Animals ; Aquaporin 1/metabolism ; Aquaporin 4/metabolism ; Aquaporin 6/metabolism ; Aquaporins/*metabolism ; Edema/pathology ; Immunohistochemistry ; Mice ; Muscle Cells/metabolism ; Myocardial Infarction/*metabolism/pathology/ultrasonography ; Myocardium/metabolism/pathology ; Time Factors

PURPOSE: To determine whether febrile seizure enhances neuroexcitability by altering synaptic transmission and whether febrile seizure-induced hyperexcitability leads to long-lasting neuronal death. METHODS: We investigated the expression of synaptic and postsynaptic proteins and the apoptosis of neuronal cells in rat pup hippocampus after hyperthermic seizure using immunoblotting and confocal microscopy. RESULTS: Hyperthermic seizure enhanced the long-term expressions of presynaptic proteins such as syntaxin, VAMP, SNAP-25 and nSec1, whereas that of NSF was decreased. The expressions of postsynaptic NMDA receptors 1, 2a and 2b were up-regulated. The expression of postsynaptic AMPA glutamate receptors 1 month after hyperthermic seizures altered by way of increasing the ratio of GluR1 to GluR2 and decreasing NSF-GluR2 interaction, which leads to the formation of Ca2+permeable AMPA receptors and enhanced toxicity. However, in spite of enhanced neuroexcitability, there was a transient increase of neuronal death in hipocampus one week after hyperthermic seizure, but returned to baseline one month later. CONCLUSION: These results demonstrate both presynaptic and postsynaptic forms of long-term enhancement of glutamate synaptic transmission after hyperthermic seizure and support the idea that early-life febrile seizure might have persistent effects on neuronal excitability in the hippocampus.
Rats ; Animals

Aging is associated with altered immune responses including dysregulation of cytokine production. Of cytokines, interleukin-1 (IL-1) family has been primarily involved with central nervous system. To evaluate the age-related different response of IL-1 family following peripheral administration of lipopolysaccharide (LPS), immunohistochemical study of IL-1beta and IL-1 receptor expression was performed on Sprague-Dawley rat brain. Experimental animals were divided into four groups; saline-treated young (3-5 months) and old (over 24 months), and LPS-treated young and old groups. After intraperitoneal (i.p.) injection of LPS, three to five rats within each group were killed at 1, 2, 4, 8 and 16 hr. After fixation in 4% neutral buffered formalin, the brain slices were paraffin-embedded. Immunohistochemical staining using labelled streptavidin biotin was performed. The results showed that IL-1beta immunoreactivity was seen in the endothelial cell of pons in both LPS-reated young and old rats, with slightly longer persistency in old group. IL-1RI immunoreactivity appeared initially in the neurons of cerebral cortex in LPS-treated old group, compared with predominantly the cerebellum in LPS-treated young group. In conclusion, our study shows that there is age-related, different neuronal localization of IL-1RI expression at different points of time after LPS treatment.
Age Factors ; Animal ; Brain Chemistry/drug effects* ; Gene Expression Regulation/drug effects ; Immunohistochemistry ; Interleukin-1/genetics ; Interleukin-1/analysis* ; Lipopolysaccharides/toxicity* ; Male ; Rats ; Rats, Sprague-Dawley ; Receptors, Interleukin-1/analysis

PURPOSE:This study was performed to elucidate that status epilepticus (SE) induces long- term neuronal damages in an immature brain and to evaluate that topiramate (TPM) has a protective effect. METHODS:We investigated the changes in a subtype expression of glutamate and gamma- amino butyric acid (GABA) receptors, and the structural integrity due to cell losses in the mouse pup hippocampus after SE using an immunoblot and confocal microscopy. RESULTS:SE induced significant cell losses with structural changes in the hippocampus 1 month later. SE up-regulated the glutamate receptor1 (GluR1) expression with an increased ratio of GluR1 to glutamate recptor2 (GluR2), leading to the formation of Ca2+ permeable alpha- amino-3-hydroxy-5-methyl-4-isoxazoleepropionic acid (AMPA) receptors for the enhanced neurotoxicity. TPM prevented the SE-induced GluR1 expression. The expression of GABAA receptors was highly increased 1 month after SE, whereas that of GABAB receptors was not changed. The TPM treatment attenuated SE-induced upregulation of GABAA receptors. SE induced significant cell losses and disruption of structural integrity in the hippocampus CA1 and CA3 regions, but the TPM treatment for 1 month in developing brains reduced the SE- induced hippocampal damage. CONCLUSION:TPM has a neuroprotective action, which might be mediated by the modulation of GluR1 and GABAA receptors.
Animals ; Brain* ; Butyric Acid ; Glutamic Acid ; Hippocampus ; Mice* ; Microscopy, Confocal ; Neurons ; Receptors, GABA ; Status Epilepticus ; Up-Regulation

BACKGROUND: Hepatocyte growth factor (HGF) is a multifunctional protein especially implicated in tissue repair and regeneration. In a murine model of chronic glomerulonepritis, subcutaneous injection of recombinant HGF inhibited renal fibrosis. Although these favorable results were accompanied by decreased expression of TGF-beta in renal tissue, the action of HGF on TGF-beta remains unclear. We studied whether HGF has an inhibitory effect on TGF-beta production in vitro. METHODS: Mouse mesangial cells grown in 5 mM glucose containing DMEM (NG, normal glucose) were exposed with 25 mM glucose medium (HG, high glucose) alone or in the presence of recombinant HGF (10 ng/mL) for 48 hours. TGF-beta in culture supernatants were measured by sandwich enzyme-linked immunosorbent assay. Secretion of fibronectin and collagen Ia was measured by western blotting and immunoprecipitaiton. TGF-beta, fibronectin and collagen Ia mRNA levels were measured by real-time PCR and internally corrected with GAPDH. RESULTS: High glucose significantly increased TGF-beta secretion (p<0.05), but HGF could not prevent high glucose-induced TGF-beta release. High glucose increased the secretion of both fibronectin and collagen Ia, which was blocked by HGF (p<0.05). CONCLUSION: In cultured murine mesangial cells HGF did not have effect on TGF-beta production, but inhibited the secretion of both fibronectin and collagen Ia. These results suggest that HGF may have indirect inhibitory effect on the action of TGF-beta.
Animals ; Blotting, Western ; Collagen* ; Enzyme-Linked Immunosorbent Assay ; Fibronectins* ; Fibrosis ; Glucose ; Hepatocyte Growth Factor ; Injections, Subcutaneous ; Mesangial Cells* ; Mice ; Real-Time Polymerase Chain Reaction ; Regeneration ; RNA, Messenger ; Transforming Growth Factor beta* ; Transforming Growth Factors

OBJECTIVE: To elucidate the role of aquaporin-4(AQP4) in cerebral edema formation, we studied the expression and subcellular localization of AQP4 in astrocytes after focal cerebral ischemia. METHODS: Cerebral ischemia were induced by permanent middle cerebral artery(MCA) occlusion in rats and estimated by the discoloration after triphenyltetrazolium chloride(TTC) immersion. Change of AQP4 expression were evaluated using western blot. Localization of AQP4 was assessed by confocal microscopy and its interaction with alpha-syntrophin was analyzed by immunoprecipitation. RESULTS: After right MCA occlusion, the size of infarct and number of apoptotic cells increased with time. The ratio of GluR1/GluR2 expression also increased during ischemia. The polarized localization of AQP4 in the endfeet of astrocytes contacting with ventricles, vessels and pia mater was changed into the diffuse distribution in cytoplasm. The interactions of AQP4 and Kir with alpha-syntrophin, an adaptor of dystrophin complex, were disrupted by cerebral ischemia. CONCLUSION: The deranged spatial buffering function of astrocytes due to mislocalized AQP4/Kir4.1 channel as well as increased assembly of Ca2+ permeable AMPA receptors might contribute to the development of edema formation and the excitotoxic neuronal cell death during ischemia.
Animals ; Apoptosis ; Aquaporin 4 ; Astrocytes ; Blotting, Western ; Brain Edema* ; Brain Ischemia* ; Cell Death ; Cerebral Infarction ; Cytoplasm ; Dystrophin ; Edema ; Immersion ; Immunoprecipitation ; Ischemia ; Microscopy, Confocal ; Neurons ; Pia Mater ; Rats* ; Receptors, AMPA ; Receptors, KIR

Cardiovascular mortality is associated with vascular calcification (VC) in hemodialysis (HD) patients. The present study was designed to find factors related with medial artery calcification on the plain radiography of feet by comparing C-reactive protein (CRP), plasminogen activator inhibitor type 1 (PAI-1) and lipid profile including oxidized low density lipoprotein (ox-LDL) and to elucidate associations among these factors in HD patients. Forty-eight HD patients were recruited for this study. VC in the feet was detected in 18 patients (37.5%) among total patients and 12 patients (85.7%) among diabetic patients. Diabetes, cardiovascular disease (CVD), pulse pressure, ox-LDL/LDL were higher and high density lipoprotein (HDL) was lower in patients with VC than in patients without VC. Negative associations were found between HDL and CRP, PAI-1. PAI-1 had positive association with ox-LDL/LDL. History of CVD was the only determinant of vascular calcification on the plain radiography of feet. Ox-LDL/LDL, HDL, CRP, and PAI-1 were closely related with one another in HD patients. History of CVD is the most important factor associated with the presence of VC and low HDL and relatively high oxidized LDL/LDL ratio may affect VC formation on the plain radiography in the feet of HD patients.
Aged ; C-Reactive Protein/metabolism ; Cardiovascular Diseases/blood/complications/diagnosis ; Female ; Foot ; Humans ; Kidney Failure, Chronic/blood/complications/diagnosis ; Lipoproteins, HDL/*metabolism ; Lipoproteins, LDL/*metabolism ; Male ; Middle Aged ; Plasminogen Activator Inhibitor 1/metabolism ; *Renal Dialysis ; Risk Factors

OBJECTIVE: Activated endothelial cells mediate the cascade of reactions in response to hypoxia for adaptation to the stress. It has been suggested that hypoxia, by itself, without reperfusion, can activate the endothelial cells and initiate complex responses. In this study, we investigated whether hypoxia-induced endothelial products alter the endothelial permeability and have a direct cytotoxic effect on nerve cells. METHODS: Hypoxic condition of primary human umbilical vein endothelial cells(HUVEC) was induced by CoCl2 treatment in culture medium. Cell growth was evaluated by 3,4,5-dimethyl thiazole-3,5-diphenyl tetrazolium bromide (MTT) assay. Hypoxia-induced products (IL-1beta, TGF-beta1, IFN-gamma, TNF-alpha, IL-10, IL-6, IL-8, MCP-1 and VEGF) were assessed by enzyme-linked immunosorbent assay. Endothelial permeability was evaluated by Western blotting. RESULTS: Prolonged hypoxia caused endothelial cells to secrete IL-6, IL-8, MCP-1 and VEGF. However, the levels of IL-1, IL-10, TNF-alpha, TGF-beta, IFN-gamma and nitric oxide remained unchanged over 48 h hypoxia. Hypoxic exposure to endothelial cells induced the time-dependent down regulation of the expression of cadherin and catenin protein. The conditioned medium taken from hypoxic HUVECs had the cytotoxic effect selectively on neuroblastoma cells, but not on astroglioma cells. CONCLUSION: These results suggest the possibility that endothelial cell derived cytokines or other secreted products with the increased endothelial permeability might directly contribute to nerve cell injury followed by hypoxia.
Anoxia ; Astrocytoma ; Blotting, Western ; Culture Media, Conditioned ; Cytokines ; Down-Regulation ; Endothelial Cells* ; Enzyme-Linked Immunosorbent Assay ; Humans ; Interleukin-1 ; Interleukin-10 ; Interleukin-6 ; Interleukin-8 ; Neuroblastoma ; Neurons* ; Nitric Oxide ; Permeability ; Reperfusion ; Stroke* ; Transforming Growth Factor beta ; Transforming Growth Factor beta1 ; Tumor Necrosis Factor-alpha ; Umbilical Veins ; Vascular Endothelial Growth Factor A

In this study, the effects of verapamil on growth rate, apoptosis, production of transforming growth factor (TGF-beta) and fibronectin were evaluated in keloid and normal human dermal fibroblasts. Both fibroblasts were primarily cultured from earlobe keloids of three female patients and treated with various concentrations of verapamil. Cell toxicity was assessed by MTT assay, growth rate and apoptosis by FACS, and the production of TGF-beta and fibronectin by ELISA and Western blot, respectively. In the MTT50, the cell growth was more suppressed in keloid fibroblasts. In the MTT90, cell growth was more stimulated in normal fibroblasts. No significant effect appeared on TGF-beta expression but an increase in extracellular fibronectin secretion was found in keloid fibroblasts. Keloid fibroblasts responded to verapamil more sensitively, and the percentage of apoptosis was higher at the MTT50l. In brief, verapamil had growth-inhibitory effect with inducing apoptosis at the MTT50, but rather growth-stimulatory effect at the MTT90. The biphasic effect of verapamil depending on the dose might explain one of the reasons of relapse after keloid treatment with verapamil. Clinical application with high concentration (2.5mg/ml) is advised unless excessive dosage is used.
Apoptosis* ; Blotting, Western ; Enzyme-Linked Immunosorbent Assay ; Female ; Fibroblasts* ; Fibronectins ; Humans ; Keloid* ; Recurrence ; Transforming Growth Factor beta ; Transforming Growth Factors ; Verapamil*