AIMTo investigate the effect of ciliary neurotrophic factor (CNTF) on the nuclear translocation of protein kinase C (PKC) following NMDA administration in the primary cultured hippocampal neurons.

METHODS(1) PKCGAMMA or PKCepsilon- immunocytochemistry staining method was used after treating neurons with NMDA or CNTF. (2) The gray of the nucleus of the PKC-positive neurons were measured under the image pattern analysis system.

RESULTS(1) After NMDA administration of different concentration and time, Nucleus appear PKCgamma and PKCepsilon activities, especially the 100 micromol/L NMDA 30 min group. (2) The gray of nucleus in CNTF + 500 micromol/L NMDA group is similar to control group.

CONCLUSIONNMDA can induce nuclear translocation of PKC in the primary cultured hippocampal neurons, and CNTF can inhibit the translocation. It suggests that the inhibition of PKC translocation induced by NMDA is one of the important reasons for the neuro-protective effects of CNTF.

Animals ; Cells, Cultured ; Ciliary Neurotrophic Factor ; pharmacology ; Hippocampus ; cytology ; N-Methylaspartate ; pharmacology ; Neurons ; drug effects ; metabolism ; Protein Kinase C ; metabolism ; Protein Transport ; Rats ; Rats, Sprague-Dawley

BACKGROUNDCerebral hyperperfusion syndrome is an important complication of carotid endarterectomy (CEA). An >100% increase in middle cerebral artery velocity (MCAV) after CEA is used to predict the cerebral hyperperfusion syndrome (CHS) development, but the accuracy is limited. The increase in blood pressure (BP) after surgery is a risk factor of CHS, but no study uses it to predict CHS. This study was to create a more precise parameter for prediction of CHS by combined the increase of MCAV and BP after CEA.

METHODSSystolic MCAV measured by transcranial Doppler and systematic BP were recorded preoperatively; 30 min postoperatively. The new parameter velocity BP index (VBI) was calculated from the postoperative increase ratios of MCAV and BP. The prediction powers of VBI and the increase ratio of MCAV (velocity ratio [VR]) were compared for predicting CHS occurrence.

RESULTSTotally, 6/185 cases suffered CHS. The best-fit cut-off point of 2.0 for VBI was identified, which had 83.3% sensitivity, 98.3% specificity, 62.5% positive predictive value and 99.4% negative predictive value for CHS development. This result is significantly better than VR (33.3%, 97.2%, 28.6% and 97.8%). The area under the curve (AUC) of receiver operating characteristic: AUC(VBI) = 0.981, 95% confidence interval [CI] 0.949-0.995; AUC(VR) = 0.935, 95% CI 0.890-0.966, P = 0.02.

CONCLUSIONSThe new parameter VBI can more accurately predict patients at risk of CHS after CEA. This observation needs to be validated by larger studies.

Aged ; Blood Pressure ; physiology ; Cerebrovascular Circulation ; physiology ; Cerebrovascular Disorders ; physiopathology ; Endarterectomy, Carotid ; Female ; Hemodynamics ; physiology ; Humans ; Male ; Middle Aged ; Prospective Studies

This study aimed to investigate the effects of concentration, intestinal section and borneol on the intestinal absorption of salvianolic acids. The experiment not only studied the intestinal absorption properties of three concentrations of rosmarinic acid, salvianolic acid B and salvianolic acid A at duodenum, jejunum and ileum, but also of salvianolic acids compatible with borneol at different concentrations using single-pass intestinal perfusion model in rat with phenol red as the marker. The results showed that salvianolic acids was stable under weak-acid condition and affected by metabolism enzyme; The Peff and Ka significantly different among three concentrations of rosmarinic acid and salvianolic acid B, whose intestinal absorption were saturated in high concentration, suggesting that the transport mechanisms of rosmarinic acid and salvianolic acid B were similar to active transport or facilitated diffusion; However, there was inconspicuousness in the Peff and Ka of salvianolic acid A at different concentrations, whose absorption was not saturated in high concentration, indicating that the transport mechanisms of salvianolic acid A was passive diffusion; The Peff and Ka in the ileum obviously higher than those in the duodenum and jejunum, namely the ileum was the best absorption section; When concentration of borneol increased, the enhancing effect of intestinal absorption of salvianolic acids increased, but significantly decreased when borneol increased to some degree. The enhancing effect of medium borneol concentration was the optimum. This implied that borneol can enhance the intestinal absorption of salvianolic acids, and the capacity of enhancing effect was influenced by the concentration of borneol.
Animals ; Benzofurans ; isolation & purification ; pharmacokinetics ; Bornanes ; administration & dosage ; pharmacokinetics ; pharmacology ; Caffeic Acids ; isolation & purification ; pharmacokinetics ; Cinnamates ; isolation & purification ; pharmacokinetics ; Depsides ; isolation & purification ; pharmacokinetics ; Dose-Response Relationship, Drug ; Duodenum ; metabolism ; Ileum ; metabolism ; Intestinal Absorption ; Jejunum ; metabolism ; Lactates ; isolation & purification ; pharmacokinetics ; Male ; Perfusion ; methods ; Plants, Medicinal ; chemistry ; Rats ; Rats, Sprague-Dawley ; Salvia miltiorrhiza ; chemistry

OBJECTIVETo investigate the cell viabilities of vascular smooth muscle cells and vascular endothelial cells stimulated by cigarette smoke extract(CSE) .

METHODSThe CSE was prepared by smoke-bubbled phosphate buffered saline(PBS) generation.After culturing cells with different concentrations of CSE, we used the cell counting kit-8 to determine the cell viability.The expression levels of c-jun and cyclinD1 were analyzed through Western blot.The c-jun plasmid was transfected to detect the change of cyclinD1 expression.

RESULTSThe smooth muscle cell viability increased when the CSE concentration ranged 0.625%-10%, whereas the endothelial cells viability decreased when exposed to the CSE concentration. After exposure to CSE for 48 hours, there was no difference in c-jun expression between toxin group and PBS group;however, the expression of p-c-jun in the smooth muscle cells significantly increased in the toxin groups than in the PBS group(P<0.05) and the expression of p-c-jun in the vascular endothelial cells significantly decreased(P<0.05) . The level of cyclinD1 significantly increased after exposed to CSE, and its expression level also increased in respond to the c-jun overexpression.

CONCLUSIONCSE can enhance the proliferation of vascular smooth muscle cells and decrease in the activity of endothelial cells proliferation, which may be explained by the phosphorylation of c-jun and the expression of cyclinD1.

Cell Proliferation ; drug effects ; Cell Survival ; Cells, Cultured ; Cyclin D1 ; metabolism ; Endothelial Cells ; drug effects ; metabolism ; Humans ; Myocytes, Smooth Muscle ; drug effects ; metabolism ; Proto-Oncogene Proteins c-jun ; metabolism ; Tobacco ; adverse effects

Carotid endarterectomy(CEA)has been proved to be an effective surgery to treat the cerebral ischemia caused by carotid atherosclerotic stenosis. However,there is still no effective mean for the early diagnosis of the CEA-related severe complications such as stroke and death. Many studies have explored the potential biomarkers for stroke alert,although there is still a long way to go for their actual application in clinical settings. The carotid atherosclerotic stenosis,the perioperative complications of CEA,and the stroke share similar pathogenic mechanisms,and some biomarkers such as S100B,matrix metalloproteinase 9,asymmetric dimethylarginine,and neuron-specific enolase have been studied in the clinical trails of CEA. This article summarizes recent advances in this field.
Biomarkers ; metabolism ; Endarterectomy, Carotid ; Humans ; Intraoperative Period ; Postoperative Complications ; prevention & control ; Risk Assessment ; S100 Calcium Binding Protein beta Subunit ; metabolism

OBJECTIVETo investigate the changes of open probability (Po) of large conductance Ca(2+)-activated K(+) channel (BK channel) in diabetic coronary smooth muscle cells and elucidate the underlying cellular electrophysiology mechanisms of coronary dysfunction.

METHODSRat coronary smooth muscle cells were isolated from control group and diabetic group. BK single channel currents were recorded by patch clamp technique in inside-out configuration. Open probabilities were calculated and compared between two groups. After exposure to DHS-1, a specific BK channel activator, Po at 0.2 and 1 µmol/L free Ca(2+) were compared between control and diabetic groups.

RESULTSIn the presence of 0.2 µmol/L free Ca(2+), the Po at baseline was significantly lower in diabetic rats than in control rats (0.0032 ± 0.0012 vs. 0.095 ± 0.036, P < 0.05). Cytoplasmic application of DSH-1 significantly increased the Po to 0.335 ± 0.096 (P < 0.05 vs. baseline) in control rats, whereas DSH-1 had no effect in diabetic rats (Po = 0.022 ± 0.018, P > 0.05 vs. baseline). In the presence of 1 µmol/L free Ca(2+), the Po at baseline was also significantly lower in diabetic rats than in control rats (0.210 ± 0.055 vs. 0.458 ± 0.077, P < 0.05). Cytoplasmic application of DHS-1 further robustly enhanced Po to 0.823 ± 0.019 (P < 0.05 vs. baseline) in control rats and to 0.446 ± 0.098 in diabetic rats (P < 0.05 vs. baseline of diabetic rats; P < 0.05 vs. control rats with DHS-1).

CONCLUSIONThe decrease of Po of BK single channel in coronary smooth muscle cells may be a potential cause for coronary dysfunction in diabetic rats.

Animals ; Coronary Vessels ; metabolism ; physiopathology ; Diabetes Mellitus, Experimental ; metabolism ; physiopathology ; Large-Conductance Calcium-Activated Potassium Channels ; metabolism ; Male ; Muscle, Smooth, Vascular ; cytology ; physiopathology ; Myocytes, Smooth Muscle ; metabolism ; Patch-Clamp Techniques ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo analyze retrospectively the quantity and activation status of the tumor infiltrating cytotoxic lymphocytes in breast cancer and the draining lymph nodes, and its relation to the clinical pathological significance.

METHODSSeventy-four breast cancer samples with their corresponding axillary lymph nodes were histologically typed and staged. Cytotxic lymphocytes were analyzed by immunohistochemistry with the monoclonal antibodies against CD8, CD56, granzyme B and perforin.

RESULTSThe number of infiltrating CD8(+) T cells in the cancerous interstitial tissue were much higher than that in the tumor parenchyma. Compared with the metastatic tumor samples, the CD8(+) T cells were more intensive in the primary tumors (35.7 +/- 16.0 vs. 23.7 +/- 9.6). The tumor infiltrating CD8(+) T cells of patients with 5 years survivals were more than that of the dead cases in this follow-up series death (32.9 +/- 14.1 vs. 20.1 +/- 9.9). There was no significant difference of activated tumor infiltrating cytotoxic T cell analyzed by using the activation marker granzyme B(+) and there was also no significant correlation between the intensity of CD8(+), CD56(+) cells and the clinicopathological stages. However, percentages of the activated cytotoxic lymphocytes in Stage I groups were significantly higher than those in stage III and IV. Moreover, the number of perforin(+) cells was significantly less than that of granzyme B(+) cells, particularly in the cancerous tissue, indicating a dysfunctional status of tumor infiltrating cytotoxic lymphocytes.

CONCLUSIONSActivated cytotoxic lymphocytes may play a significant role against the tumor progression and is associated with a favorable prognosis to some extent. However, a putative dysfunctional status of cytotoxic lymphocytes at tumor site may compromise the host immunity against cancer.

Adult ; Aged ; Axilla ; Breast Neoplasms ; metabolism ; pathology ; CD56 Antigen ; metabolism ; CD8 Antigens ; metabolism ; Female ; Follow-Up Studies ; Granzymes ; metabolism ; Humans ; Immunohistochemistry ; Lymph Nodes ; metabolism ; pathology ; Lymphatic Metastasis ; Lymphocytes, Tumor-Infiltrating ; metabolism ; pathology ; Middle Aged ; Neoplasm Staging ; Perforin ; metabolism ; Retrospective Studies ; Survival Rate ; T-Lymphocytes, Cytotoxic ; metabolism ; pathology

OBJECTIVETo investigate in vivo inhibitory effect of histone deacetylase (HDAC) inhibitor valproic acid (VPA) on xenografted Kasumi-1 tumor in nude mice and its mechanism.

METHODSXenografted Kasumi-1 tumor mouse model was established by subcutaneous inoculation of Kasumi-1 cells. Xenotransplanted nude mice were assigned into control or VPA treatment groups. Volume of the xenografted tumors was measured and compared between the two groups. Terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling (TUNEL) was applied to detection of tumor cell apoptosis. The gene expression of GM-CSF, HDAC1, Ac-H3 and survivin was studied with semi-quantitative RT-PCR and Western blotting. ChIP method was used to assay the effects of VPA on acetylation of histone H3 within GM-CSF promoter region.

RESULTS(1) VAP significantly inhibited xenografted Kasumi-1 tumor growth. The calculated inhibition rate was 57.25%. (2) Morphologic study showed that VPA induced differentiation and apoptosis of Kasumi-1 tumor cells. The apoptosis index of VAP treatment group [(3.661 +/- 0.768)%] was significantly higher than that of control group [(0.267 +/- 0.110)%]. (3) Comparing to those in control group, the level of nuclear HDAC1 protein was significantly decreased, the Ac-H3 protein expression level was increased, the mRNA and protein expression levels of GM-CSF and acetylation of histone H3 were remarkably increased, and the gene expression level of survivin significantly decreased in VPA treatment group.

CONCLUSIONVAP significantly inhibits xenografted Kasumi-1 tumor growth and induces tumor cell differentiation and apoptosis. The mechanism may be decrease of survivin gene expression, inhibition of nuclear expression of HDAC, promotion of histone protein acetylation level and acetylation of histone H3 within GM-CSF promoter region, and increase of GM-CSF transcription.

Animals ; Apoptosis ; drug effects ; Cell Line, Tumor ; Histone Deacetylase Inhibitors ; pharmacology ; Humans ; Mice ; Mice, Nude ; Valproic Acid ; pharmacology ; Xenograft Model Antitumor Assays

OBJECTIVETo investigate the effects of two histone deacetylase (HDAC) inhibitors, valproic acid (VPA) and TSA, on the expression of vascular endothelial growth factor (VEGF) and its receptor KDR of the leukemia cell line Kasumi-1 cells, and to explore their potential mechanism in leukemia angiogenesis.

METHODKasumi-1 cells were treated with VPA and TSA at different concentrations for 3 days. The mRNA and protein expression levels of VEGF and KDR were determined by semi-quantitative RT-PCR and Western blot, and the bFGF mRNA by semi-quantitative RT-PCR.

RESULTSAs compared with that of control groups, VPA at 3 mmol/L downregulated the VEGF mRNA expression level for VEGF(121) from 0.632 ± 0.014 to 0.034 ± 0.004 and for VEGF(165) from 0.526 ± 0.021 to 0.015 ± 0.001, for KDR mRNA from 0.258 ± 0.034 to 0.038 ± 0.000, and for bFGF mRNA from 0.228 ± 0.017 to 0.086 ± 0.015. TSA downregulated the VEGF mRNA and KDR mRNA at concentration of 100 nmol/L, but its effect on bFGF mRNA only at higher concentration.

CONCLUSIONHDAC inhibitors might inhibit the leukemia angiogenesis by regulating the expression of VEGF and its recptor.

Angiogenesis Inducing Agents ; Cell Line ; Histone Deacetylase Inhibitors ; pharmacology ; Humans ; RNA, Messenger ; genetics ; Valproic Acid ; pharmacology ; Vascular Endothelial Growth Factor A

OBJECTIVETo conduct a proteomic analysis of human colorectal carcinoma cell lines LS174T and SW480 by two-dimensional electrophoresis (2-DE) and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS).

METHODSThe total proteins of human colorectal carcinoma cell lines LS174T and SW480 were separated with 2-DE using immobilized pH gradient strips and analyzed by MALDI-TOF-MS to obtain peptide mass fingerprints (PMFs). Proteins were identified by using Mascot software to search protein databases.

RESULTSGood resolution 2-DE maps were obtained. Some proteins including immunoglobulin heavy chain variable region (IgVH) and co-stimulatory molecule B7-1 were identified. IgVH and B7-1 were confirmed by electrospray ionization tandem spectrometry (ESI-MS/MS) and immunocytochemistry.

CONCLUSIONThere are IgVH and B7-1 expressions in human colorectal carcinoma cell lines LS174T and SW480. Results obtained will help to elucidate the mechanisms of tumor immune escape.

B7-1 Antigen ; biosynthesis ; genetics ; Cell Line, Transformed ; Colorectal Neoplasms ; genetics ; immunology ; metabolism ; Humans ; Immunoglobulin Heavy Chains ; biosynthesis ; genetics ; Immunoglobulin Variable Region ; biosynthesis ; genetics ; Neoplasm Proteins ; biosynthesis ; genetics ; immunology ; Peptide Mapping ; Proteome ; isolation & purification