In vitro anti-platelet aggregative effect of procyanidins isolated from grape seeds

AIM: To study the possible anti-platelet aggregative mechanisms of procyanidins (PC) isolated from grape seeds in vitro. METHODS: Platelet-rich plasma (PRP) and platelet-poor plasma (PPP) were prepared from the blood of healthy volunteers. PC,diphenylene iodonium(DPI,a nonspecific NADPH oxidase inhibitor) and apocynin (a specific NADPH oxidase inhibitor) were used to observe the effects on collagen-induced platelet maximum aggregation rate using platelet aggregometer. The influences of PC on platelet NADPH oxidase activity, NO content and superoxide anion (O₂^-·) level were evaluated by chemiluminescence spectrometer. The role of PC in the expression of activated platelet markers (PAC-1 and CD62P) was observed by flow cytometry. RESULTS: PC (100 μmol/L), apocynin (10 μmol/L) and DPI (100 μmol/L) significantly inhibited collagen-induced maximum platelet aggregation rate (P<0.01). In collagen-activated platelets, NO content reduced and O₂^-· level increased,both of which were recovered by PC at concentration of 100 μmol/L (P<0.05). PC also obviously inhibited NADPH oxidase activity (P<0.01), and significantly down-regulated PAC-1 and CD62P expression (P< 0.05) in platelets. CONCLUSION: Procyanidins isolated from grape seeds have the anti-platelet aggregation function through inhibiting NADPH oxidase activity, further influencing platelet NO and O₂^-· levels.     

Effects of hypertriglyceridemia and fenofibrate on CD40L expression in platelets

AIM: To observe the effects of hypertriglyceridemia and fenofibrate on CD40L expression in platelets in vitro and in vivo. METHODS: In vivo experiments, according to its own strict standards, 20 patients were respectively selected for hypertriglyceridemia group and control group, before and after treatment of fenofibrate for hypertriglyceridemia patients. The CD40 ligand positive rates of platelets by flow cytometry and plasma soluble CD40 ligand by ELISA were examined under the same conditions as control group. The CD40L and sCD40L in each group were compared. In in vitro experiments, all 6 objects plasma was chosen in the same condition except for triglyceridemia, after the co-incubation of these plasma with the same healthy platelets was performed and the interference with wy14643, the CD40 ligand positive rate of platelets by flow cytometry and total platelets CD40 ligand protein content by Western blotting were examined under the same conditions in all objects. The CD40L positive rate and total CD40L content in each group were compared, respectively. RESULTS: The platelet CD40L positive rate and plasma sCD40L concentration in hypertriglyceridemia group were significant higher than those in control group (P<0.01). Followed the TG concentration decreased, the platelet CD40L positive rate and plasma sCD40L concentration decreased after the treatment of fenofibrate, the same as the total platelets CD40L content which was significant higher in hypertriglyceridemia group than that in control group in vitro (P<0.05). No effect of wy14643 on the total CD40L content expression was observed in vitro. CONCLUSION: Hypertriglyceridemia plasma stimulates immune-activation of platelets both in vitro and in vivo. sCD40L may mainly come from CD40L on platelet membrane. PPARα activator of fenofibrate may inhibit the immune-activation of platelets by reducing the concentration of plasma TG, but PPARα activator WY14643 cant inhibit the expression of CD40L and CD40L in vitro.     

Establishment and assessment of a method for acid elution of platelet HLA class I antigens

AIM: To establish an effective method for acid elution of platelet HLA class I antigens, and to evaluate the optimal condition and the feasibility of clinical application of the acid-elution technique. METHODS: Platelets were treated with citric acid buffer at different pH levels (pH=2, 3, 5, 7). Expression of HLA class I antigens and P-selectin (CD62P) on the platelet surface was analyzed by multicolor flow cytometry. The proportion of early apoptotic platelets was detected by Annexin V staining. The maximum platelet aggregation rate was determined by electrical impedance aggregometry.RESULTS: With the decrease in the pH levels of the citric acid buffer (from pH=7 to pH=2), the expression of HLA class I antigens on the platelet surface was remarkably decreased. However, the rates of platelets activation (CD62P expression) and early apoptosis (Annexin V expression) were both significantly increased. Compared with PBS, treatment of the platelets with citric acid buffer at pH 3.0 remarkably reduced the expression of platelet HLA-class I antigens (P<0.05). Although the rates of the platelet activation and apoptosis were also significantly increased (P<0.05), the aggregation of platelet was not remarkably reduced (P>0.05).CONCLUSION: Acid elution of platelet HLA-class I antigens with citric acid buffer at pH 3.0 at 0 ℃ can be use as an attempt to produce HLA-eluted platelets. This technique of acid-elution needs further improvement and standardization before clinical use.     

Effects of IL-6 and IL-11 on differentiation of cord blood CD34+ cells towards megakaryocytes

AIM: To investigate the effects of interleukin-6 (IL-6) and interleukin-11 (IL-11) on differentiation of cord blood CD34+ cells towards megakaryocytes and platelet production in vitro.METHODS: The CD34+ cells from fresh umbilical cord blood samples were cultured in serum-free culture medium with thrombopoietin (TPO) 50 μg/L,IL-3 10 μg/L,stem cell factor (SCF) 50 μg/L as control groups,then 10 μg/L IL-6 or IL-11 or IL-6＋IL-11 respectively was added as treatment groups.Mononuclear cells (MNCs) in cultured cells were detected by cell counter,megakaryocytes (CD41+ cells) and platelets were measured by flow cytometry,respectively.Platelet agglutination after thrombin induced was observed by microscopy and flow cytometry.RESULTS: Compared with the control group,the number of MNCs was not significantly different（P>0.05）,but the numbers of CD41+ cells and platelets were increased significantly (P<0.05) in treatment groups.There were more platelet particles in treatment groups than those in control group by microscopy and the results also showed that the cytoplasmic fragments from the cultures responded to thrombin induction.CONCLUSION: It is concluded that both IL-6 and IL-11 induce the cord blood CD34+ cells to differentiate towards megakaryocytes and produce platelets.     

Antitumor effect of heat-killed lactobacilli adhered to cervical carcinoma cells

AIM: To observe the immune property of cervical carcinoma and antitumour effect of heat-killed lactobacilli adhered to HeLa cells. METHODS: Flow cytometry was used to analyze the expression of immune molecules on HeLa cells after adhered with heat-killed lactobacilli. MTT was used to analyze the cytotoxity of lymphocytes on cervical carcinoma cells. RESULTS: Adherence of heat-killed lactobacilli to HeLa cells did not affect the expression of MHC-I molecules (P>0.05), but enhanced the expression of CD80, CD86 significantly (P<0.05) in HeLa cells. Adherence of heat-killed lactobacilli to HeLa cells also enhanced the cytotoxity of NK cells. Compared to control, the cytotoxity of CTL to HeLa carcinoma cells was significantly enhanced by the adherence of heat-killed lactobacilli to HeLa cells (P<0.05). CONCLUSION: Heat-killed lactobacilli strengthen the antitumor effect on HeLa cells.     

Effects of different group B streptococci strains on platelet activation

AIM: To explore the ability of different group B streptococci (GBS) strains on inducing platelet activation. METHODS: Six strains of GBS, separated from the septic patients with thrombocytopenia, were used as the inducers. Light transmission aggregometry was used to measure platelet aggregation. Scanning electron microscopy (SEM) was performed to investigate the interaction of platelets with bacteria. The expression of platelet CD62P, Toll-like receptor 2 (TLR2) and TLR4 was determined by flow cytometry and Western blotting. Furthermore, the activity of platelet TLR2 (or TLR4) was blocked by anti-TLR2 (or anti-TLR4) monoclonal antibody, and the platelet aggregation induced by GBS was detected. RESULTS: Only 3 of 6 GBS strains isolated from the septic patients induced platelet aggregation and up-regulated the expression of CD62P and TLR2 in the platelets (P < 0.05), but not TLR4. Incubation with anti-TLR2 antibody, but not anti-TLR4 antibody, significantly blocked platelet aggregation induced by GBS.CONCLUSION: Some GBS strains from the patients are able to trigger platelet activation in vitro, and platelet TLR2 may play an important role in the interaction between GBS and platelets.     

P-selectin on activated platelets promotes hematogenous metastasis of primary lung cancer via P-selectin glycoprotein ligand-1

AIM: To investigate the relationship between platelets/P-selectin on activated platelets and clinico-pathological features, hematogenous metastasis and prognosis of lung cancer, and to explore the effect of P-selectin and P-selectin glycoprotein ligand-1 (PSGL-1) interaction on the hematogenous metastasis-related integrin β3 (ITGB3). METHODS: The expression of P-selectin on activated platelets was detected by flow cytometry, and its effects on lung cancer and the risk of hematogenous metastasis were analyzed. The expression of PSGL-1 in different types of lung cancer tissues and cell lines was determined by Western blot. By co-culturing platelets and lung cancer cells in vitro, the effects of up- and down-regulation of PSGL-1 on invasion and migration abilities of lung cancer cells were observed. RESULTS: The peripheral blood platelet counts and P-selectin expression on activated platelets in the patients with lung cancer were significantly increased (P<0.05). The P-selectin expression on activated platelets was significantly associated with hematogenous metastasis of lung cancer (r=0.256, P<0.05). The strongest expression of PSGL-1 was found in the lung adenocarcinoma samples, next in the lung squamous-cell carcinoma samples, and the weakest in small-cell lung cancer samples. P-selectin promoted transmembrane invasion of lung cancer cells. Inhibition of P-selectin and its ligand PSGL-1 reduced ITGB3 expression, invasion and migration of lung cancer cells. CONCLUSION: The P-selectin level on activated platelets is significantly associated with hematogenous metastasis of lung cancer, which is related to the binding of P-selectin and its ligand PSGL-1. Up-regulation of ITGB3 level after their binding might be one of the mechanisms of the remodeling of extracellular matrix to facilitate hematogenous metastasis of lung cancer.     

《园艺学报》征稿简则

AIM:To investigate the prognostic value of biomarkers on predicting recurrence of hepatocellular carcinoma (HCC) after orthotopic liver transplantation (OLT). METHODS:Fifty patients with HCC who underwent OLT between April 2002 and November 2005 with a minimum clinical follow up of 12 months were included in this retrospective study. We examined immunohistochemical expression of E-cadherin with β-catenin, Ki-67 proliferative index and analyzed the correlation between these biomarkers with recurrence and survival, along with the main clinical-pathological variables.RESULTS:Only at univariate analysis, TNM, portal vein tumor thrombi were valuable on predicting recurrence and survival time (P<0.05), and preoperative serum AFP correlate to recurrence only at multivariate analysis (OR=2.552, P<0.05). Lower membrane expression of E-cadherin, nuclear β-catenin localization and high Ki-67 index showed notable significance on predicting recurrence and survival time at univariate analysis, as well at multivariate analysis, all P<0.01. The membrane expression of β-catenin did not correlate to the prognosis (P>0.05).CONCLUSION:These three biomarkers might have potential as a tumor prognostic marker for predicting recurrence of HCC after OLT and perhaps are better than the clinical-pathological variables.     

Effect of proliferating cell nuclear antigen specific antisense oligonucleotide on differentiation of cord blood CD34+ cells

AIM:To study the effect of proliferating cell nucler antigen antisense oligonucleotide on ex vivo expansion of cord blood CD34+ hematopoietic stem/progenitor cells. METHODS:CD34+ cells were purified from fresh cord blood by immunomagnetic beads. CD34+ cells were incubated in liquid culture system with different concentrations of PCNA-ASODN. Using flow cytometry, the number of different kinds of stem/progenitor cells and PCNA expression were measured after CD34+ cell incubation. RESULTS:PCNA was lowly expressed in low experiential group, with a positive rate of (27.2±3.6)% and (19.0±1.5)%, the positive rate of control group was (53.8±8.3)% (P<0.01), a high significant difference was observed. Just in low concentration of PCNA-ASODN, the percentage of CD34+cells were increased to (33.4±3.2)%, CD34+CD38-cells expanded (57.8±9.9) folds, and the percentage of CD34+cells were (25.2±2.6)% (P<0.01), the CD34+CD38-cells expanded (43.5±7.4) folds (P<0.05), it has significant difference compared with the control group. CONCLUSION:Low concentration of PCNA-ASODN decreases PCNA expression effectively, slows down differentiation of CD34+cells during ex vivo expansion procedure, and improves the expansion efficiency.     

Effect of nitric oxide on adiponectin-induced inhibition of platelet aggregation in hyperlipidemia rats

AIM: To investigate the mechanism that adiponectin inhibits platelet aggregation via nitric oxide (NO) signaling pathway. METHODS: Adult rats were fed with normal or high-fat diet for 14 weeks. Their platelets were immediately isolated and treated with or without recombinant full-length adiponectin (rAPN). The platelet aggregation, NO and superoxide production, endothelial nitric oxide synthase (eNOS)/inducible NOS (iNOS) expression, and antioxidant capacity were determined. RESULTS: Treatment with rAPN inhibited platelet aggregation induced by hyperlipidemia (P<0.05). Interestingly, total NO, a crucial molecule depressing platelet aggregate and thrombus formation, was significantly reduced, rather than increased in rAPN-treated platelets. Treatment with rAPN significantly decreased superoxide production by 62% (P<0.05) and increased antioxidant capacity by 38% (P<0.05) in hyperlipidemic platelets. Importantly, hyperlipidemia-induced reduction of eNOS phosphorylation and increase in iNOS expression were markedly reversed by rAPN treatment (P<0.05 and P<0.01, respectively). CONCLUSION: Adiponectin is an adipokine that inhibits platelet aggregation by enhancing eNOS activation and attenuating oxidative/nitrative stress including blockage of iNOS expression and superoxide production.     

Clinical significance and prognostic value of β-catenin expression in colonic adenocarcinoma

AIM: To observe the expression of β-catenin in colonic adenocarcinoma and to investigate its clinical significance and prognostic value. METHODS: The integrated clinical and follow-up data of 52 patients, who were undergone radical operation, were retrospectively analyzed from June 2000 to June 2004 in our hospital. The paraffin-embedded tissues of 52 colonic adenocarcinoma specimens and 20 cases of normal paraneoplastic colonic mucous tissues were detected for β-catenin expression by immunohistochemical method. The relationships between β-catenin and clinical variables and prognostic value were statistically analyzed. RESULTS: β-catenin was normally expressed in all 20 normal cases. In the cases of colonic adenocarcinoma, the abnormal expression rate of β-catenin was 71%. The abnormal expression of β-catenin did not correlated with gender, age, histological differentiation and blood CEA (P>0.05), but it was correlated with lymph node metastasis and clinical stage (P<0.05). The abnormal expression of β-catenin were also correlated with the postoperative survive time (P<0.05). CONCLUSION: The abnormal expression of β-catenin is correlated with the lymph node metastasis and clinical stage, and the abnormal expression is an important adverse prognostic factor for survival in the patients with colonic adenocarcinoma. β-catenin may be a molecular prognostic marker in the patients with colonic adenocarcinoma.     

PP2A activation in mesenteric arteries of angiotensin Ⅱ-induced hypertensive rats

AIM: To investigate the mechanism of protein phosphatase 2A (PP2A) activation in mesenteric arteries of angiotensinⅡ(AngⅡ)-induced hypertensive rats. METHODS: Adult male Sprague-Dawley (SD) rats were subjected to AngⅡinfusion (500 ng·kg^-1·min^-1) using osmotic minipump up to 14 d to established the hypertension model. The rats (n=40) were randomly divided into 4 groups:control group (n=10), AngⅡgroup (n=10), candesartan (CAN; AngⅡtype 1 receptor blocker)+AngⅡgroup (n=10) and CAN group (n=10). The rats in CAN+AngⅡgroup and CAN group were administered with candesartan ester at the dose of 10 mg·kg^-1·d^-1 by gavage on the first day after implantation of osmotic minipump. The rats were sacrificed on the 15th day after minipump implantation. Serum and mesenteric arteries were collected. Systolic blood pressure was measured by tail-cuff method. The serum levels of AngⅡ were measured by ELISA. The protein levels of endothelial nitric oxide synthase (eNOS), phosphorylated eNOS (Ser1177), PP2A catalytic subunit (PP2Ac), phosphorylated PP2Ac (Tyr307) and PP2A inhibitor 2 (I₂^PP2A) in the mesenteric arteries were determined by Western blot. The activity of PP2A in the arteries was detected using PP2A activity assay kit. RESULTS: Compared with control group, the systolic blood pressure in AngⅡgroup was significantly increased(P<0.05), while those in CAN+AngⅡgroup and CAN group were significantly decreased (P<0.05). The serum levels of AngⅡ in AngⅡ group and CAN+AngⅡ group were significantly higher than that in control group (P<0.05). Compared with control group, the phosphorylation levels of eNOS Ser1177 were decreased in AngⅡgroup (P<0.05), but the activity of PP2A was significantly increased (P<0.05), and Pearson correlation analyses showed a negative correlation between PP2A activity and eNOS S1177 phosphorylation (r=-0.842, P<0.05). Compared with AngⅡgroup, the phosphorylation levels of eNOS Ser1177 in CAN+AngⅡgroup were significantly increased (P<0.05), but the activity of PP2A was reduced (P<0.05). Compared with control group, the protein levels of phosphorylated PP2Ac (Tyr307) and I₂^PP2A in the mesenteric arteries were decreased in AngⅡgroup (P<0.05), but increased in CAN+AngⅡgroup (P<0.05). No significant difference in all above-mentioned measures between control group and CAN group, nor in the levels of total eNOS and PP2Ac protein expression among all the groups was observed. CONCLUSION: AngⅡmay reduce the protein levels of phosphorylated PP2Ac (Tyr307) and I₂^PP2A in the mesenteric arteries of AngⅡ-induced hypertensive rats through AngⅡ/AngⅡ type 1 receptor-mediated signaling pathway, resulting in the activation of PP2A, then leading to down-regulation of eNOS S1177 phosphorylation, which ultimately mediates the occurrence of vascular endothelial dysfunction.     

Effect of progesterone on SH-SY5Y cells injured by ATP

AIM: To investigate the neuroprotective effect of progesterone against adenosine triphosphate (ATP)-injured human neuroblastoma SH-SY5Y cells.METHODS: The SH-SY5Y cells in the logarithmic phase were divided into different groups according to the progesterone and ATP concentrations. The cell viability was measured by CCK-8 assay. The membrane permeability was detected using fluorescent dye YO-PRO-1. Cytosolic Ca²⁺ concentration was measured with fluorescent dye Fluo-3/AM. The expression of purinergic P2X₇ receptor was assessed by Western blot.RESULTS: The viability of the SH-SY5Y cells was significantly decreased (P<0.05) and YO-PRO-1 uptake was obviously increased (P<0.05) in a concentration-dependent manner compared with control group when SH-SY5Y cells were treated with ATP at 1, 3, 5 and 7 mmol/L for 2 h. The viability reduction of the SH-SY5Y cells induced by ATP was obviously counteracted by treatment with progesterone at 3, 10 and 30 nmol/L for 30 min (P<0.05) as compared with ATP group. YO-PRO-1 fluorescence enhancement induced by ATP in SH-SY5Y cells was significantly reduced (P<0.05) by progesterone (30 nmol/L) or P2X₇ receptor antagonist KN-62 (500 nmol/L) pretreatment for 30 min, and no obvious difference between treatments with progesterone and KN-62 was observed. Cytosolic Ca²⁺ fluorescence intensity in normal group was a little, but that in ATP group was increased (P<0.05). Progesterone or KN-62 pretreatment significantly decreased the cytosolic fluorescence intensity of Ca²⁺ induced by ATP (P<0.05). However, no obvious difference between treatments with progesterone and KN-62 was found. The expression of P2X₇ receptor in ATP group was significantly higher than that in control group (P<0.05), and progesterone inhibited ATP-induced P2X₇ receptor expression (P<0.05).CONCLUSION: Progesterone inhibits P2X₇ receptor expression, membrane pore formation, intracellular Ca²⁺ increase and cell death induced by ATP, so progesterone may protect SH-SY5Y cells against ATP-induced injuries.     

Effect of fraction F of naja naja atra venom on protein tyrosine phosphorylation in platelet activation

AIM: To observe the mechanism of intracellular signal transduction that fraction F of naja naja atra venom inhibits platelet aggregation. METHODS: Tests were divided into six groups: (1) blank group; (2) control group and (3)-(6) ADP plus fraction F group (doses of fraction F were 100, 30, 10, 3 mg/L, respectively). Protein tyrosine phosphorylation in platelets was assayed by Western blotting and platelet aggregation was assayed by nephelometer. RESULTS: Fraction F significantly inhibited molecular masses (MW) 76, 66 and 37.5 kD protein tyrosine phosphorylation in platelet that induced by ADP in a dose-dependent manner, in which 30 and 100 mg/L dose group showed obviously different effects when compared to control group (P<0.05). Inhibition of platelet aggregation was positive correlated with protein tyrosine phosphorylation in platelets （r=0.9367, P<0.01）. CONCLUSIONS: Fraction F of naja naja atra venom affected intracellular signal transduction pathway in platelets by inhibiting MW 76, 66 and 37.5 kD protein tyrosine phosphorylation. The result suggests that this effect may be one of the anti-thrombus mechanisms of fraction F.     

Role of ERK5 in platelet activation in vitro and arterial thrombosis in vivo

AIM: To investigate the role of extracellular signal-regulated kinase 5 (ERK5) in platelet aggregation in vitro and arterial thrombosis in vivo. METHODS: The expression and phosphorylation levels of ERK5 in human platelet were detected by Western blot. The effects of ERK5 selective inhibitor XMD8-92 on platelet aggregation and dense granule secretion were detected by Chrono-Log aggregometer. The effect of ERK5 on in vivo thrombosis was analyzed using an FeCl₃ artery thrombosis model. The effects of XMD8-92 on protein kinase B (PKB/Akt) and phosphatase and tensin homolog deleted on chromosome ten (PTEN) phosphorylation levels were determined by Western blot. RESULTS: ERK5 was stably expressed in human platelets and its phosphorylation level increased significantly after platelet activation (P<0.05). XMD8-92, a selective inhibitor of ERK5, inhibited platelet aggregation and dense granule secretion in response to several platelet stimulators (P<0.05). The results of Western blot showed that XMD8-92 inhibited Akt phosphorylation level by down-regulating PTEN Ser370 phosphorylation and enhancing PTEN activity. The pathway was further confirmed using platelet specific PTEN deficiency mice. The first occlusion time was obviously extended in the mice intravenously given XMD8-92 in the FeCl₃-induced carotid artery injury model. CONCLUSION: ERK5 plays a role in platelet activation and arterial thrombosis by influencing PTEN and Akt phosphorylation.     

Effects of peripheral-type benzodiazepine receptors in platelet membrane on patients with depression after first attack of cerebral infarction

AIM：To evaluate the changes of peripheral-type benzodiazepine receptors (PBRs) in platelet membrane in post-stroke depression (PSD) patients and to investigate the effects of PBRs on PSD. METHODS：Forty-three patients with PSD, fifty-nine patients with first-ever cerebral infarction and fifty-six healthy volunteers participated in this study. Platelet membrane in venous blood was prepared. Binding assay of the radioactive PBRs antagonist ［3H］PK11195 to platelet membrane was performed. RESULTS：A significant difference of ［3H］PK11195 binding was found among the 3 groups (P<0.01). Compared with the healthy volunteers ［(298.2±25.1） pmol/(g protein)］, a highly significant increase in ［3H］PK11195 binding was observed in platelet membrane in the patients with first attack of cerebral infarction ［(1 410.8±41.4） pmol/(g protein), P<0.01］. Compared with the patients with first attack of cerebral infarction, a significant reduction in ［3H］PK11195 binding was detected in platelet membrane in the patients with PSD ［(361.7±30.6) pmol/(g protein), P<0.01］. In the patients with PSD, no significant difference of ［3H］PK11195 binding was found between men and women (P>0.05). ［3H］PK11195 binding was related to the score of Hamilton depression rating scale (r=-0.44, P<0.01) but not to the duration of cerebral infarction (r=0.27,P>0.05). CONCLUSION:PBRs binding activity in platelet membrane decreases in the patients with PSD and affects the degree of depression.     

Effect of E-selectin pretreatment on cerebral ischemia-reperfusion injury in rats

AIM:To study the effect of nasal mucosal tolerance to E-selectin on cerebral ischemia-reperfusion injury.METHODS:Two different doses (single and booster) of E-selectin or PBS were dropped into membrana mucosa nasi of rats. The middle cerebral artery occlusion (MCAO) model referring to Zea Longa method with modifications was performed 48 h after the last dose of E-selectin or PBS. After 2 h ischemia and 22 h reperfusion, the numbers of CD3+CD4+T-lymphocyte and CD3+CD8+T lymphocyte subgroup in the blood were examined with flow cytometry. Rats were killed, then part of the animals was used to measure the cerebral infarction volume by TTC staining. mRNA expressions of E-selectin, ICAM-1 and lymphocyte function-associated antigen-1(LFA-1) were determined by RT-PCR and activity of SOD was determined by xanthinoxidanse method in ischemic cortex of the other part of animals. RESULTS:The ratio of the numbers of CD3+CD4+T-lymphocytes and CD3+CD8+T-lymphocytes increased in E-selectin single pretreatment group (P<0.05). Compared to other groups, E-selectin booster pretreatment group showed decreased CD3+CD8+T-lymphocytes (P<0.05), increased ratio of the numbers of CD3+CD4+T-lymphocytes and CD3+CD8+T-lymphocytes (P<0.05), reduced cerebral infarction volume by 40.87% (P<0.05), heightened activity of SOD (P<0.05), lowed E-selectin mRNA and ICAM-1 mRNA expression (P<0.05), and less tendency of LFA-1 mRNA expression.CONCLUSION:E-selectin induces cerebral ischemic tolerance and relieves cerebral ischemia-reperfusion injury. The mechanisms are related to the changes in the ratio of CD4+T-lymphocyte and CD8+T-lymphocyte. The heightened activity of SOD, the lowed mRNA expressions of E-selectin and ICAM-1, as well as the less tendency of LFA-1 mRNA expression are also involved.     

Effect of berberine on the adhesion between human umbilical vein endothelial cells and human pulmonary carcinoma cells

AIM: To study the mechanism of berberine on the adhesion between human pulmonary carcinoma cells (PG cells) and HUVECs. METHODS: The effect of berberine (2.5-40 mg/L) on the proliferation of HUVECs was detected by MTT method. Further, PG cells were treated with berberine at doses of 2.5, 5, 10 mg/L for 6, 12, 24 h. The adhesion between PG cells and HUVECs was determined by rose bengal staining. The expression of CD44s on PG cells were determined by fluorescence antibody staining. Fluorescence anisotropy imaging system was used to assay the fluidity of PG cell membrane. RESULTS: Berberine at concentrations of 2.5, 5, 10 mg/L were the safety doses to the proliferation of HUVECs treated for 6, 12, 24 h. Berberine inhibited the adhesion between PG cells and HUVECs significantly in a dose-dependent manner (P<0.05 or P<0.01). Meanwhile, berberine increased the expression of CD44s on PG cells (P<0.05 or P<0.01). Berberine decreased the fluidity of PG cell membrane in a dose-dependent manner after 24 h incubation. CONCLUSION: Berberine inhibits the adhesion between PG cells and HUVECs by regulating the expression of adhesion molecules and the fluidity of cell membrane on PG cells.