Thromboxane A2 receptor contributes to the activation of rat pancreatic stellate cells induced by 8-epi-prostaglandin F2α

Background:Pancreatic stellate cells(PSCs)activation plays a critical role in the development of chronic pancreatitis.Previous studies confirmed that thromboxane A2 receptor(TxA2r)was overexpressed in activated PSCs in rats.The purpose of this study was to investigate the role of TxA2r in the activation of PSCs induced by 8-epi-prostaglandin F2α(8-epi-PGF2α).Methods:TxA2r expression in both quiescent and activated PSCs was detected by immunocytochemistry and immunoblot assay.Isolated PSCs were treated with 8-epi-PGF2α(10^-6,10^-7,10^-8 mol/L)for 48 h,and SQ29548(10^-4,10^-6,and 10^-7 mol/L),a TxA2r-specific antagonist,for 48 h,respectively,to identify the drug concentration with the best biological effect and the least cytotoxicity.Then isolated PSCs were treated with SQ29548(10^-4 mol/L)for 2 h,followed by 10^-7 mol/L 8-epi-PGF2α for 48 h.Real-time polymerase chain reaction was performed to detect the messenger RNA(mRNA)levels of a-smooth muscle actin(a-SMA)and collagen I.Comparisons between the groups were performed using Student’s t test.Results:TxA2r was up-regulated in activated PSCs in vitro compared with quiescent PSCs(all P<0.001).Compared with the control group,different concentrations of 8-epi-PGF2α significantly increased mRNA levels of a-SMA(10^-6 mol/L:2.23±0.18 vs.1.00±0.07,t=10.70,P<0.001;10^-7 mol/L:2.91±0.29 vs.1.01±0.08,t=10.83,P<0.001;10^-8 mol/L,1.67±0.07 vs.1.00±0.08,t=11.40,P<0.001)and collagen I(10^-6 mol/L:2.68±0.09 vs.1.00±0.07,t=24.94,P<0.001;10^-7 mol/L:2.12±0.29 vs.1.01±0.12,t=6.08,P<0.001;10^-8 mol/L:1.46±0.15 vs.1.00±0.05,t=4.93,P=0.008).However,different concentrations of SQ29548 all significantly reduced the expression of collagen I(10^-4 mol/L:0.55±0.07 vs.1.00±0.07,t=10.47,P<0.001;10^-6 mol/L:0.56±0.10 vs.1.00±0.07,t=6.185,P<0.001;10^-7 mol/L:0.27±0.04 vs.1.00±0.07,t=15.41,P<0.001)and a-SMA(10^-4 mol/L:0.06±0.01 vs.1.00±0.11,t=15.17,P<0.001;10^-6 mol/L:0.28±0.03 vs.1.00±0.11,t=11.29,P<0.001;10^-7 mol/L:0.14±0.04 vs.1.00±0.11,t=12.86,P<0.001).After being treated with SQ29548(10^-4 mol/L)and then 8-epi-PGF2α(10^-7 mol/L),the mRNA levels of a-SMA(0.20±0.08 vs.1.00±0.00,t=17.46,P<0.001)and collagen I(0.69±0.13 vs.1.00±0.00,t=4.20,P=0.014)in PSCs were significantly lower than those of the control group.Conclusions:The results show that 8-epi-PGF2α promoted PSCs activation,while SQ29548 inhibited PSCs activation induced by 8-epi-PGF2α.The result indicated that TxA2r plays an important role during PSC activation and collagen synthesis induced by 8-epi-PGF2α in vitro.This receptor may provide a potential target for more effective antioxidant therapy for pancreatic fibrosis.

Apigenin is an anoikis sensitizer with strong anti-metastatic properties in experimental breast cancer

Loss of susceptibility to anoikis signals is a crucial step in metastasis.Anoikis resistance therefore represents a promising adjuvant therapeutic target for cancer management.In this study,we have conducted a rationalized screening to search for novel leading anoikis sensitizer from daily foods.Among 19 tested dietary phytochemicals,the best results were obtained with apigenin,a natural component of celery.Phenotypically,apigenin sensitized breast cancer cells to anoikis,lowered the number of circulating tumor cells,and protected against breast cancer metastasis to lung in mice.Mechanistically,we demonstrated that the thromboxane A_(2)(TXA_(2))-TXA_(2)receptor(TP)axis has a critical role in acquired anoikis resistance by activating PI3K-Akt signaling pathway.Blockage of TXA_(2)signaling up-regulated p53 as well as its target gene p21,caused a G1 phase arrest,and finally led to apoptosis in breast cancer cells.TXA_(2)level was positively correlated with breast cancer cell anoikis rate,and apigenin significantly inhibited TXA_(2)biosynthesis in vitro and in vivo.Collectively,we identified apigenin as a potent anoikis sensitizer with anti-metastatic properties in a mouse model of breast cancer,and these findings might provide a rationale for introducing apigenin supplementation to breast cancer patients.

Associations between thromboxane A synthase 1 gene polymorphisms and the risk of ischemic stroke in a Chinese Han population

Thromboxane A synthase 1 （TBXAS1） catalyses the synthesis of thromboxane A2 （TXA2）, which plays an important role in the pathogenesis of ischemic stroke. Thus, the TBXAS1 gene was investigated as a candidate gene involved in the formation of atherosclerosis. This case-control study collected peripheral blood specimens and clinical data of 370 ischemic stroke patients and 340 healthy controls in the Northern Chinese Han population from October 2010 to May 2011. Two TBXAS1 single-nucleotide polymorphisms, rs2267682 and rs10487667, were analyzed using a SNaPshot Multiplex sequencing assay to explore the relationships between the single-nucleotide polymorphisms in TBXAS1 and ischemic stroke. The TT genotype frequency and T allele frequency of rs2267682 in the patients with ischemic stroke were significantly higher than those in the controls （P 〈 0.01 and P = 0.02）. Furthermore, compared with the GG ＋ GT genotype, the TT rs2267682 genotype was associated with increased risk of ischemic stroke （odds ratio （OR） = 1.80, 95% confidence interval （CI）： 1.16–2.79, P 〈 0.01）. Multivariate logistic analysis with adjustments for confounding factors revealed that rs2267682 was still associated with ischemic stroke （OR = 1.94,95% CI ： 1.13–3.33, P = 0.02）. The frequency of the T-G haplotype in the patients was significantly higher than that in the controls according haplotype analysis （OR = 1.49, 95% CI： 1.10–2.00, P 〈 0.01）. These data reveal that the rs2267682 TBXAS1 polymorphism is associated with ischemic stroke. The TT genotype of TBXAS1 and T allele of rs2267682 increase susceptibility to ischemic stroke in this Northern Chinese Han population. The protocol has been registered with the Chinese Clinical Trial Registry （registration number： ChiCTR-COC-17013559）.

Targeting brain microvascular endothelial cells: a therapeutic approach to neuroprotection against stroke

Brain microvascular endothelial cells form the interface between nervous tissue and circulating blood, and regulate central nervous system homeostasis. Brain microvascular endothelial cells differ from peripheral endothelial cells with regards expression of specific ion transporters and receptors, and contain fewer fenestrations and pinocytotic vesicles. Brain microvascular endothelial cells also synthesize several factors that influence blood vessel function. This review describes the morphological characteristics and functions of brain microvascular endothelial cells, and summarizes current knowledge regarding changes in brain microvascular endothelial cells during stroke progression and therapies. Future studies should focus on identifying mechanisms underlying such changes and developing possible neuroprotective therapeutic interventions.

Clinical Implication of the Changes of cAMP, TXA_2 and PGI_2 in CSF of Asphyxiated Newborns

To evaluate the changes of 3', 5'-cyclic adenosine monophosphate (cAMP), thrombox-ane A2(TXA2) and prostacyclin (PGI2) in cerebrospinal fluid (CSF) in the asphyxiated newborn and explore their roles in hypoxic-ischamic brain damage (HIBD). Thirty-six full term newborns were divided into 3 groups, including 12 with moderate-severe hypoxic-ischaemic encephalopathy (HIE), 13 with mild HIE, 11 without HIE (control group). The levels of cAMP, TXB2(TXA2 metabolite) and 6-keto-PGF1α(PGI2 metabolite) in CSF and plasma were measured 36-72 h after birth by RIA, and the concentrations were expressed as nM/L (cAMP), ng/L(TXB2 and 6-keto-PGF1α). The infants were followed-up at 6 and 12 month of age and Mental Development Index (MDI) and Psychomotor Development Index (FDD were measured using Bayley Scales of Infant Development (BSID). The CSF cAMP level in moderate-severe HIE group was 8. 60±2. 40, significantly lower than that of the mild HIE group (14. 83±2. 84) and the control group (24. 43±2. 39)(for both P<0. 01). The levels of TXB2 and 6-keto-PGF1α in CFS in the moderate-severe HIE group (206. 06±29. 74, 168. 47± 23. 02, respectively) were significantly higher than in the mild HIE group (83. 37±28. 57, 131. 42± 16. 57, respectively, P<0. 01) and the control group (41. 77±21. 58, 86. 23±13. 05, respectively, P<0. 01). The level changes of cAMP,TXB2 and 6-keto-PGF1α in plasma in all groups were similar to those in CSF, but no significant difference was found between mild HIE group and the control group (P>0. 05). The follow-up results showed that MDI and PDI of the moderate-severe HIE group were the lowest (84. 79±13. 34, 83. 50±13. 28, respectively), followed by mild HIE group (102.19±7. 02, 99. 94±9. 08, respectively) , with the control group being the highest (116. 63± 12.08, 116. 69±10. 87, respectively). Univariate analysis showed some significant difference (the moderate-severe HIE group vs. the mild HIE group or the control group, P<0. 01; the mild HIE group vs. the control group P<0. 05). The results suggested that the concentration of cAMP, TXA2 and T/K ratio in CSF after neonatal asphyxia might be sensitive markers in evaluating the severity of brain damage in early stage and predicting the future outcome.

Protective Effect of Herba Dracocephali on Lipoperoxidation Damage in Patients with Coronary Heart Disease

In order to study the effects of Herba Dracocephali (HD) in treating coronary heart disease(CHD) and antiperoxidative damage. 40 patients of CHD were treated with HD and the changes ot plasma su-peroxide dismutase (SOD) , lipid peroxide (LPO) , thromboxane B2 (TXB2) and 6-keto-prostaglandin F1 (6-keto-PGF1 ) as well as platelet LPO (PL-LPO) , SOD (PL-SOD) and selenium-glutathion peroxidase (SGP)were measured before and after treatment. Results showed that TXB2 , LPO and PL-LPO increased, whileSOD, PL-SOD decreased signiticantly in patients. After treatment by HD, the TXB2 , LPO, PL-LPO loweredand 6-keto-PGF1 , SOD, PL-SOD and SGP elevated obviotjsly. Electron microscopy showed that the ultra-structore of platelets improved and the activity ot adenylate cyclase increased markedly. The results suggest-ed that HD is a traditional Chinese herbal antioxidant and is hopeful tor treating CHD.

Thrombosis and its significance after experimental pulmonary thromboembolism

Objective To study thrombosis and its significance after acute experimental pulmonary thromboembolism. Methods The acute pulmonary thromboembolism (PTE) model of rabbits was established by intravenous injection of autologous blood clots (0.04 g/kg) which were stabilized in temperature-controlled (70℃) distilled water for 10 min. The process of thrombosis was observed grossly and microscopically. The Quick's method was used to examine the coagulability of blood and radioimmunoassay was employed to measure the level of plasma thromboxane A 2 and endothelin. Results Thrombotic propensity was observed at 1 h, fresh thrombus started to form and the blood coagulation system was activated at 24 h following clots infusion. Emboli were completely or partly dissolved at 5 d and appeared to organize at both 10 d and 14 d after clots were infused. Venous plasma thromboxane A 2 concentration began to increase at 5 min (2489.59±714.68 ng/L) and reached its maximum at 15 min (2545.46±590.58 ng/L) then declined at 60 min after clot infusion (P<0.001, respectively, vs 626.59±510.02 ng/L of pre-clot). The level of endothelin in both arterial and venous blood increased at 5 d post-clot infusion (840.74±154.19 ng/L, 230.35±52.39 ng/L, respectively) compared to the one before infusion (602.66±453.26 ng/L, 148.01±53.28 ng/L, respectively, P<0.05).Conclusions Thrombosis occurs after autologous-blood-clot-induced PTE. The interactions between thrombus formation, fibrinolysis and organization determines the consequences of emboli. Abnormalities of endothelin metabolism and the increment of thromboxane A 2 may play an important role in PTE.