[Effect of Kanli Granule on Myocardial Mechanics in Pressure Overload Induced Diastolic Heart Failure Rats].

OBJECTIVE: To observe the effect of Kanli Granule (KG) on myocardial mechanics in pressure overload induced diastolic heart failure (DHF) rats. METHODS: Totally 60 male Wistar rats were divided into the sham-operation group, the model group, the KG group, and the Valsartan group according to random digit table, 15 in each group. The pressure overload induced DHF model was established in all groups except the sham-operation group using abdominal aortic constriction surgery. Totally 7 rats died after modeling (with the mortality of 10. 67%) , and the rest 53 finished the following test. Rats in the KG group were administered with KG extract (calculated as 6. 75 g crude drug/kg) by gastrogavage. Rats in the Valsartan group were administered with Valsartan (7.2 µg/g) by gastrogavage. Equal volume of double distilled water was administered to rats in the model group and the sham-operation group by gastrogavage. All rats were intervened for 32 weeks. The response of isolated heart papillary muscle tonus to isoprenaline (ISO) and adenylate cyclase (Forskolin) was respectively observed. The enhancement phenomenon after resting development force (DF) of isolated heart papillary muscle tonus, and changes of DF in different Ca²âº concentrations were observed. RESULTS: (1) In the ISO response test: Compared with the sham-operation group, the amplifications of DF, ±df/dt, -df/dt were obviously elevated in the model group (P < 0.05). Compared with the model group, the amplifications of DF and ±df/dt were obviously lowered in the KG group (P < 0.01), and the amplification of ±df/dt was also reduced in the Valsartan group (P < 0.01). (2) In the Forskolin response test: Compared with the sham-operation group, the amplifications of DF and ±df/dt obviously increased in the model group (P < 0.05). Compared with the model group, the amplifications of DF and ±df/dt were obviously reduced in the KG group (P < 0.01), and the amplification of DF was also reduced in the Valsartan group (P < 0.05). (3) In post-resting DF enhancement test: Compared with the sham-operation group, the amplification of DF showed gradually decreasing tendency along with prolonged resting time in the model group, and they were obviously lowered at all time points (P < 0.05). Compared with the model group, the amplification of DF was gradually increasing along with prolonged resting time in the KG group. The amplification of DF at post-resting 240 s was obviously larger in the KG group than in the model group (P < 0.05). The amplification of post-resting DF still showed gradually decreasing tendency along with prolonged resting time in the Valsartan group, with increased amplifications of DF at post-resting 60 s and 120 s (P < 0. 05) (4) The amplifications of DF in different Ca²âº concentrations: Compared with the sham-operation group, the amplifications of DF were significantly elevated in different Ca²âº concentrations (1.75, 3.5, 7.0 mmol/L ) (P < 0.05, P < 0.01). Compared with the model group, there was no statistical difference in amplification of DF in different Ca²âº concentrations in the KG group (P > 0.05). The amplifications of DF in different Ca²âº concentrations were significantly reduced in the Valsartan group (P < 0.05). CONCLUSIONS: The ISO response and the Forskolin response were enhanced in isolated heart papillary muscle tonus of pressure overload induced DHF rats; enhanced post-resting DF was reduced; DF in different supra-physiologic levels of Ca²âº was still enhanced. KG could significantly improve excessive enhancement of pressure overload induced DHF rats in ISO response and Forskolin response, and improve enhancement of post-resting myocardium.

Kurarinone Synergizes TRAIL-Induced Apoptosis in Gastric Cancer Cells.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been identified as a promising anti-tumor agent against in a variety of cancers. However, gastric cancer cells are less sensitive than other cancer cells to TRAIL-induced apoptosis. Here, we combined TRAIL with kurarinone, a natural compound, to induce apoptosis in gastric cancer cell lines SGC7901. After the cells were treated with TRAIL and/or kurarinone, the cell viability and apoptosis were examined by MTT and flow cytometry, respectively. The expression of apoptosis-associated proteins was determined by western blot and q-RT-PCR. Kurarinone at low concentration significantly potentiated the cytotoxic effect of TRAIL by enhancing apoptosis as well as cell cycle arrest at G2/Mphase. The enhancement of apoptosis TRAIL induced by kurarinone involved downregulation of anti-apoptotic proteins Mcl-1 and c-FLIP as well as inhibition of STAT3 signaling. Moreover, we found that STAT3 inhibitor could synergistically enhanced TRAIL-induced apoptosis, similar to kurarinone. Kurarinone synergizes TRAIL-induced apoptosis in human gastric cancer cells. The synergistic effect between these two drugs is associated with downregulation of Mcl-1 and c-FLIP via inhibiting STAT3 signaling. The combination of TRAIL and kurarinone might be an effective regimen for the treatment of advanced gastric cancer.

Tangeretin, a citrus polymethoxyflavonoid, induces apoptosis of human gastric cancer AGS cells through extrinsic and intrinsic signaling pathways.

Tangeretin, a natural polymethoxyflavone present in citrus peel oil, is known to have anticancer activities in breast cancer, colorectal carcinoma and lung carcinoma, yet, the underlying mechanisms of tangeretin in human gastric cancer AGS cells have not been investigated to date. In the present study, the apoptotic mechanisms of tangeretin in AGS cells were explored. It was observed that tangeretin increased the apoptotic rates of AGS cells following treatment with tangeretin for 48 h in a dose-dependent manner by Annexin V-FITC and PI double staining. In addition, characteristic apoptotic morphology such as nuclear shrinkage and apoptotic bodies was observed after Hoechst 33258 staining. Flow cytometric assay showed that treatment of AGS cells with tangeretin decreased the mitochondrial membrane potential (MMP) in a dose-dependent manner, which indicated that mitochondrial dysfunction was involved in the tangeretin-induced apoptosis. Caspase-3, -8 and -9 activities were increased by tangeretin in a dose-dependent manner. Western blotting showed that the protein levels of pro-apoptotic proteins including cleaved caspase-3, cleaved caspase-8, cleaved caspase-9, Bax, Bid, tBid, p53, p21/cip1, Fas and FasL were significantly upregulated by tangeretin. In addition, PFT-α (a p53 inhibitor) reduced the apoptotic rates and the expression of p53, p21, caspase-3 and caspase-9 induced by tangeretin, indicating that tangeretin-induced apoptosis was p53-dependent. In conclusion, these results suggest that tangeretin induces the apoptosis of AGS cells mainly through p53-dependent mitochondrial dysfunction and the Fas/FasL-mediated extrinsic pathway.

Curcumin induces apoptosis in human gastric carcinoma AGS cells and colon carcinoma HT-29 cells through mitochondrial dysfunction and endoplasmic reticulum stress.

In the present study, we investigate the effect of curcumin, a major active component isolated from rhizomes of Curcuma longa, on the cytotoxicity of three human carcinoma cell lines (AGS, HT-29 and MGC803) in gastrointestinal tract and a normal gastric epithelial cell line GES-1, and the mechanism of curcumin-induced apoptosis. The results indicated that curcumin inhibited the gastrointestinal carcinoma cell growth in a dose-dependent manner and cytotoxicity was more towards the gastric carcinoma cell AGS and colon carcinoma cell HT-29 compared to normal gastric cell GES-1, and increased externalization of phosphatidylserine residue was observed by Annexin V/PI staining in the two cell lines. Treatment of AGS and HT-29 cells with curcumin enhanced the cleavage of procaspase-3, -7, -8 and -9. Meanwhile, curcumin induced endoplasmic reticulum (ER) stress and mitochondrial dysfunction as evidenced by up-regulation of CCAAT/enhancer binding protein homologous protein (CHOP), phosphorylation of JNK and down-regulation of SERCA2ATPase, release of cytochrome c, decrease of Bcl-2 and reduction of mitochondrial membrane potential in both AGS and HT-29 cells. Overexpression of bax, total JNK, phospho-FADD and total FADD were also observed in curcumin-treated HT-29 cells. Moreover, curcumin decreased cytosolic and ER Ca(2+), but increased mitochondrial Ca(2+) in the two cell lines. 2-Aminoethoxydiphenyl borate, an antagonist of inositol 1, 4, 5-triphosphate receptor, partly blocked curcumin-induced cytosolic Ca(2+) decrease in AGS and HT-29 cells. Additionally, carbonyl cyanide m-chlorophenylhydrazone, an inhibitor of mitochondrial Ca(2+) uptake, reversed curcumin-triggered AGS and HT-29 cells growth inhibition. siRNA to CHOP markedly reduced curcumin-induced apoptosis. These results suggest that curcumin can impact on ER stress and mitochondria functional pathways in AGS and HT-29 cells, death receptor pathway was also involved in curcumin-treated HT-29 cells, thus identifying specific well-defined molecular mechanisms that may be targeted by therapeutic strategies.

[Effects of sinensetin on proliferation and apoptosis of human gastric cancer AGS cells].

OBJECTIVE: To study the effects and mechanisms of sinensetin on proliferation and apoptosis of human AGS gastric cancer cells. METHOD: MTT assay was used to detect the growth inhibition rates of human AGS gastric cancer cells treated with sinsesectin in different concentrations and times. The cell cycle distribution was measured by flow cytometry. The apoptosis was examined by Annexin-FITC/PI staining and DNA fragment analysis. The apoptosis morphology was observed by inverted fluorescence microscope after Hoechst 33342 staining. The protein expressions of p21 and p53 were detected by western blot. RESULT: MTT assay showed that sinensetin inhibited the growth of AGS gastric cancer cells in a dose- and time-dependent manner. Sinensetin blocked AGS cells in G2/ M and increased the apoptosis rates of AGS cells in a dose-dependent manner. DNA ladder was observed in cells treated with 60 micromol x L(-1) sinensetin for 48 h. The typical apoptotic morphological changes including cell nucleus shrinkage, chromatin condensation and apoptotic bodies were observed when treated with different dose of sinensetin. Western blot showed that sinensetin increased expressions of p53 and p21 in a dose-dependent manner. CONCLUSION: Sinensetin could inhibit human AGS gastric cancer cells proliferation and induce cell cycle block in G2/M phase and apoptosis. The up regulation of p53 and p21 protein might be one of the mechanisms.

Traditional Chinese formula, lubricating gut pill, stimulates cAMP-dependent CI(−) secretion across rat distal colonic mucosa.

AIM OF THE STUDY: Lubricating gut pill (LGP), a traditional Chinese formula, had been conformed to improve the loperamide-induced rat constipation by stimulation of Cl(-) secretion, but its mechanism has not been fully explored. Thus, the purpose of this study was to identify the action sites of LGP-stimulated Cl(-) secretion across rat distal colonic mucosa. MATERIALS AND METHODS: Rat distal colonic mucosa was mounted in Ussing chambers and short circuit current (I(SC)), apical Cl(-) current and basolateral K(+) current were recorded. Intracellular cyclic adenosine monophosphate (cAMP) content and protein kinase A (PKA) activity were determined with ELISA kit and the non-radioactive PepTag test, respectively. RESULTS: LGP at 800µg/ml elicited a sustained increase in Cl(-) secretory response, which was inhibited by CFTR(inh)172, a cystic fibrosis transmembrane conductance regulator (CFTR) inhibitor. Permeabilizing apical membrane with nystatin revealed that LGP-stimulated basolateral K(+) current was significantly inhibited by KCNQ1 K(+) channel inhibitor chromanol 293B. LGP-stimulated I(SC) was markedly reduced by pretreatment with cis-N-[2-phenylcyclopentyl]-azacyclotridec-1-en-2amine (MDL-12,330A) and N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide (H-89), but not with inhibitors of Ca(2+)-dependent signaling pathway. Treatment of tissue with LGP resulted in an increase in intracellular cAMP level and the activation in protein kinase A. The E-prostanoid(4) (EP)(4) receptor antagonist L-161,982 completely eliminated LGP-induced response. CONCLUSIONS: The results showed that LGP enhances Cl(-) and fluid secretion via prostanoid receptor signaling and also cAMP and protein kinase A pathway, subsequently triggering the activation of apical Cl(-) channels mostly CFTR and basolateral cAMP-dependent K(+) channel.

[Effects of Coptis chinensis and Evodia rutaecarpa water extract on DMH-induced precancerous lesion of colon].

OBJECTIVE: To investigate the effects of Coptis chinensis and Evodia rutaecarpa water extract on precancerous lesion of colon induced by DMH and proliferation and apoptosis changes of colon mucosa crypts. METHOD: Precancerous lesion of colon was induced by DMH. The changes of proliferation and apoptosis of colon mucosa crypts were detected by morphological analysis. The numbers of aberrant crypt foci (ACF) were measured by feulgen staining. RESULT: C. chinensis and E. rutaecarpa water extract could significantly inhibit the formation of ACF in model animals. The proliferative crypts were increased obviously in middle and distal colon, and decreased by C. chinensis and E. rutaecarpa water extract. The apoptosis crypts were increased in distal colon but not middle colon. C. chinensis and E. rutaecarpa water extract could promote apoptosis of both middle and distal colon. CONCLUSION: C. chinensis and E. rutaecarpa water extract could significantly inhibit the formation of ACF in model animals. These results indicated that C. chinensis and E. rutaecarpa water extract maybe have an inhibitory and clinically therapeutic effect on colon cancer, which were partly resulted from inhibiting proliferation and promoting apoptosis of crypts in middle and distal colon.

Structural shifts of gut microbiota as surrogate endpoints for monitoring host health changes induced by carcinogen exposure.

This study monitored structural shifts of gut microbiota of rats developing precancerous mucosal lesions induced by carcinogen 1,2-dimethyl hydrazine (DMH) treatment using PCR-denaturing gradient gel electrophoresis (DGGE) and 454 pyrosequencing on the 16S rRNA gene V3 region. Partial least square discriminant analysis of DGGE fingerprints showed that the gut microbiota structure of treated animals was similar to that of the controls 1 and 3 weeks after DMH treatments, but significantly different 7 weeks after DMH treatments, when a large number of aberrant crypt foci (ACF) developed in their colons. Martens' uncertainty test, followed by anova test (P<0.05) identified Ruminococcus-like and Allobaculum-like bacteria as key variables for discrimination of DMH-treated rats from controls. Real-time PCR confirmed the significant increase of the Ruminococcus obeum and the Allobaculum-like bacteria in DMH-treated rats. UniFrac analysis based on V3 pyrosequencing further validated that the gut microbiota structures of treated and control animals were similar at an early stage, but segregated after ACF formation. Thirteen operational taxonomic units including Ruminococcus-like and Allobaculum-like bacteria were identified as key variables for the discrimination of DMH-treated rats from controls. Dynamic analysis of gut microbiota may become a noninvasive strategy for monitoring host health changes induced by carcinogen exposure.

Traditional Chinese formula, lubricating gut pill, improves loperamide-induced rat constipation involved in enhance of Cl- secretion across distal colonic epithelium.

AIM OF THE STUDY: Lubricating gut pill (LGP), a traditional Chinese formula, was widely used for the treatment of chronic constipation, especially in the elderly, in China. However, it is unclear whether LGP-induced laxative and/or lubricating effect is involved in water and electrolytes transport in distal colonic epithelium. MATERIALS AND METHODS: The present study was designed to evaluate the effect of LGP on Cl(-) secretion across rat distal colonic epithelium mounted in Ussing chambers, and on a rat constipation model induced by loperamide, respectively. RESULTS: Application of LGP in the apical side elicited a sustained increase in short circuit current (I(SC)) response in a concentration-dependent manner. Evidence that LGP-stimulated I(SC) was due to Cl(-) secretion is based on inhibition of current by (a) a Na(+)-K(+)-2Cl(-) cotransporter inhibitor bumetanide, (b) removal of Cl(-) ions in bath solution, and (c) the cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channel blocker DPC, suggesting that a apical cAMP-dependent Cl(-) channel was activated. LGP-stimulated I(SC) was also strongly inhibited by pretreatment with clotrimazole, indicating that the basolateral K(+) channel was also involved in maintaining this cAMP-dependent Cl(-) secretion. Pretreatment of tissues with indomethacin, but not atropine, tetrodotoxin or hexamethonium, inhibited LGP-induced response. In a rat constipation model, oral administration with LGP was significantly restored number of fecal pellets, water content and mucus secretion compared with loperamide-treated group alone. CONCLUSIONS: LGP enhances Cl(-) secretion that is mostly mediated through the release of cyclooxygenase metabolites, by which provided an osmotic force for the subsequent laxative action observed in the rat constipation model.

[Effects of Astragalus membranaccus on cardiac function and SERCA2a gene expression in myocardial tissues of rats with chronic heart failure].

OBJECTIVE: To investigate the effects of Astragalus membranaccus (As) on cardiac function and SERCA2a gene expression in left ventricular tissues of rats with chronic heart failure. METHODS: Heart failure was induced by clipping the abdominal aorta 60 male SD rats were divided into four groups: sham-operated (Sham), aortic stenosis (Model), Model + As (20 g/kg) and Model + Captopril (0.05 g/kg). The drugs were administered orally from the 13th week after surgery. Rats were examined after 12 weeks' treatment with drugs. The parameters of hemodynamics including LVSP, LVEDP, and +/- LVdp/dt(max) were measured. SERCA2a mRNA and protein expressions in left ventricular tissues were determined by half-quantitative RT-PCR and Western blot normalized to abundance of GAPDH mRNA and portein, respectively. RESULTS: LVSP and LVEDP were obviously enhanced (P < 0.01 or P < 0.001) in model rats in vivo. Both Captopril and As prevented the increase of LVSP (P < 0.05 or P < 0.01) and LVEDP (P < 0.05 or P < 0.01). RT-PCR and Western blot results demonstrated that SERCA2a gene expression was downregulated (P < 0.05) significantly in model group compared with sham group. As upregulated SERCA2a gene expression (P < 0.05), whereas Captopril had no effect on that. CONCLUSION: As can ameliorate abnormity of cardiac function, especially diastoilc function in rats with pressure overload-induced heart failure, and that may be partly related to its up-regulation of SERCA2a gene expressions in left ventricular tissues.

[Effect of astragalus on calcium accumulation and SERCA2a gene expression in myocardial tissues in rats with pressure overload-induced left ventricular hypertrophy].

OBJECTIVE: To investigate the effect of astragalus (As) on calcium accumulation and SERCA2a gene expression in left ventricular tissues in rats with pressure overload-induced cardiac hypertrophy. METHOD: cardiac hypertrophy was induced by clipping the abdominal aorta in rats. Male SD rats were allocated to six groups: sham-operrated (Sham), aortic stenosis (Model), model +As-L (5 g x kg(-1) x d(-1)), model+As-M (10 g x kg(-1) x d(-1)), model+As-H (20 g x kg(-1) x d(-1)) and model + captopril (0.05 mg x kg(-1) x d(-1), a positive control). The drugs were administered orally from the 13 th week after surgery. Rats were examined after 12 week treatment with drugs. The cardiac hypertrophy was evaluated by left ventricular mass index (LVMI, left ventricular weight/ body weight). The calcium content in left ventricular tissue was measured by atomic absorption spectrometry. SERCA2a mRNA and protein expressions in left ventricular tissues were determined by half-quantitative RT-PCR and Western blot normalized to abundance of GAPDH mRNA and protein, respectively. RESULT: The increase of LVMI was dose-dependently lessened by As (P < 0.01, P < 0.001). The effect of As-H was similar to that of Captopril. As markedly attenuated calcium accumulation in myocardial tissure (P < 0.01). RT-PCR and Western blot results demonstrated that SERCA2a gene expressions were downregulated (P < 0.05) significantly in model group compared with sham group. As-H upregulated SERCA2a gene expressions (P < 0.05), whereas Captopril had no effect on that. CONCLUSION: The inhibition of As on left ventricular hypertrophy induced by pressure overload in rats may partly contribute to its attenuation of calcium accumulation and up-regulation of SERCA2a gene expressions in left ventricular tissues.

Rutaecarpine induces chloride secretion across rat isolated distal colon.

The present study evaluated the effect of rutaecarpine (Rut) on Cl(-) secretion across rat distal colonic mucosa. Basolateral application of Rut elicited an increase in short-circuit current (I(SC)) response in a concentration-dependent manner. Evidence that Rut-stimulated I(SC) was due to Cl(-) secretion is based on 1) inhibition of current by bumetanide; 2) Cl(-) channel blockers diphenylamine-2-carboxylate, 5-nitro-2-(3-phenylpropylamino)-benzoic acid, and glibenclamide; and 3) removal of Cl(-) ions in bath solution. Determination of neurogenic blockers on Rut-induced I(SC) indicated that pretreatment of tissues with tetrodotoxin or indomethacin, but not atropine or hexamethonium, inhibited Rut-induced response. Treatment with Rut led to release and synthesis of prostaglandin E(2) in rat colonic mucosa. Rut-stimulated I(SC) was markedly reduced by pretreatment with MDL-12,330A [cis-N-[2-phenylcyclopentyl]-azacyclotridec-1-en-2-amine] and N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide (H-89), but not with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester, bisindolylmaleimide, and thapsigargin. Elimination of the extracellular Ca(2+) also did not alter Rut response. Rut treatment resulted in the increase in intracellular cAMP levels and the activation of protein kinase A. Depolarizing the basolateral membrane with high K(+) showed that Rut-stimulated apical Cl(-) current was largely prevented by cystic fibrosis transmembrane conductance regulator (CFTR) inhibitors. Permeabilizing apical membrane with nystatin revealed that Rut-stimulated basolateral K(+) current was specifically inhibited by Ba(2+) ions and chromanol 293B. The evidence derived from present study suggests that Rut-stimulated Cl(-) secretion is mediated by generation of endogenous prostaglandin E(2) and that it also involves the stimulation of cAMP and protein kinase A pathways, which subsequently lead to the activation of apical Cl(-) channels, mostly the CFTR and basolateral cAMP-dependent K(+) channels.

[Effect of jinfu kang to experimental precancerous colon lesions and urinary metabolites in rat].

OBJECTIVE: : To profile urinary metabolite variations from 1, 2-dimethylhydrazine (DMH)-induced precancerous colon rats, Jinfu Kang treated rats and healthy controls. METHOD: We used ethyl chloroformate derivatization and gas chromatography-mass spectrometry (GC-MS) based metabonomic method to analyze rat urines. RESULT: The time-dependent variations of metabolite profile showed a progressive deviation of the metabolism in the model group from the initial pattern over time and a systemic recovery of the metabolism in the treatment group, which is consistent with the histological results. The in-depth analysis indicated that the disorder of tricarboxylic acid cycle (TCA), tryptophan metabolism, polyamine metabolism and gut flora structure were associated with DMH intervention. CONCLUSION: Metabolic study revealed that Jinfu Kang can effectively reverse metabolic departures in DMH-induced precancerous colon rat, which is consistent with pathological results.

Erianin induces a JNK/SAPK-dependent metabolic inhibition in human umbilical vein endothelial cells.

BACKGROUND: Erianin is a natural product derived from Dendrobium chrysotoxum, with promising antitumor activity. MATERIALS AND METHODS: To evaluate the metabolic effect of erianin, a cytosensor assay for acidification rate, MTT assay, measurement of lactate, glucose and ATP were performed in human umbilical vein endothelial cells (HUVECs) exposed to 1-100 nM erianin. JNK/SAPK activity was detected by Western blot. RESULTS: Twelve- or 24- hour incubation with erianin induced a dose-dependent metabolic inhibition, as indicated by reduced acidification rate and cell viability, with an endothelium-selectivity. Erianin caused decreases in lactate production, glucose consumption and intracellular ATP level. Pretreatment with the JNK/SAPK inhibitor SP600125 significantly abolished these inhibitory responses, and especially restored the erianin-induced decreases in ATP and the erianin-induced phosphorylation of JNK/SAPK with dose- and time- dependence. CONCLUSION: Erianin inhibited endothelial metabolism in a JNK/SAPK-dependent manner. This mechanism may be involved in the potential antitumnor and antiangiogenic actions of erianin.

Effects of calycosin on the impairment of barrier function induced by hypoxia in human umbilical vein endothelial cells.

The purpose of the present study was to examine the effects of calycosin, an isoflavonoid isolated from Astragali Radix, on the impairment of barrier function induced by hypoxia in cultured human umbilical vein endothelial cells. Hypoxia induced an increase in endothelial cell monolayer permeability, indicating endothelial cell barrier impairment. Endothelial barrier dysfunction induced by hypoxia was accompanied by decreases in cytosolic ATP concentration and cAMP level, the development of actin stress fibers and intercellular gap formation, suggesting that the decreases in cytosolic ATP and cAMP levels and rearrangements of F-actin could be associated with an increase in permeability of endothelial monolayers. Application of calycosin inhibited the hypoxia-induced increase in endothelial permeability in a dose-dependent fashion, which is compatible with inhibition of lactate dehydrogenase release, decrease of the fall in ATP and cAMP contents, and improvement of F-actin rearrangements. These findings indicate that calycosin protected endothelial cells from hypoxia-induced barrier impairment by increasing intracellular energetic sources and promoting regeneration of the cAMP level, as well as improving cytoskeleton remodeling.