Astragali Radix Isoflavones Synergistically Alleviate Cerebral Ischemia and Reperfusion Injury Via Activating Estrogen Receptor-PI3K-Akt Signaling Pathway.

Isoflavones are major neuroprotective components of a medicinal herb Astragali Radix, against cerebral ischemia-reperfusion injury but the mechanisms of neuroprotection remain unclear. Calycosin and formononetin are two major AR isoflavones while daidzein is the metabolite of formononetin after absorption. Herein, we aim to investigate the synergistic neuroprotective effects of those isoflavones of Astragali Radix against cerebral ischemia-reperfusion injury. Calycosin, formononetin and daidzein were organized with different combinations whose effects observed in both in vitro and in vivo experimental models. In the in vitro study, primary cultured neurons were subjected to oxygen-glucose deprivation plus reoxygenation (OGD/RO) or l-glutamate treatment. In the in vivo study, rats were subjected to middle cerebral artery occlusion to induce cerebral ischemia and reperfusion. All three isoflavones pre-treatment alone decreased brain infarct volume and improved neurological deficits in rats, and dose-dependently attenuated neural death induced by l-glutamate treatment and OGD/RO in cultured neurons. Interestingly, the combined formulas of those isoflavones revealed synergistically activated estrogen receptor (estrogen receptors)-PI3K-Akt signaling pathway. Using ER antagonist and phosphatidylinositol 3-kinase (PI3K) inhibitor blocked the neuroprotective effects of those isoflavones. In conclusion, isoflavones could synergistically alleviate cerebral ischemia-reperfusion injury via activating ER-PI3K-Akt pathway.

Glycyrrhizin Prevents Hemorrhagic Transformation and Improves Neurological Outcome in Ischemic Stroke with Delayed Thrombolysis Through Targeting Peroxynitrite-Mediated HMGB1 Signaling.

Peroxynitrite (ONOO-) and high mobility group box 1 protein (HMGB1) are important cytotoxic factors contributing to cerebral ischemia-reperfusion injury. However, the roles of ONOO- in mediating HMGB1 expression and its impacts on hemorrhagic transformation (HT) in ischemic brain injury with delayed t-PA treatment remain unclear. In the present study, we tested the hypothesis that ONOO- could directly mediate the activation and release of HMGB1 in ischemic brains with delayed t-PA treatment. With clinical studies, we found that plasma nitrotyrosine (NT, a surrogate marker of ONOO-) was positively correlated with HMGB1 level in acute ischemic stroke patients. Hemorrhagic transformation and t-PA-treated ischemic stroke patients had increased levels of nitrotyrosine and HMGB1 in plasma. In animal experiments, we found that FeTmPyP, a representative ONOO- decomposition catalyst (PDC), significantly reduced the expression of HMGB1 and its receptor TLR2, and inhibited MMP-9 activation, preserved collagen IV and tight junction claudin-5 in ischemic rat brains with delayed t-PA treatment. ONOO- donor SIN-1 directly induced expression of HMGB1 and its receptor TLR2 in naive rat brains in vivo and induced HMGB1 in brain microvascular endothelial b.End3 cells in vitro. Those results suggest that ONOO- could activate HMGB1/TLR2/MMP-9 signaling. We then addressed whether glycyrrhizin, a natural HMGB1 inhibitor, could inhibit ONOO- production and the antioxidant properties of glycyrrhizin contribute to the inhibition of HMGB1 and the neuroprotective effects on attenuating hemorrhagic transformation in ischemic stroke with delayed t-PA treatment. Glycyrrhizin treatment downregulated the expressions of NADPH oxidase p47 phox and p67 phox and iNOS, inhibited superoxide and ONOO- production, reduced the expression of HMGB1, TLR2, MMP-9, preserved type IV collagen and claudin-5 in ischemic brains. Furthermore, glycyrrhizin significantly decreased the mortality rate, attenuated hemorrhagic transformation, brain swelling, blood-brain barrier damage, neuronal apoptosis, and improved neurological outcomes in the ischemic stroke rat model with delayed t-PA treatment. In conclusion, peroxynitrite-mediated HMGB1/TLR2 signaling contributes to hemorrhagic transformation, and glycyrrhizin could be a potential adjuvant therapy to attenuate hemorrhagic transformation, possibly through inhibiting the ONOO-/HMGB1/TLR2 signaling cascades.

Baicalin can scavenge peroxynitrite and ameliorate endogenous peroxynitrite-mediated neurotoxicity in cerebral ischemia-reperfusion injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Baicalin is one of the principal flavonoids isolated from the dried root of Scutellaria baicalensis Georgi that has long been used to treat ischemic stroke. However, its neuroprotective mechanisms against cerebral ischemia injury are poorly understood. AIM OF THE STUDY: To explore the neuroprotective mechanisms of baicalin against cerebral ischemia reperfusion injury. MATERIAL AND METHODS: In chemical systems, we conducted electron paramagnetic resonance (EPR) spin trapping experiments to evaluate the scavenging effects of baicalin on superoxide and nitric oxide, and mass spectrometry (MS) studies on the reaction of baicalin and peroxynitrite. In cellular experiments, we investigated the effects of baicalin against extraneous and endogenous peroxynitrite mediated neurotoxicity in SH-SY5Y cells treated with peroxynitrite donor, synthesized peroxynitrite and exposed to oxygen glucose deprivation and reoxygenation (OGD/RO) in vitro. Moreover, we studied the neuroprotective effects of baicalin by using a rat model of middle cerebral artery occlusion in vivo. FeTMPyP, a peroxynitrite decomposition catalyst, was used as positive control. Cell viability and apoptotic cell death was accessed by MTT assay and TUNEL assay respectively; 3-nitrotyrosine formation and infarction volume were detected by immunostaining experiments and TTC staining respectively. RESULTS: Baicalin revealed strong antioxidant ability by directly scavenging superoxide and reacting with peroxynitrite. Baicalin protected the neuronal cells from extraneous and endogenous peroxynitrite-induced neurotoxicity. In ischemia-reperfused brains, baicalin inhibited the formation of 3-nitrotyrosine, reduced infarct size and attenuated apoptotic cell death, whose effects were similar to FeTMPyP. CONCLUSIONS: Baicalin can directly scavenge peroxynitrite and the peroxynitrite-scavenging ability contributes to its neuroprotective mechanisms against cerebral ischemia reperfusion injury.

Effect of fluoride exposure on bone metabolism indicators ALP, BALP, and BGP.

OBJECTIVE: To analyze the changes in serum alkaline phosphatase (ALP) and bone alkaline phosphatase (BALP) activity and changes in osteocalcin (BGP) content following fluoride exposure and, thereby, determine the reference indications of fluoride-induced changes in bone metabolism. METHODS: In the animal study, rats were allowed free access to drinking water containing different concentrations (10, 150, or 400 mg/L) of sodium fluoride. Serum ALP and BALP activity and serum BGP content were assessed at three exposure time-points. In the spot study, serum ALP and BALP activity and serum BGP content were assessed in workers exposed to fluoride in their working environment for different periods of time. RESULTS: Compared with the control group, on days 15 and 30, the activity of serum ALP in the low- and medium-dose group was significantly higher (p < 0.05), while in the high-dose group it was significantly lower (p < 0.05). Only on day 30 was the activity of serum BALP in the medium-dose group significantly higher than that of the control group (p < 0.05). BGP content was lower in the high-dose group than in the control group (p < 0.05) on days 30 and 90, but it was higher in the medium-dose group on day 90. Compared with the control group, BGP content in the fluoride-exposed group was higher (p < 0.05). In the spot study, serum ALP activity and serum BGP content in the medium working-age group were higher than that in the short working-age group (p < 0.05). However, serum ALP activity and serum BGP content were lower in the long working-age group than in the medium working-age group (p < 0.05). CONCLUSIONS: Our results suggest that serum fluoride and urinary fluoride can be used as reference indications to provide an overall reflection of the body's fluoride-load and fluoride exposure level. Serum ALP activity and serum BGP content can be used as important reference indications for diagnosing bone metabolism changes resulting from fluoride exposure.

[Effect of Kangxianling Decoction on expression of hepatocyte growth factor mRNA and phosphorylations of extracellular signal-regulated protein kinase 1/2 and p38 in renal tissue of rats with unilateral ureteral obstruction].

OBJECTIVE: To study the effect of Kangxianling Decoction (KXLD), a compound traditional Chinese herbal medicine, on expression of hepatocyte growth factor (HGF) mRNA and phosphorylations of extracellular signal-regulated protein kinase 1/2 (ERK 1/2) and p38 in renal tissue of rats with unilateral ureteral obstruction (UUO). METHODS: Eighteen male SD rats were randomly divided into 3 groups: sham-operated group, untreated group and KXLD-treated group. A rat model of renal interstitial fibrosis was established by UUO. Rats with UUO were sacrificed after intragastric administration of KXLD for 14 days, and the parameters such as serum creatinine (SCr), blood urea nitrogen (BUN) and hydroxyproline in the kidney of rats in 3 groups were analyzed. The expression of HGF mRNA in kidney tissue was determined by reverse transcription polymerase chain reaction. The expressions of c-Met protein, ERK1/2 protein, p38 protein and the phosphorylations of ERK1/2 and p38 were determined by Western blotting method. RESULTS: The levels of SCr, BUN and hydroxyproline in the untreated group were significantly increased as compared with those in the sham-operated group (P<0.05). The expression of HGF mRNA in the untreated group was significantly down-regulated. The expression of c-Met protein and the phosphorylations of ERK1/2 and p38 in the kidney tissue of rats with UUO in the untreated group were significantly up-regulated. After intervention with KXLD, the phosphorylations of ERK1/2 and p38 were all significantly inhibited except for c-Met expression. The HGF mRNA was increased in KXLD-treated group. CONCLUSION: KXLD can decrease the level of collagen in the obstructed kidney of rats with UUO and alleviate the renal interstitial fibrosis in rats with UUO through enhancing the HGF mRNA expression and inhibiting the phosphorylations of ERK1/2 and p38.

[The effects of Tongxinluo Supermicro Powder on nuclear factor-kappaB, intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 expression in aorta of rabbits fed with high-lipid diet].

OBJECTIVE: To observe the effects of Tongxinluo Supermicro Powder on the nuclear factor-kappaB (NF-kappaB), inter-cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expression in aorta of rabbits fed with high-lipid diet. METHODS: Healthy male New Zealand rabbits were randomly divided into 4 groups (n = 8 each): control group, model group, atorvastatin group (3 mg x kg(-1) x d(-1) per gavage), and Tongxinluo group (0.31 g x kg(-1) x d(-1) per gavage). At the end of 6 weeks, the expression of NF-kappaB, ICAM-1 and VCAM-1 were observed by immunochemistry methods, Western blotting and reverse transcription polymerase chain reaction (RT-PCR). RESULT: The nuclear translocation of NF-kappaB in aortic endothelial cells and the gene expressions of NF-kappaB, ICAM-1 and VCAM-1 at protein and mRNA levels of the model group was significantly increased compared that in the control group (all P < 0.05), these effects could be significantly attenuated by atorvastatin and Tongxinluo Supermicro Powder (P < 0.01 vs. model group). CONCLUSIONS: Similar as atorvastatin, Tongxinluo Supermicro Powder could relieve the process of atherosclerosis by decreasing the nuclear translocation of NF-kappaB and reducing the expression of ICAM-1, VCAM-1 in this model.

[Effect of Kangxianling on Smads signal pathway molecules in rats with adriamycin induced nephropathy].

OBJECTIVE: To study the effect and mechanism of Kangxianling (KXL, a Chinese compound recipe) in treating adriamycin (ADR) induced renal fibrosis rats. METHODS: Forty-five male SD rats were randomly divided into 4 groups, the normal group (n = 7), the sham operative group (n = 8), the model group (n = 15), and the treatment group (n = 15). Model of renal interstitial fibrosis was established in rats by unilateral nephrectomy and intravenous injection of ADR twice at a 30-day interval, and the rats in the treatment group treated with KXL once a day for 72 days. Body weight, serum creatinine (SCr), blood urea nitrogen (BUN) levels and endogenous creatinine clearance rate (CCr) of animals were analyzed at the end of the 4th and the 8th week after operation. Rats were sacrificed after 72 days of treatment and their kidney obtained for pathological examination with HE and PASM staining. And protein expression levels of transforming growth factor beta (TGF-beta) receptor I (TbetaR I), TGF-beta receptor II (TbetaR II), Smad2 and Smad7 were determined by Western blotting. RESULTS: Levels of SCr and BUN in animals of the model group were significantly higher and CCr lower than those in the normal group (P < 0.05). Pathological examination of kidney in the model group showed thickened glomerular/tubular basement membrane with segmental sclerosis and hyaline degeneration; atrophy of the renal tubule around the sclerotic glomeruli and part of them disappeared; hypertrophy of partial glomeruli with surrounding severe dilated tubules; obvious glomeruli centering phenomena; severe tubular epithelial cell degeneration, necrosis with protein cast; fibrous tissue proliferation and large amount of inflammatory cell interstitial infiltration. The protein expression of TbetaR I and Smad2 in kidney tissue of the model group were significantly up-regulated, while that of TbetaR II and Smad7 unchanged. After KXL intervention, level of BUN lowered, SCr tended to normal and the endogenous SCr was raised to some degree. The renal pathological status in the treatment group was significantly improved and with markedly lowering of TbetaR I and Smad2 protein expression. CONCLUSION: KXL could inhibit the protein expression of TbetaR I and Smad2 in kidney tissue, so as to alleviate the renal fibrosis induced by adriamycin and improve the renal function.

Static magnetic fields neither prevent nor diminish symptoms and signs of delayed onset muscle soreness.

OBJECTIVE: To determine whether application of a commercially available static magnetic field would alter the signs and/or symptoms of delayed onset muscle soreness (DOMS) produced by exhaustive eccentric exercise. DESIGN: A double-blinded, randomized, and placebo-controlled study, with subjects serving as their own controls. SETTING: An outpatient physical therapy and performance center. PARTICIPANTS: Twenty-three healthy volunteers (18 women; mean age, 30 y; range, 18-40 y; 5 men; mean age, 29 y; range, 19-39 y). INTERVENTION: After exhaustive eccentric exercise of both the right and left elbow flexor muscle groups, subjects received daily treatment with either a 350G magnet or a placebo device for 5 consecutive days. MAIN OUTCOME MEASURES: Outcome variables, including anthropometric measurements, perceived discomfort, and muscle force production, were compared using linear mixed models. RESULTS: Arm circumference, relaxed elbow flexion angle, and pain increased, whereas active elbow flexion angle and maximal isometric torque decreased transiently before returning to near baseline. No significant difference in outcome variables existed between the treated and control arms. Participants reported less pain in both treated and control arms after each session, suggesting a placebo effect. CONCLUSIONS: Static magnetic fields were no more effective than placebo in preventing DOMS.

Echinacea purpurea therapy for the treatment of the common cold: a randomized, double-blind, placebo-controlled clinical trial.

BACKGROUND: Echinacea purpurea stimulates the immune response and is promoted to reduce symptom severity and the duration of upper respiratory tract infections. We sought to determine the efficacy of a standardized preparation of E purpurea in reducing symptom severity and duration of the common cold. METHODS: A randomized, double-blind, placebo-controlled design was used. Patients received either 100 mg of E purpurea (freeze-dried pressed juice from the aerial portion of the plant) or a lactose placebo 3 times daily until cold symptoms were relieved or until the end of 14 days, whichever came first. Symptoms (sneezing, nasal discharge, nasal congestion, headache, sore or scratchy throat, hoarseness, muscle aches, and cough) were scored subjectively by the patient and recorded daily in a diary. Kaplan-Meier curves were used to estimate the survival function of time to resolution in each group. The Wilcoxon rank sum test was used to compare time to resolution between the 2 groups. RESULTS: One hundred twenty-eight patients were enrolled within 24 hours of cold symptom onset. Group demographic distribution was comparable for sex, age, time from symptom onset to enrollment in the study, average number of colds per year, and smoking history. No statistically significant difference was observed between treatment groups for either total symptom scores (P range,.29-.90) or mean individual symptom scores (P range,.09-.93). The time to resolution of symptoms was not statistically different (P =.73). CONCLUSIONS: Some studies have concluded that Echinacea effectively reduces the symptoms and duration of the common cold. We were unable to replicate such findings. Further studies using different preparations and dosages of E purpurea are necessary to validate previous claims.

[Study on metabolism of preventive anti-fluorine agent on rats].

In order to study the metabolism of preventive anti-fluorine agent, 30 rats are randomly divided into high-dose, low-dose and a control groups. The high dose (400 mg/kg.d) and low dose (16 mg/kg.d) are orally administrated respectively, and the content of boron and/or zinc in urine, dung, serum, bone, liver, muscles, brain tissues is determined. The results showed that during the administration of this agent, the content of boron in urine and the content of zinc in dung increases obviously in both high-dose and low-dose groups and their discharge rate is consistent with the dose given. The content of boron and zinc in bone, liver, and zinc in serum, muscles, brain tissues increases evidently compared to that in control group but decreases rapidly after administration of the agent. The findings revealed that there is a rapid metabolism of boron and zinc in the body of rats. The highest content of the agent is observed in bones. The content ranks second in liver and muscles but no accumulative effects are observed.