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Background Accumulation of free fatty acids (FFAs) in hepatocytes is a hallmark of liver dysfunction and non-alcoholic fatty liver disease (NAFLD). Excessive deposition of FFAs alters lipid metabolism pathways increasing the oxidative stress and mitochondrial dysfunction. Attenuating hepatic lipid accumulation, oxidative stress, and improving mitochondrial function could provide potential targets in preventing progression of non-alcoholic fatty liver (NAFL) to non-alcoholic steatohepatitis (NASH). Earlier studies with Picrorhiza kurroa extract have shown reduction in hepatic damage and fatty acid infiltration in several experimental models and also clinically in viral hepatitis. Thus, the effect of P. kurroa's phytoactive, picroside II, needed mechanistic investigation in appropriate in vitro liver cell model. Objective(s) To study the effect of picroside II on FFAs accumulation, oxidative stress and mitochondrial function with silibinin as a positive control in in vitro NAFLD model. Materials and methods HepG2 cells were incubated with FFAs-1000μM in presence and absence of Picroside II-10 μM for 20 hours. Results HepG2 cells incubated with FFAs-1000μM lead to increased lipid accumulation. Picroside II-10μM attenuated FFAs-induced lipid accumulation (33%), loss of mitochondrial membrane potential (ΔΨm), ATP depletion, and production of reactive oxygen species (ROS). A concomitant increase in cytochrome C at transcription and protein levels was observed. An increase in expression of MnSOD, catalase, and higher levels of tGSH and GSH:GSSG ratios underlie the ROS salvaging activity of picroside II. Conclusion Picroside II significantly attenuated FFAs-induced-lipotoxicity. The reduction in ROS, increased antioxidant enzymes, and improvement in mitochondrial function underlie the mechanisms of action of picroside II. These findings suggest a need to develop an investigational drug profile of picroside II for NAFLD as a therapeutic strategy. This could be evaluated through the fast-track path of reverse pharmacology.

Picroside II (P‐II), one of the main active components of scrophularia extract, which have anti‐oxidative, anti‐inflammatory effects, but its effect on hyperhomocysteinemia (HHcy) induced endothelial injury remains to be determined. Here, we test whether P‐II protects HHcy‐induced endothelial dysfunction against oxidative stress, inflammation and cell apoptosis. In vitro study using HUVECs, and in hyperhomocysteinemia mouse models, we found that HHcy decreased endothelial SIRT1 expression and increased LOX‐1 expression, subsequently causing reactive oxygen species generation, up‐regulation of NADPH oxidase activity and NF‐κB activation, thereby promoting pro‐inflammatory response and cell apoptosis. Blockade of Sirt1 with Ex527 or siRNASIRT1 increased LOX‐1 expression, whereas overexpression of SIRT1 decreased LOX‐1 expression markedly. P‐II treatment significantly increased SIRT1 expression and reduced LOX‐1 expression, and protected against endothelial cells from Hcy‐induced oxidative injury, inflammation and apoptosis. However, blockade of SIRT1 or overexpression of LOX‐1 attenuated the therapeutic effects of P‐II. In conclusion, our results suggest that P‐II prevents the Hcy induced endothelial damage probably through regulating the SIRT1/LOX‐1 signaling pathway.

The inflammatory response is important in the pathogenesis of myocardial ischemia/reperfusion (I/R) injury. Picroside II, the primary active constituent of Picrorhizae, has been reported to protect the myocardium from I/R-induced injury, however, the exact mechanism underlying these protective effects remains unclear. The aim of the present study was to investigate the mechanism underlying the protective effects of picroside II on I/R-induced myocardial injury. Adult male Sprague-Dawley rats underwent 1 h left coronary artery occlusion followed by 3 h reperfusion. Picroside II was administered (10 mg/kg) via the tail vein 30 min prior to left coronary artery occlusion. The results revealed that pretreatment of picroside II could significantly alleviate I/R-induced myocardial injury concomitantly with a decrease in inflammatory factor production. In addition, picroside II was also able to decrease high mobility group box 1 (HMGB1) expression, and release and downregulate the expression of the receptor for advanced glycation end products (RAGE), toll-like receptor (TLR)-2 and TLR-4. Furthermore, picroside II was able to inhibit nuclear factor-κB (NF-κB) activation. The results indicated that the protective effect of picroside II on I/R-induced myocardial injury was associated, at least partly, with inhibition of the inflammatory response by suppressing the HMGB1-RAGE/TLR-2/TLR-4-NF-κB signaling pathway.

Chronic obstructive pulmonary disease (COPD) is a major inflammatory lung disease characterized by irreversible and progressive airflow obstruction. Although corticosteroids are often used to reduce inflammation, steroid therapies are insufficient in patients with refractory COPD. Both serum amyloid A (SAA) and IL-33 have been implicated in the pathology of steroid-resistant lung inflammation. Picroside II isolated from Pseudolysimachion rotundum var. subintegrum (Plantaginaceae) is a major bioactive component of YPL-001, which has completed phase-2a clinical trials in chronic obstructive pulmonary disease patients. In this study, we investigated whether picroside II is effective in treating steroid refractory lung inflammation via the inhibition of the SAA-IL-33 axis. Picroside II inhibited LPS-induced SAA1 expression in human monocytes, which are resistant to steroids. SAA induced the secretion of IL-33 without involving cell necrosis. Picroside II, but not dexamethasone effectively inhibited SAA-induced IL-33 expression and secretion. The inhibitory effect by picroside II was mediated by suppressing the mitogen-activated protein kinase (MAPK) p38, ERK1/2, and nuclear factor-&kappa, B pathways. Our results suggest that picroside II negatively modulates the SAA-IL-33 axis that has been implicated in steroid-resistant lung inflammation. These findings provide valuable information for the development of picroside II as an alternative therapeutic agent against steroid refractory lung inflammation in COPD.

Picrorhiza kurroa, the dried rhizome of Picrorhiza kurroa Royle ex Benth, is a famous Chinese herb that has been traditionally used in China. Picroside II (PII), a glycoside derivative, is the main bioactive constituent of Picrorhiza kurroa. In the past several decades, bioactive components from Picrorhiza kurroa have attracted the attention of researchers due to their promising therapeutic effects. A large number of studies have demonstrated the therapeutic potential of PII for the prevention and treatment of some diseases, such as organic ischemia/reperfusion (I/R) injury, liver damage, inflammation, cancer metastasis and angiogenesis. In the present paper, we aimed to provide an overview of the pharmacology of PII, focusing on its anti-oxidant, anti-inflammatory and anti-apoptotic activities. Meanwhile, the plant tissue distribution and pharmacokinetic properties were also described. Due to its beneficial pharmacological effects in I/R injury, PII may serve as a promising therapeutic agent for organic I/R injury prevention.

Picroside II possesses a wide range of pharmacological effects and has been demonstrated to ameliorate cerebral ischemia and reperfusion (I/R) injury. However, its effects on renal I/R injury remain unclear. In the present study, the role of picroside II in attenuating oxidative stress and the inflammatory response in a rat model of renal I/R injury was investigated. Sprague Dawley rats were subjected to 45 min of ischemia followed by 24 h of reperfusion. Prior to reperfusion, the rats were treated with picroside II or an equal volume of phosphate-buffered saline. Renal function and histological changes were compared and the relevant parameters of oxidative stress and inflammation were detected. The expression of toll-like receptor 4 (TLR4) and nuclear factor κB (NF-κB; p65) were assessed by immunohistochemistry and western blotting. It was observed that renal function was significantly improved by treatment with picroside II. Morphological analysis indicated that picroside II clearly reduced tissue damage and the expression of TLR4 and NF-κB. Reverse transcription-quantitative polymerase chain reaction demonstrated that picroside II inhibited the increase of tumor necrosis factor (TNF)-α, interleukin (IL)-1β and intercellular adhesion molecule (ICAM)-1 expression induced by I/R injury. Western blot analysis indicated that the expression levels of TLR4 and NF-κB were significantly downregulated in the picroside II group compared with those in the I/R group. These results indicate that picroside II treatment suppressed the TLR4/NF-κB signaling pathway, protecting renal tissue against I/R-induced oxidative stress and inflammatory response.

Objective: Diabates mellitus, is a chronic metabolic disorder accompanied by an increase in oxidative stress. Ischemia-reperfusion injury is a cascade of events initiated by tissue ischemia. The cellular damage produced by reperfusion leads to an active inflammatory response. This study was performed to investigate the effect of picroside II on myocardial ischemiareperfusion injury in rats with streptozotocin-induced diabetes. Methods: Animals were equally (n:6) divided for five groups as follows; Control (C), diabetes [D], diabetes+picroside II [DP], diabetes+I/R [DIR], and diabetes+I/R+ picroside II [DIRP]. In DIR group, a left anterior descending artery branch was occluded for 60 minutes, the reperfused for 120 minutes. In DIRP group, picroside II was administrated via 10 mg/kg intraperitoneal route 30 minutes before ligating the left anterior descending artery. At the end of the study, myocardial tissues were taken for total oxidant status and total antioxidant status level determinations. Results: Total oxidant status levels were significantly higher in DIR group, when compared with C, DP, and DIRP groups (p:0.001, p:0.019, and p:0.031, respectively). Total antioxidant status levels were significantly higher in DIR group, when compared with C, DP, and DIRP groups (p:0.006, p:0.024, and p:0.007, respectively). Conclusion: These results indicate that administration of picroside II may have protective effects against I/R injury., Amaç: Diyabetes mellitus, oksidatif stres artışının eşlik ettiği kronik metabolik bir hastalıktır. İskemi-reperfüzyon (İ/R) hasarı doku iskemisi tarafından başlatılan olayların bir kaskadıdır. Reperfüzyon sonucunda oluşan hücre hasarı inflamatuar yanıtı aktive eder. Bu çalışma, Streptozosin kaynaklı diyabeti olan ratlarda miyokard İ/R hasarı üzerine Pikrozid II’nin etkisini araştırmak amacıyla yapıldı. Yöntem: Hayvanlar, beş gruba (n=6) olacak şekilde ayrıldı; Kontrol grubu (K), Diyabet grubu [D], Diyabet + Pikrozid II grubu [DP], Diyabet + İ/R grubu [DİR] ve Diyabet + İ/R + Pikrozid II grubu [DİRP]. DİR grubunda, sol ön inen arter dalı iskemi amaçlı 60 dakika süre ile kapatılmış ardından reperfüzyon için akım sağlanarak 120 dakika beklenmiştir. DİRP grubuna sol ön inen arter dalı kapatılmadan 30 dakika önce Pikrozid II, 10 mg/kg olmak üzere intraperitoneal olarak verildi. Çalışmanın sonunda, miyokard doku örnekleri total oksidan durum ve total antioksidan durum seviyesinin ölçülmesi için alındı. Bulgular: Toplam oksidan durum seviyeleri DİR grubunda, diğer gruplarla karşılaştırıldığında (K, DP, DİRP) anlamlı yüksek bulunmuştur (sırasıyla p:0.001, p:0.019, ve p:0.031). Total antioksidan durum seviyeleri DİR grubunda, diğer gruplarla karşılaştırıldığında (K, DP, DİRP ) anlamlı olarak yüksek bulunmuştur (sırasıyla p:0.006, p:0.024, ve p:0.007). Sonuç: Bu bulgular Pikrozid II’nin İ/R hasarına karşı koruyucu etkiye sahip olabileceğini göstermektedir.

Yiğit Kılıç,1 Abdullah Özer,1 Tolga Tatar,1 Mustafa Hakan Zor,1 Mehmet Kirişçi,2 Hakan Kartal,3 Ali Doğan Dursun,4 Deniz Billur,5 Mustafa Arslan,6 Ayşegül Küçük7 1Department of Cardiovascular Surgery, Gazi University Medical Faculty, Ankara, 2Department of Cardiovascular Surgery, Kahramanmaras Sutcu Imam Medical Faculty, Kahramanmaras, 3Department of Cardiovascular Surgery, Ardahan State Hospital, Ardahan, 4Department of Physiology, Ankara University Medical Faculty, 5Department of Histology and Embryology, Ankara University Medical Faculty, 6Department of Anaesthesiology and Reanimation, Gazi University Medical Faculty, Ankara, 7Department of Physiology, Dumlupinar University Medical Faculty, Kütahya, Turkey Introduction: Many structural and functional damages are observed in cells and tissues after reperfusion of previously viable ischemic tissues. Acute ischemia reperfusion (I/R) injury of lower extremities occurs especially when a temporary cross-clamp is applied to the abdominal aorta during aortic surgery. Research regarding the treatment of I/R injury has been increasing day-by-day. In this study, we aimed to investigate the effect of picroside II on skeletal muscle of rats experiencing simulated I/R.Materials and methods: Twenty-four male Wistar albino rats weighing between 210 and 300g were used in this study. Rats were randomly divided into 4 groups of 6 rats each (control, I/R, control + picroside II, and I/R+picroside II). The infrarenal section of the abdominal aorta was occluded with an atraumatic microvascular clamp in I/R group. The clamp was removed after 120 minutes and reperfusion was provided for a further 120 minutes. Picroside II (10 mgkg–1) was administered intraperitoneally to the animals in control+picroside II and I/R+picroside II groups. At the end of the study, skeletal muscle tissue was obtained for the determination of total oxidant status (TOS) and total antioxidant status (TAS) levels. Apoptosis was evaluated by TUNEL experiment.Results: TOS levels were significantly higher in I/R group than that of control and I/R+picroside II groups (P=0.014, P=0.005, respectively). TAS levels were significantly higher in I/R group than that of control and I/R+picroside II groups (P=0.007 P=0.005, respectively). TUNEL assay revealed that picroside II reduced cell necrosis.Conclusion: The results of this study demonstrated that picroside II plays a critical role to prevent I/R injury. Even though our results were found to be satisfactory, it should be encouraging to those who want to conduct future research on this topic. Keywords: ischemia reperfusion, picroside II, hind limb skeletal muscle, TOS, TAS

Picroside II could reduce the oxidative damage induced by cerebral ischemia-reperfusion and improve nerve behavior function of rats. The aim of this study was to optimize the therapeutic dose and time window of picrosede II by orthogonal test in cerebral ischemic injury in rats. The forebrain ischemia models were established by bilateral common carotid artery occlusion (BCCAO) methods. The successful models were randomly grouped according to orthogonal experimental design and treated by injecting picroside II intraperitonenally at different ischemic time with different dose. The concentration nitric oxide (NO) in serum and brain tissue were determined by nitratase reductase assay. The activity of glutathione peroxidase (GSHPx) in serum and tissue were determined by chemical colormetry. The optimized composition of the therapeutic dose and time window of picroside II in cerebral ischemic injury was (1) ischemia 1.5h with 20mg/kg body weight according to the concentrations of NO in serum and brain tissue; (2) ischemia 1.5h with 10mg/kg and ischemia 2.0h with 20mg/kg body weight according to the activities of GSHPx in serum and brain tissue. From the principle of lowest therapeutic dose with longest time window, the optimized composition of the therapeutic dose and time window of picroside II in cerebral ischemic injury is ischemia 1.5- 2.0h with 10-20mg/kg body weight.

The aim is to optimize the anti-inflammatory effect and the therapeutic dose and time window of picrosede II by orthogonal test in cerebral ischemic injury in rats. The forebrain ischemia models were established by bilateral common carotid artery occlusion (BCCAO) methods in 30 Wistar rats. The successful models were randomly divided into sixteen groups according to orthogonal experimental design and treated by injecting picroside II intraperitoneally at different ischemic time with different dose. The concentrations of aquaporins 4 (AQP4), matrix metalloproteinases9 (MMP9) and cyclooxygenase 2 (COX2) in serum and brain tissue were determined by enzyme linked immunosorbent assay to evaluate the therapeutic effect of picroside II in cerebral ischemic injury. The best therapeutic time window and dose of picroside II in cerebral ischemic injury were 1) ischemia 2.0 h with 20 mg/kg and 1.5 h with 20 mg/kg body weight according to the concentration of AQP4 in serum and brain tissue; 2) ischemia 1.5 h with 20 mg/kg and ischemia 2.0 h with 20 mg/kg according to the concentrations of MMP9 in serum and brain tissue; and 3) ischemia 1.5 h with 10 mg/kg and ischemia 1.5 h with 20 mg/kg according to the concentrations of COX2 in serum and brain tissue respectively. According to the principle of the lowest therapeutic dose with the longest time window, the optimized therapeutic dose and time window were injecting picroside II intraperitoneally with 10 - 20 mg/kg body weight at ischemia 1.5 - 2.0 h in cerebral ischemic injury.

The letter highlights the difference in the pharmacokinetic profile of picroside II in Sprague-Dawley rats as reported by Upadhyay et al., in two separate studies. Both the studies have been conducted by the same author, in the same laboratory set-up, using the similar animal species and sex as well as the same dose level of kutkin (Dose: 100 mg/kg bodyweight, which was equivalent to 45 mg/kg of picroside I and 55 mg/kg of picroside II). However, the C max and AUC 0-t observed in the first study were 9-fold and 5-fold higher, respectively, as compared to the recently reported study. The only difference between these two studies was the instrument used for analysing the plasma samples. In the first study, the analyses of the plasma samples were done using HPLC-UV whereas the second study used LC-ESI-MS system for plasma sample analysis. The pharmacokinetic parameters shall not change significantly with the change in analytical instrument. In my opinion, the probable cause for the observed higher C max and AUC 0-t values in the first study may be due to the interference of some metabolite(s)/impurity that got eluted at the same retention time as that of picroside II and contributed to the higher values. The future prospects shall focus on identifying the metabolite(s)/impurity that have co-eluted with picroside II during initial HPLC-UV analysis and contributed to the higher C max and AUC 0-t values by performing parallel analysis of plasma samples using HPLC-PDA and LC-ESI-MS.

PicrosideⅡ, one of the most effective components extracted from picrorhiza, has been reported with neuronal protective activity. To isolate the picroside Ⅱ and similar structure moleculars from picrorhiza effectively, molecularly imprinted polymer (MIP) was prepared by precipitation polymerization with Picroside Ⅱ as template molecule and TRIM as cross-linkers. The MIP was with high affinity and excellent stereo-selectivity toward Picroside Ⅱ and similar structure molecular. The purity of picroside is 70% of target sample. This research might offer a novel method for the enrichment and determination of target compounds from the Traditional Chinese Medicine. A model of middle cerebral artery occlusion and reperfusion (MCAO/R) in rats was used to determine the protective effects of Picroside II sample of this experiment on cerebral ischaemia/reperfusion in vivo. Results indicate the picroside II sample can protect neurons against acute ischemic stroke injury, and the neurological deficit scores of dosage group is lower than that of model group.

Picroside-I and picroside-II are known bioactive metabolites in Picrorhiza species. In the present study a simple, precise method has been established for the simultaneous determination of picrosides (picroside-I and picroside-II) in two different Picrorhiza species, P. kurroa and P. scrophulariiflora. This method was also validated for accuracy, precision, robustness, limit of detection and quantification, repeatability and recovery, according to International Conference of Harmonization guidelines. Separation and quantification was achieved by HPTLC using as the mobile phase chloroform–methanol (88:12, v/v) on precoated silica gel 60F254 aluminum plates. Densitometric determination was carried out at wavelength λmax 254 nm in UV absorbance mode. Comparative study also revealed that picroside-I and picroside-II are higher in P. scrophulariiflora than P. kurroa. Picroside-I content was found to be 1.258 and 1.611%, and picroside-II was estimated as 0.481 and 0.613% in P. kurroa and P. scrophulariiflora, respectively. Antioxidant potential of these two Picrorhiza species was also studied using DPPH. At a concentration of 0.1 mg/mL the scavenging activities of P. kurroa and P. scrophulariiflora were found to 37.70 and 34.30%, respectively. Copyright © 2011 John Wiley & Sons, Ltd.

The aim of this study was to explore the effect of picroside II on neuronal apoptosis and the expression of caspase-3 and poly ADP-ribose polymerase (PARP) following middle cerebral artery occlusion/reperfusion in male Wistar rats. Picroside II (10 mg/kg) was administered intravenously into the tail vein of the animals. The neurological function deficits were evaluated with the Bederson’s test and the cerebral infarction volume was visualized with tetrazolium chloride (TTC) staining. The apoptotic cells were counted by in situ terminal deoxynucleotidyl transferase-mediated biotinylated deoxyuridine triphosphate nick end labeling (TUNEL) assay. The immunohistochemistry stain and enzyme linked immunosorbent assay (ELISA) was used to determine the expressions of caspase-3 and PARP in brain tissue. The results indicated that rats in the control group showed neurological function deficit and cerebral infarction in ischemic hemisphere after two hours ischemia followed by 22 hours reperfusion. Caspase-3 and PARP expressions were also profound in the cortex, the striatum and the hippocampus, along with increased apoptotic cells in this group. Bederson's score, infarction volume, and expressions of caspase-3 and PARP, as well as apoptosis in the treatment group were, however, significantly decreased compared to those in the control group indicating that intravenous treatment with picroside II might be beneficial to inhibit neuronal apoptosis and, thus, to improve the neurological function of rats upon cerebral ischemia reperfusion injury.

Renal ischemia and reperfusion (I/R) injury, which commonly occurs in kidney transplantation, is the leading cause of acute kidney injury. Picroside II possesses a wide range of pharmacological effects, including anti-apoptosis effects. In the present study, the ability of picroside II to attenuate apoptosis in a rat model of renal I/R injury was investigated. Sprague-Dawley rats were subjected to 45 min of ischemia followed by 24 h of reperfusion. Prior to reperfusion, the rats were treated with picroside II or an equal volume of phosphate-buffered saline. It was observed that renal function was significantly improved by the treatment with picroside II. Morphological analysis indicated that picroside II markedly reduced tissue damage and the expression of cleaved caspase-3. Reverse transcription-quantitative polymerase chain reaction and western blotting revealed that the expression levels of Bax and poly(ADP-ribose) polymerase-1 (PARP-1) were upregulated in the I/R group, whereas those of Bcl-2 were downregulated. However, the treatment with picroside II inhibited these changes induced by renal I/R injury. In conclusion, picroside II has potent anti-apoptotic activity against renal I/R injury.

Background To explore the neuroprotective effect and optimize the therapeutic dose and time window of picroside II by orthogonal test and the expression of myelin basic protein (MBP) in cerebral ischemic injury in rats. Bilateral common carotid artery occlusion (BCCAO) was used to establish forebrain ischemia models. The successful rat models were grouped according to orthogonal experimental design and injected picroside II intraperitoneally at different ischemic time with different doses. Myelin sheath fast green staining(FGS) and transmission electron microscopy (TEM) were used to observe nerve fiber myelin; the expression of MBP was tested qualitatively and quantitatively by immunohistochemical assay (IHC) and Western blot (WB); Reverse transcription polymerase chain reaction (RT-PCR) was used to detect the transcription level of MBP mRNA. Results The protective effect of picroside II was presented by increasing the expression of MBP and decreasing demyelination after cerebral ischemic injury. The best therapeutic time window and dose was (1) ischemia 2.0 h with picroside II 10 mg/kg body weight according to the results of FGS, IHC and WB; (2) ischemia 1.5 h with picroside II 20 mg/kg according to the analysis of RT-PCR. Conclusion Given the principle of the longest time window and the lowest therapeutic dose, the optimized therapeutic dose and time window should be injecting picroside II intraperitoneally with 10-20 mg/kg body weight at ischemia 1.5-2.0 h in cerebral ischemic injury.

Faruk Metin Çomu,1 Yiğit Kılıç,2 Abdullah Özer,2 Mehmet Kirişçi,3 Ali Doğan Dursun,4 Tolga Tatar,2 Mustafa Hakan Zor,2 Hakan Kartal,2 Ayşegül Küçük,5 Hakan Boyunağa,6 Mustafa Arslan71Department of Physiology, Kirikkale University Medical Faculty, Kirikkale, 2Department of Cardiovascular Surgery, Gazi University Medical Faculty, Ankara, 3Department of Cardiovascular Surgery, Necip Fazil State Hospital, Kahramanmaras, 4Department of Physiology, Ankara University Medical Faculty, Ankara, 5Department of Physiology, Dumlupinar University Medical Faculty, Kütahya, 6Department of Biochemistry, Kirikkale University Medical Faculty, Kirikkale, 7Department of Anaesthesiology and Reanimation, Gazi University Medical Faculty, Ankara, TurkeyBackground: Ischemia reperfusion injury (I/R) in hind limb is a frequent and important clinical phenomenon. Many structural and functional damages are observed in cells and tissues in these kinds of injuries. In this study, we aimed to evaluate the effect of picroside II on lipid peroxidation and erythrocyte deformability during I/R in rats.Methods: Rats were randomly divided into four groups – each containing six animals (sham, I/R, sham + picroside II, and I/R + picroside II). The infrarenal section of the abdominal aorta was occluded with an atraumatic microvascular clamp in I/R groups. The clamp was removed after 120minutes and reperfusion was provided for a further 120 minutes. Picroside II (10 mg·kg-1) was administered intraperitoneally to the animals in the appropriate groups (sham + picroside II, I/R + picroside II groups). All rats were euthanized by intraperitoneal administration of ketamine (100 mg·kg-1) and taking blood from the abdominal aorta. Erythrocytes were extracted from heparinized complete blood samples. Buffer (PT) and then erythrocytes (PE) were passed through the filtration system and the changes in pressure were measured to investigate the role of serum malondialdehyde and nitric oxide (NO) in lipid peroxidation and erythrocyte deformability index.Results: Deformability index was significantly increased in the I/R group compared to groups sham, sham + picroside-II, and I/R + picroside-II (P

This study compared the hepatic glucuronidation of Picroside II in different species and characterized the glucuronidation activities of human intestinal microsomes (HIMs) and recombinant human UDP-glucuronosyltransferases (UGTs) for Picroside II. The rank order of hepatic microsomal glucuronidation activity of Picroside II was ratmousehumandog. The intrinsic clearance of Picroside II hepatic glucuronidation in rat, mouse and dog was about 10.6-, 6.0- and 2.3-fold of that in human, respectively. Among the 12 recombinant human UGTs, UGT1A7, UGT1A8, UGT1A9 and UGT1A10 catalyzed the glucuronidation. UGT1A10, which are expressed in extrahepatic tissues, showed the highest activity of Picroside II glucuronidation (K(m) = 45.1 μM, V(max) = 831.9 pmol/min/mg protein). UGT1A9 played a primary role in glucuronidation in human liver microsomes (HLM; K(m) = 81.3 μM, V(max) = 242.2 pmol/min/mg protein). In addition, both mycophenolic acid (substrate of UGT1A9) and emodin (substrate of UGT1A8 and UGT1A10) could inhibit the glucuronidation of Picroside II with the half maximal inhibitory concentration (IC(50)) values of 173.6 and 76.2 μM, respectively. Enzyme kinetics was also performed in HIMs. The K(m) value of Picroside II glucuronidation was close to that in recombinant human UGT1A10 (K(m) = 58.6 μM, V(max) = 721.4 pmol/min/mg protein). The intrinsic clearance was 5.4-fold of HLMs. Intestinal UGT enzymes play an important role in Picroside II glucuronidation in human.

A new iridoid, pikuroside (1), was isolated from the roots of Picrorhiza kurroa, together with three known iridoids, picroside I (2), picroside II (3), and 6-feruloyl catalpol (4). The structure of 1 was established by interpretation and full assignments of NMR spectral data. Pikuroside (1) had no antiinflammatory activity, although the crude extract and picroside II (3) demonstrated moderate activity.

Nan Wu, Wenna Li, Wenqi Shu, Dalin Jia Department of Cardiology, The First Affiliated Hospital of China Medical University, Shenyang, People's Republic of China Abstract: The aim of this study was to determine the effect of picroside II on myocardial ischemia reperfusion injury in rats and to explore its underlying mechanism. Isolated rat hearts underwent 30 minutes of global ischemia followed by 120 minutes of reperfusion. Different doses of picroside II (1 µM, 10 µM, and 100 µM) were given 20 minutes before ischemia. Phosphoinositide 3-kinase inhibitor (wortmannin) and nitric oxide synthase (NOS) inhibitor (L-NG-nitroarginine methyl ester) were given 10 minutes before picroside II treatment. The cardiac function, myocardial infarct size, apoptosis, myocardial nitric oxide content, the expressions of Bcl-2 and Bax, and the activation of the phosphoinositide 3-kinase/Akt/endothelial NOS pathway were evaluated. Treatment with 10 µM and 100 µM picroside II significantly improved postischemic myocardial function, reduced myocardial infarct size, inhibited apoptosis, increased myocardial NO content, upregulated Bcl-2, downregulated Bax, and increased the phosphorylation of Akt and endothelial NOS, but cardioprotection was not shown in the 1 µM picroside II treatment group and was abrogated by wortmannin and L-NG-nitroarginine methyl ester. Furthermore, cardioprotection in the 100 µM picroside II treatment group was superior to that in the 10 µM picroside II treatment group. In conclusion, the data reveals that picroside II has a significant protective effect on myocardial ischemia reperfusion injury in a dose-dependent manner, which was mediated by upregulating the phosphoinositide 3-kinase/Akt/endothelial NOS pathway to increase nitric oxide production and regulating the expressions of Bcl-2 and Bax to inhibit apoptosis. Keywords: picroside II, ischemia reperfusion injury, apoptosis, nitric oxide, endothelial nitric oxide synthase

A preliminary study from our research group showed that picroside II inhibited neuronal apoptosis in ischemic penumbra, reduced ischemic volume, and improved neurobehavioral function in rats with cerebral ischemia. The aim of the present study was to validate the neuroprotective effects of picroside II and optimize its therapeutic time window and dose in a rat model of cerebral ischemia. We found that picroside II inhibited cell apoptosis and reduced the expression of neuron-specific enolase, a marker of neuronal damage, in rats after cerebral ischemic injury. The optimal treatment time after ischemic injury and dose were determined, respectively, as follows: (1) 2.0 hours and 10 mg/kg according to the results of toluidine blue staining; (2) 1.5 hours and 10 mg/kg according to early apoptotic ratio by flow cytometry; (3) 2.0 hours and 10 mg/kg according to immunohistochemical and western blot analysis; and (4) 1.5 hours and 10 mg/kg according to reverse transcription polymerase chain reaction. The present findings suggest that an intraperitoneal injection of 10 mg/kg picroside II 1.5-2.0 hours after cerebral ischemic injury in rats is the optimal dose and time for therapeutic benefit.