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β-Glucuronidase-inhibitory and hepatoprotective effects of Reduohanxiao-tang (Yuldahanso-tang), which has been used for liver diseases and stroke, on carbon tetrachloride (CCl₄)-induced hepatotoxicity of rats were investigated. Reduohanxiao-tang potently inhibited β-glucuronidases. Serum aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactic acid dehydrogenase (LDH) levels of the CCl₄ group orally treated with Reduohanxiao-tang (100 mg/kg) were lowered to 54%, 71.5% and 66.1% of the CCl₄-treated control group, respectively. Among the ingredients of the Reduohanxiao-tang, the rhizomes of Pueraria thunbergiana and it Scutellaria baicalensis potently inhibited β-glucuronidases and protected against CCl₄-induced liver injury. Orally administered puerarin, which is a main component of Pueraria thunbergiana, showed potent hepatoprotective activity, but did not inhibit β-glucuronidase. However, daidzein, which is produced from puerarin by human intestinal bacteria, potently inhibited β-glucuronidase. These results suggest that β-glucuronidase inhibition by herbal medicines may protect a gainst CCl₄-induced liver injury.

In order to study the pharmacokinetics of puerarin and ginsenoside Rg1 of cerebral blood nutrition (CBN) and its relationship with pharmacodynamics of platelet aggregation induced by ADP in rat, the blood samples after injection were collected. The concentrations of puerarin and ginsenoside Rg1 in plasma were determined by RP-HPLC, and the platelet aggregations were observed simultaneously. The data showed that there was distinct statistic significance (p < 0.01) for puerarin processing, which was a single compartment model with quick elimination rate (t_1/2β = 18 min) and MRT (26 min), while ginsenoside Rg1 processing was a double compartment model with rapid distribution rate (t_1/2α = 8 min), slow elimination rate (t_1/2β = 11 hours) and MRT (3.3 hours). Effects of anti-platelet aggregation were presented at 5–10 min, 45–90 min and 6–8 hours after injection separately, and the corresponding concentrations of puerarin were 25–21 μg/ml, 4.5–0.8 μg/ml and 0 μg/ml, ginsenoside Rg1 were 7.6–6.7 μg/ml, 1.2–0.6 μg/ml and 1.8–0.5 μg/ml. The two components presented a positive correlation between their concentrations and the effect of anti-platelet aggregation in 5–10 min after CBN injection (coefficient of correlation were 0.999 and 0.995). However, it was noted that the effect was still stronger even when concentrations of puerarin and ginsenoside Rg1 in plasma decreased. Therefore, puerarin and ginsenoside Rg1 not only had different pharmacokinetics, but also had a positive correlation with platelet aggregation, just in 5–10 min after CBN injection.

In order to investigate the mechanism of the therapeutic effect of puerarin on non-alcoholic fatty liver disease, a non-alcoholic fatty disease male rat model was induced by a high fat diet, all rats were randomly divided into a blank group, model group, simavastatin group and puerarin group. After 4 weeks of drug treatment, the liver was slided to investigate pathological morphology. Elisa was used to measure the total cholesterol (TC), triglyeride (TG) in liver, and leptin content in serum. RT-PCR and Western blotting were employed to detect liver leptin mRNA receptor expression and P-JAK2, P-STAT3 expression levels in the liver respectively. The results showed that puerarin significantly decreased the TG, TC content in liver of the non-alcoholic fatty disease rats, ameliorated steatosis in liver, lowered liver inflammatory reaction, decreased leptin level in serum, and enhanced the expression of leptin receptor mRNA and P-JAK2/P-STAT3 level. All the results demonstrated that puerarin can exhibit therapeutic effect on non-alcoholic fatty liver disease by improving leptin signal transduction through JAK2/STAT3 pathways.

To provide experimental and theoretical basis for the clinical application of Puerarin in acute alcohol poisoning, 30 Wistar rats were randomized into 3 groups as follows: (1) Group A (control) underwent normal sodium (N.S.) peritoneal injection (i.p.) and intragastric administration (i.g.); (2) Group B (alcohol) underwent an equivalent dosage of N.S. i.p. and 40% ethanol (8000 mg/kg. d).ig for 5 days; (3) Group C (Puerarin) underwent Puerarin 200 mg/kg. d. ip, and an equivalent dosage of ethanol for 5 days. The left lobes of livers were sampled, and the levels of MDA, SOD and GPX in plasma and liver homogenate were detected. The level of MDA in plasma and liver homogenate in the alcohol group was obviously higher than that in the control group (p < 0.05, respectively), while that in the Puerarin group was significantly lower than in the alcohol group (p < 0.05, respectively). The levels of SOD and GPX were opposite to that of MDA. Under a light microscope, the livers of the rats in the alcohol group showed unclear structure of hepatic lobules, stiffness of hepatic sinusoids, diffused lipid degeneration of hepatic cells, cellular swelling, and focal necrosis, while the structure remained clear in the Puerarin group. Under the electron microscope, lipid degeneration, cell organ decrease, enlargement of endoplasmic reticulum, reduced quantity of hepatins and swelling of mitochondria were observed in cells of the model group. However, the pathologistic changes were slight in the Puerarin group. In conclusion, Puerarin may have the function of inhibiting the oxidative stress induced by acute alcoholism.

This study investigates the effect of puerarin (PR) on peripheral nerve regeneration in vitro and in vivo. PR at concentrations of 1, 10, and 100 μM significantly promoted survival and outgrowth of cultured Schwann cells, as compared to the controls treated with culture medium only. in vivo study, peripheral nerve regeneration was evaluated across a 15-mm gap in the sciatic nerve of rats using a silicone rubber nerve chamber filled with PR solution. The control group chambers were filled with normal saline only. At the end of eight weeks, animals in the PR groups, especially at a concentration of 1 μM, had a significantly higher density of myelinated axons, greater evoked action potential area, and a larger nerve conductive velocity, as compared to the controls. All experimental results indicate that PR treatment promotes nerve growth and is a promising herbal medicine for recovery of regenerating peripheral nerves.

Several studies demonstrate that estradiol can prevent arterial calcification. However, little is known regarding the effect of puerarin, a phytoestrogen extracted from Radix Puerariae, on arterial calcification. The aim of the present study was to determine whether puerarin reduced osteoblastic differentiation of calcifying vascular smooth muscle cells (CVSMCs). The CVSMCs were isolated from mice aorta and treated with different concentrations of puerarin. The alkaline phosphatase (ALP) activity, osteocalcin secretion and Runx2 expression were determined. To examine whether estrogen receptors (ERs) PI3K and Akt play a role in this effect, ICI182789, phosphoinositide 3-kinase (PI3K) inhibitor, LY294002, or the Akt inhibitor, 1L-6-hydroxymethyl-chiro-inositol 2-(R)-2-O-methyl-3-O-octadecylcarbonate (HIMO) was used. Our results showed puerarin could inhibit ALP activity, osteocalcin secretion and Runx2 expression in CVSMCs. Puerarin could induce the activation of Akt. Furthermore, pretreatment of ICI182780, LY294002, HIMO could abolish the effect of puerarin on ALP activity in CVSMCs. Our experiment demonstrated that puerain could attenuate the osteoblastic differentiation of VSMCs through the ER/PI3K-Akt signal pathway.

Puerarin is an isoflavonoid isolated from the Chinese herb, Kudzu roots (also known as Gegen), which has been widely used for the treatment of hypertensive diseases and diabetic mellitus in traditional Chinese medicine. Dahl salt-sensitive (DS) rat is a genetic model of salt-sensitive hypertension with cardiovascular injury and vascular insulin resistance. Here, we investigated whether puerarin improved vascular insulin resistance and attenuated cardiac and aortic remodeling in salt-sensitive hypertension. DS rats were given a normal (NS) or high salt diet (HS) for five weeks. An additional group of DS rats was pretreated with puerarin and NS for 10 days, then switched to HS plus puerarin for five weeks. HS for five weeks increased systolic blood pressure (SBP), cardiac hypertrophy and fibrosis, and aortic hypertrophy with increased the expression of phosphor-ERK1/2 in the aorta and heart; puerarin attenuated cardiac and aortic hypertrophy, cardiac fibrosis and phosphor-ERK1/2 with a mild reduction in SBP. Hypertensive rats also manifested impairment of acetylcholine- and insulin-mediated vasorelaxation and insulin-mediated Akt and eNOS phosphorylation associated with the activation of NF$\kappa$B/TNF$\alpha$/JNK pathway. Puerarin improved acetylcholine- and insulin-mediated vasorelaxation and insulin-stimulated Akt/NO signaling with the inhibition of the NF$\kappa$B inflammatory pathway. Our results demonstrated that in salt-sensitive hypertension, puerarin improved vascular insulin action with cardiovascular beneficial effects. Our results found that the underlying mechanisms may involve its inhibition of NF$\kappa$B/JNK and ERK1/2 pathway. These results suggest that puerarin could be used as a new antihypertensive agent to expand our armamentarium for the prevention and treatment of end-organ damage in individuals with hypertension and metabolic diseases.

Puerarin is a traditional Chinese medicine with beneficial effects of reduced depression-like behaviors in mice with stress. Previous studies also show that puerarin can produce neuroprotective effect via activating the Akt or increased brain-derived neurotrophic factor (BDNF) expression. Interestingly, BDNF and Akt downstream target, mammalian target of rapamycin (mTOR) mediate the fast-acting antidepressant properties of ketamine. Until now, the involvement of the mTOR signaling pathway or BDNF on puerarin-induced antidepressant effect remains unknown. We aimed to investigate whether the antidepressant-like effect induced by puerarin would associate mTOR signaling pathway and BDNF release. The antidepressant-like effects of puerarin were evaluated using the forced swim test. The activation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionaic acid receptor (AMPAR)-mTOR signaling pathway and release of BDNF in the prefrontal cortex were determined. We also investigated the effect of puerarin on AMPAR trafficking through measuring the PKA phosphorylation of AMPAR subunit GluR1. Our present results show that puerarin exerted antidepressant-like responses that was mediated by AMPAR-induced mTOR signaling pathway and associated with increased BDNF release. Moreover, a significant increase in the GluR1 phosphorylation at its PKA site was noted following puerarin treatment. Our findings are the first to demonstrate that the antidepressant-like actions of puerarin require AMPAR–mTOR signaling pathway activation, are associated with an increased BDNF level and facilitate AMPAR membrane insertion. These findings provide preclinical evidence that puerarin may possess antidepressant property which is mediated by the glutamatergic system.

Diabetes mellitus (DM) has become one of the most challenging public health problems globally. The increasing prevalence and mortality rates call for more effective therapeutic agents, especially for DM complications. Traditional herbs have a long clinical application history for DM treatment. Puerarin is a natural isoflavone from Pueraria lobata (Wild.) Ohwi which has been consumed both as a functional food and herb in Eastern Asia countries. Documented data has shown that puerarin has cardio-protective, neuroprotective, anti-oxidative, anti-inflammatory and many other effects. In this review, we will summarize the beneficial effects and underlying mechanisms of puerarin on DM and complications. Puerarin may directly benefit DM by decreasing blood glucose levels, improving insulin resistance, protecting islets, inhibiting inflammation, decreasing oxidative stress and inhibiting Maillard reaction and advanced glycation end products (AGEs) formation. Furthermore, puerarin may also benefit DM indirectly by retarding and improving a series of DM complications, such as cardiovascular complications, diabetic nephropathy, diabetic retinopathy, diabetic neuropathy, etc. However, comprehensive studies of its effect and mechanisms are needed. In addition, its efficacy is relatively low, which is partially due to its pharmacokinetics profiles. Though puerarin shows low toxicity to experimental animals, its safety on human remains to be clarified. Collectively, we suggest that puerarin might be a potential adjuvant agent for the treatment of DM and DM complications in future.

Puerarin is an isoflavonoid isolated from the root of Pueraria lobata (Gegen in Chinese) that has been widely used to treat cardiovascular and cerebrovascular diseases in China. Here, we investigated the hypotensive effects and mechanisms of puerarin in spontaneously hypertensive rats. The qPCR array technique was used to determine the expression of hypertension-related genes. Then, the differentially expressed genes were analyzed using the STRING database. The systolic blood pressure and diastolic blood pressure of rats decreased after the administration of puerarin for nine weeks. Puerarin, but not losartan, also slowed the heart rate of rats. NO and cGMP levels were improved by puerarin. Eighteen differentially expressed hypertension-related genes were identified by comparing the model group with the control group and the high-dose puerarin group with the model group. NO and cGMP levels were increased by high-dose puerarin. High-dose puerarin increased the levels of the phosphorylated eNOS protein and decreased AT1 and Cav1 levels. Based on our results, eNOS was a key target in the mechanism by which puerarin reduced blood pressure, and puerarin represents a potential antihypertensive agent.

Type 1 diabetes (T1D) is an autoimmune and inflammatory disease with excessive loss of pancreatic islet $\beta$-cells. Accumulating evidence indicated that endoplasmic reticulum (ER) stress played a critical role in $\beta$-cells loss, leading to T1D. Therefore, promoting the survival of pancreatic $\beta$cells would be beneficial for patients with T1D. Puerarin is a natural isoflavone that has been demonstrated to be able to decrease blood glucose in patients with T1D. However, it remains unknown whether puerarin improves ER stress to prevent $\beta$-cells from apoptosis. Here, we sought to investigate the role of puerarin in ER stress-associated apoptosis and explore its underlying mechanism in the mouse insulinoma cell line (MIN6). Flow cytometry and cell counting kit-8 (CCK8) experiments showed that puerarin caused a significant increase in the viability of MIN6 cells injured by H₂O₂. Furthermore, the protein kinase R-like ER kinase (PERK) signal pathway, a critical branch of ER stress response, was found to be involved in this process. Puerarin inhibited the phosphorylation of PERK, subsequently suppressed the phosphorylation of eukaryotic initiation factor 2$\alpha$ (eIF2$\alpha )$, then decreased the activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP) expression, ultimately attenuating ER stress to prevent MIN6 cells from apoptosis. In addition, puerarin inhibited the activation of Janus kinase 2 (JAK2)/signal transducer and activators of transcription 3 (STAT3), which suppressed the PERK signal cascade with decreased ATF4 and CHOP levels. Taken together, our results firstly demonstrated that puerarin could prevent MIN6 cells from apoptosis at least in part by inhibiting the PERK-eIF2$\alpha$-ATF4-CHOP axis under ER stress conditions, which might be mediated by inactivation of the JAK2/STAT3 signal pathway. Therefore, investigating the mechanism underlying the effects of puerarin might highlight the potential roles of puerarin developing into an antidiabetic drug.

During menopause, the sharp decline in estrogen levels leads to an increased risk of cardiovascular disease in women. The inflammatory response and oxidative stress are reportedly involved in the development of cardiovascular disorders postmenopause. In this study, we evaluated the cardioprotective effects of puerarin, a phytoestrogen derived from the root of Pueraria lobate, and investigated its underlying molecular mechanisms. Puerarin alleviated cytotoxicity and the production of reactive oxygen species (ROS) in lipopolysaccharide (LPS)- and hydrogen peroxide-stimulated H9c2 cardiomyoblasts. Puerarin scavenges free radicals and reduces apoptosis, thereby suppressing NADPH oxidase-1 and Bax activation to attenuate the production of ROS and restore Bcl-2 expression. Additionally, puerarin inhibited the expression of inducible nitric oxide synthase, cyclooxygenase-2, and nitric oxide production and decreased the hypertrophic phenotype under LPS stimulation. Treatment with puerarin reduced the levels of malondialdehyde and restored glutathione levels when facing oxidative stress. Mechanistically, puerarin inhibited both the LPS-induced Toll-like receptor 4/NF-$\kappa$B and mitogen-activated protein kinase signaling pathways. Furthermore, it reversed both the LPS-mediated downregulation of Akt activation and heme oxygenase-1 (HO-1) expression. The cardioprotective effects of puerarin were abolished by inhibitors of Akt and HO-1 and the estrogen receptor antagonist fulvestrant (ICI). This indicated that the estrogen receptor mediated by these two molecules plays important roles in conferring the anti-inflammatory and anti-oxidative functions of puerarin. These results demonstrate the therapeutic potential of puerarin for treating heart disease in postmenopausal women through Akt and HO-1 activation.