ANGPTL3 is a novel HDL component that regulates HDL function.

BACKGROUND: Angiopoietin-like protein 3 (ANGPTL3) is secreted by hepatocytes and inhibits lipoprotein lipase and endothelial lipase activity. Previous studies reported the correlation between plasma ANGPTL3 levels and high-density lipoprotein (HDL). Recently ANGPTL3 was found to preferentially bind to HDL in healthy human circulation. Here, we examined whether ANGPTL3, as a component of HDL, modulates HDL function and affects HDL other components in human and mice with non-diabetes or type 2 diabetes mellitus. METHODS: HDL was isolated from the plasma of female non-diabetic subjects and type-2 diabetic mellitus (T2DM) patients. Immunoprecipitation, western blot, and ELISA assays were used to examine ANGPTL3 levels in HDL. Db/m and db/db mice, AAV virus mediated ANGPTL3 overexpression and knockdown models and ANGPTL3 knockout mice were used. The cholesterol efflux capacity induced by HDL was analyzed in macrophages preloaded with fluorescent cholesterol. The anti-inflammation capacity of HDL was assessed using flow cytometry to measure VCAM-1 and ICAM-1 expression levels in TNF-α-stimulated endothelial cells pretreated with HDL. RESULTS: ANGPTL3 was found to bind to HDL and be a component of HDL in both non-diabetic subjects and T2DM patients. Flag-ANGPTL3 was found in the HDL of transgenic mice overexpressing Flag-ANGPTL3. ANGPLT3 of HDL was positively associated with cholesterol efflux in female non-diabetic controls (r = 0.4102, p = 0.0117) but not in female T2DM patients (r = - 0.1725, p = 0.3224). Lower ANGPTL3 levels of HDL were found in diabetic (db/db) mice compared to control (db/m) mice and were associated with reduced cholesterol efflux and inhibition of VCAM-1 and ICAM-1 expression in endothelial cells (p < 0.05 for all). Following AAV-mediated ANGPTL3 cDNA transfer in db/db mice, ANGPTL3 levels were found to be increased in HDL, and corresponded to increased cholesterol efflux and decreased ICAM-1 expression. In contrast, knockdown of ANGPTL3 levels in HDL by AAV-mediated shRNA transfer led to a reduction in HDL function (p < 0.05 for both). Plasma total cholesterol, total triglycerides, HDL-c, protein components of HDL and the cholesterol efflux function of HDL were lower in ANGPTL3-/- mice than ANGPTL3+/+ mice, suggesting that ANGPTL3 in HDL may regulate HDL function by disrupting the balance of protein components in HDL. CONCLUSION: ANGPTL3 was identified as a component of HDL in humans and mice. ANGPTL3 of HDL regulated cholesterol efflux and the anti-inflammatory functions of HDL in T2DM mice. Both the protein components of HDL and cholesterol efflux capacity of HDL were decreased in ANGPTL3-/- mice. Our findings suggest that ANGPTL3 in HDL may regulate HDL function by disrupting the balance of protein components in HDL. Our study contributes to a more comprehensive understanding of the role of ANGPTL3 in lipid metabolism.

A Prediction Model for Sight-Threatening Diabetic Retinopathy Based on Plasma Adipokines among Patients with Mild Diabetic Retinopathy.

Background: Accumulating evidence has suggested a link between adipokines and diabetic retinopathy (DR). This study is aimed at investigating the risk factors for sight-threatening DR (STDR) and establishing a prognostic model for predicting STDR among a high-risk population of patients with type 2 diabetes mellitus (T2DM). Methods: Plasma concentrations of adipokines were determined by enzyme-linked immunosorbent assay. In the case-control set, principal component analysis (PCA) was performed to select optimal predictive cytokines for STDR, involving severe nonproliferative DR (NPDR) and proliferative DR. Support vector machine (SVM) was used to examine the possible combination of baseline plasma adipokines to discriminate the patients with mild NPDR who will later develop STDR. An individual prospective cohort with a follow-up period of 3 years was used for the external validation. Results: In both training and testing sets, involving 306 patients with T2DM, median levels of plasma adiponectin (APN), leptin, and fatty acid-binding protein 4 (FABP4) were significantly higher in the STDR group than those in mild NPDR. Except for adipsin, the other three adipokines, FABP4, APN, and leptin, were selected by PCA and integrated into SVM. The accuracy of the multivariate SVM classification model was acceptable in both the training set (AUC = 0.81, sensitivity = 71%, and specificity = 91%) and the testing set (AUC = 0.77, sensitivity = 61%, and specificity = 92%). 110 T2DM patients with mild NPDR, the high-risk population of STDR, were enrolled for external validation. Based on the SVM, the risk of each patient was calculated. More STDR occurred in the high-risk group than in the low-risk group, which were grouped by the median value of APN, FABP4, and leptin, respectively. The model was validated in an individual cohort using SVM with the AUC, sensitivity, and specificity reaching 0.77, 64%, and 91%, respectively. Conclusions: Adiponectin, leptin, and FABP4 were demonstrated to be associated with the severity of DR and maybe good predictors for STDR, suggesting that adipokines may play an important role in the pathophysiology of DR development.

Central role of cardiac fibroblasts in myocardial fibrosis of diabetic cardiomyopathy.

Diabetic cardiomyopathy (DCM), a main cardiovascular complication of diabetes, can eventually develop into heart failure and affect the prognosis of patients. Myocardial fibrosis is the main factor causing ventricular wall stiffness and heart failure in DCM. Early control of myocardial fibrosis in DCM is of great significance to prevent or postpone the progression of DCM to heart failure. A growing body of evidence suggests that cardiomyocytes, immunocytes, and endothelial cells involve fibrogenic actions, however, cardiac fibroblasts, the main participants in collagen production, are situated in the most central position in cardiac fibrosis. In this review, we systematically elaborate the source and physiological role of myocardial fibroblasts in the context of DCM, and we also discuss the potential action and mechanism of cardiac fibroblasts in promoting fibrosis, so as to provide guidance for formulating strategies for prevention and treatment of cardiac fibrosis in DCM.

The Microbiota-Dependent Treatment of Wuzhuyu Decoction for Chronic Migraine Model Rat Associated with Anxiety-Depression Like Behavior.

We previously found that Wuzhuyu Decoction (WZYD) could affect central and peripheral 5-HT to relieve hyperalgesia in chronic migraine (CM) model rats, possibly related to gut microbiota. However, the exact role of gut microbiota has not been elucidated. Accumulating evidence points to the possibility of treating central nervous system disease via the gut-brain axis. In our study, the inflammatory soup-induced CM model rats presented depression- and anxiety-like behaviors which both related to insufficient 5-HT. It was found that antibiotic administration caused community dysbiosis, and proteobacteria became the main dominant bacteria. The bacteria related to short-chain fatty acids and 5-HT generation were reduced, resulting in reduced levels of 5-HT, tryptophan hydroxylase, and secondary bile acids. Functional prediction-revealed sphingolipid signaling pathway in CM rats was significantly decreased and elevated after WZYD treatment. The effect of WZYD could be weakened by antibiotics. The CM rats exhibited anxiety- and depression-like behavior with 5-HT and number of neurons decreased in the CA1 and CA2 regions of hippocampal. The treatment of WZYD could recover to varying degrees. Antibiotics combined with WZYD attenuate the effect of WZYD on increasing the 5-HT content and related protein expression in the brain stem, plasma and colon, reducing CGRP, c-Fos, and inflammatory factors. And antibiotics also led to colon length increasing and stool retention, so that the antimigraine effect was weakened compared with WZYD. This experiment revealed that gut microbiota mediated WZYD treatment of CM rats with anxiety-depression like behavior.

Mesenchymal Stem Cells Promote Wound Healing and Effects on Expression of Matrix Metalloproteinases-8 and 9 in the Wound Tissue of Diabetic Rats.

Diabetic foot ulcer (DFU) is a multifactorial complication of diabetes, mainly manifested as infection, ulcer, or destruction of deep tissue, and there is currently no effective treatment. Several preclinical and clinical studies have proved that the transplantation of mesenchymal stem cells (MSCs) improved wound healing. In this study, we evaluated the therapeutic efficacy of human umbilical cord (hUC-MSCs) in DFU rat model. One dose of hUC-MSCs (1 × 106 cells) was subcutaneously injected around wounds in male Sprague-Dawley rats. Wound healing was evaluated macroscopically (wound closure) every 3 days. In addition, we measured growth factors and specific proteins [matrix metalloproteinases (MMPs)-9 and MMP-8] on Day 14 post hUC-MSC transplantation. Results showed significant differences in the wound healing kinetics of lesions that received hUC-MSCs compared to lesions that received vehicle (phosphate buffered saline; P < 0.05). Enzyme-linked immunosorbent assay analyses indicated that MMP-9 protein contents were significantly upregulated in DFU animals, while MMP-8 was downregulated compared to the diabetic rats (P < 0.05). After MSC treatment, the level of MMP-9 and MMP-8 decreased and increased compared to the vehicle group, respectively. These findings suggest that hUC-MSC transplantation can ameliorate the healing process of DFU rats and a potential mechanism through which MSCs enhance DFU wound healing by decreasing MMP-9 expression and increasing MMP-8 expression. This study represents a promising opportunity to gain insight into how MSCs mediate wound healing.

Role of Dipeptidyl Peptidase 4 Inhibitors in Antidiabetic Treatment.

In recent years, important changes have occurred in the field of diabetes treatment. The focus of the treatment of diabetic patients has shifted from the control of blood glucose itself to the overall management of risk factors, while adjusting blood glucose goals according to individualization. In addition, regulators need to approve new antidiabetic drugs which have been tested for cardiovascular safety. Thus, the newest class of drugs has been shown to reduce major adverse cardiovascular events, including sodium-glucose transporter 2 (SGLT2) and some glucagon like peptide 1 receptor (GLP1) analog. As such, they have a prominent place in the hyperglycemia treatment algorithms. In recent years, the role of DPP4 inhibitors (DPP4i) has been modified. DPP4i have a favorable safety profile and anti-inflammatory profile, do not cause hypoglycemia or weight gain, and do not require dose escalation. In addition, it can also be applied to some types of chronic kidney disease patients and elderly patients with diabetes. Overall, DPP4i, as a class of safe oral hypoglycemic agents, have a role in the management of diabetic patients, and there is extensive experience in their use.

SGLT2 inhibition restrains thyroid cancer growth via G1/S phase transition arrest and apoptosis mediated by DNA damage response signaling pathways.

BACKGROUND: Although the prognosis for most patients with papillary thyroid cancer (PTC) is good, the present treatment is ineffective for 5-10% patients. Several studies found sodium-glucose cotransporter 2 (SGLT2) inhibitors may inhibit the growth of tumors. However, whether SGLT2 inhibitors have therapeutic effect on thyroid cancer remains unclear. MATERIALS AND METHODS: The levels of SGLT2 in PTC and normal thyroid tissue were assessed by immunohistochemistry and clinical dataset analysis. Cell growth was detected by the CCK-8 and colony formation. Glucose uptake into thyroid cancer cell was evaluated by 2-DG uptake assay. Glycolysis were analyzed by Seahorse XF Extracellular Flux Analysis. RNA-seq were used to screen differentially expressed genes of cells treated with/without canagliflozin (a SGLT2 inhibitor). Furthermore, flow cytometry, western blot, and gene set enrichment analysis were employed to elucidate cell cycle, apoptosis and the underlying mechanism of the anticancer effect of canagliflozin. The effect of canagliflozin on thyroid cancer growth was further confirmed in vivo through xenograft formation assay. RESULTS: SGLT2 inhibition attenuated the growth of thyroid cancer cells in vitro and in vivo. Canagliflozin inhibited glucose uptake, glycolysis and AKT/mTOR signaling activation, and increased AMPK activation in thyroid cancer cell. Furthermore, canagliflozin inhibited G1/S phase transition and cyclin D1, cyclin D3, cyclin E1, cyclin E2, and E2F1 expression levels in thyroid cancer cell. In addition, canagliflozin increased apoptosis of thyroid cancer cell. Further investigation revealed that canagliflozin could increase γ-H2AX expression levels and DNA damage response signaling ATM/CHK2 activation. In thyroid cancer patients, SGLT2 was increased in thyroid cancer and positively related to cyclin D3. CONCLUSIONS: SGLT2 inhibition may limit glucose uptake resulting in energetic crisis, following oxidative stress mediated DNA damage and cell cycle arrest, which resulted to the increased cell apoptosis and decreased proliferation of thyroid cancer cells, suggesting a potential use for SGLT2 inhibitors as thyroid cancer therapeutics.

Wuzhuyu Decoction relieves hyperalgesia by regulating central and peripheral 5-HT in chronic migraine model rats.

BACKGROUND: Chronic migraine (CM) is a highly disabling and burdensome disease. Wuzhuyu decoction (WZYD), a clinical used formula to treat and prevent episodic migraine and CM, has been reported to relieve the hyperalgesia of CM and increase brainstem and blood serotonin (5-hydroxytryptamine, 5-HT) in migraine model rats in previous studies; yet the mechanism is unclear. PURPOSE: This study aimed to observe the hyperalgesia relief effect of WZYD and investigate the mechanistic association with the regulation on central and peripheral 5-HT. METHODS: WZYD with different doses (3.372, 1.686 and 0.843 g/kg∙d) and the positive drug - sumatriptan (5.83 mg/kg∙3 d) were intragastrically administered in inflammatory soup (IS)-induced CM model rats, respectively. Hyperalgesia was assessed by facial mechanical withdrawal threshold and tail-flick latency. 5-HT was determined by ELISA. Western blot analysis, immunohistochemistry and immunofluorescence determination, and 16S rRNA gene sequencing were performed. RESULTS: WZYD significantly relieved the hyperalgesia by elevating the facial mechanical withdrawal threshold and tail-flick latency. In WZYD groups, increased 5-HT and decreased calcitonin gene-related peptide in both the brainstem and plasma, downregulated TNF-α, IL-1ß, and c-fos expression in the brainstem were observed in dose-dependent manner. Interestingly, 5-HT in colon tissues were also observed, which is associated with upregulating tryptophan hydroxylase, serotonin transporter and Piezo1 expression and increasing 5-HT and chromogranin A in enterochromaffin cells. Disorder of the microbiota, function and metabolism was correlated with 5-HT synthesis. WZYD could regulate the abundance of Anaerostipes and Acidifaciens. CONCLUSION: WZYD has the pharmacological effect on relieving hyperalgesia in CM model rats, possibly by affecting central and peripheral 5-HT.

Therapeutic Effect of Matrine on Collagen-Induced Arthritis Rats and Its Regulatory Effect on RANKL and OPG Expression.

OBJECTIVE: To investigate the effect of matrine on rats with collagen-induced arthritis (CIA) and its regulatory effect on receptor activator of nuclear factor-κB ligand (RANKL) and osteoprotegerin (OPG) expression. METHODS: Wistar rats (n = 6) and CIA rats (n = 30) were randomly divided into six groups: healthy, CIA control, low/medium/high matrine (25, 50, or 100 mg/kg, once per day for six weeks), and methotrexate (MTX) (2 mg/kg, once per week for six weeks). The degree of joint damage was evaluated by X-ray and HE staining. Bone marrow suppression was assessed by routine blood analysis. In addition, the levels of serum RANKL and OPG in the rats were measured by ELISA. RESULTS: The level of joint swelling and degree of joint damage assessed by ankle swelling measurements, AI score, X-ray, and HE staining were alleviated in the CIA rats treated with MTX or different doses of matrine. Furthermore, no obvious inhibitory effect was observed on the bone marrow of the CIA rats, regardless of the dose of matrine or treatment with 2 mg/kg MTX (P > 0.05). The levels of OPG in serum and the ratio of OPG/RANKL were higher, and RANKL expression was lower in the low/medium/high matrine group compared with that of the CIA control group. The serum levels of OPG and OPG/RANKL ratio increased with the matrine dose, while the opposite was observed for RANKL expression. CONCLUSION: Matrine treatment was associated with a lower degree of bone destruction, increased OPG expression and OPG/RANKL ratio, and decreased RANKL expression in CIA rats. Thus, matrine may represent a novel drug candidate for the treatment of RA.

[UPLC-MS/MS determination of active components from Wuzhuyu Decoction in plasma and brain tissues of nitroglycerin-induced migraine rats].

A sensitive and specific ultra-performance liquid chromatography-mass spectrometry(UPLC-MS/MS) method was deve-loped for analysis of rutaecarpine(Ru), evodiamine(Ev), rutaevine(Rv), limonin(Li), ginsendside Rb_1(Rb_1), ginsendside Re(Re) in rat plasma and brain tissues of nitroglycerin-induced migraine rats. Male healthy Sprague-Dawley(SD) rats were orally given multiple dose of optimized(OS) and un-optimized Wuzhuyu Decoction(UNOS), and their blood samples and brainstem were collected at different time points after injection of nitroglycerin(10 mg·kg~(-1)) into the frontal region. The drug concentrations of the 6 analytes in plasma and brainstem were determined by UPLC-MS/MS method. Subsequently, the main pharmacokinetics parameters of plasma were calculated by using Phoenix WinNolin 5.2.1 software. The methodological test showed that all of analytes in both plasma and brainstem homogenate exhibited a good linearity within the concentration range(r>0.994 7). The intra-day and inter-day accuracy, precision, matrix effect, stability of the investigated components meet the requirements for biopharmaceutical analysis. The developed method was successfully applied in pharmacokinetic studies on abovementioned ingredients in rat plasma and brain stem. The plasma pharmacokinetic parameters of active ingredients in two different Wuzhuyu Decoction group were compared, it was found that Rb_1 had higher t_(1/2), T_(max), C_(max), AUC_(0-24 h) and AUC_(0-∞ )in OS group. Meanwhile, Ev had higher t_(1/2) and T_(max) but lower C_(max), AUC_(0-24 h) and AUC_(0-∞), Ru has higher t_(1/2 )but lower C_(max), AUC_(0-24 h) and AUC_(0-∞ )in OS group. The brain tissue distribution of each component were compared between the two groups, the component with higher content in OS, such as Ru at 30 min and 2 h after administration, Ev at 30 min, Rb_1 at 30 min and Rb_1 at 2 h after administration have lower brain tissue distribution than those in UNOS group, while the component with higher content in UNOS, such as Rv at 30 min, 2 h and 12 h after administration had higher brain tissue distribution than those in OS group.

Dynamic Alterations in the Gut Microbiota of Collagen-Induced Arthritis Rats Following the Prolonged Administration of Total Glucosides of Paeony.

Rheumatoid arthritis (RA) is a common autoimmune disease linked to chronic inflammation. Dysbiosis of the gut microbiota has been proposed to contribute to the risk of RA, and a large number of researchers have investigated the gut-joint axis hypothesis using the collagen-induced arthritis (CIA) rats. However, previous studies mainly involved short-term experiments; very few used the CIA model to investigate changes in gut microbiota over time. Moreover, previous research failed to use the CIA model to carry out detailed investigations of the effects of drug treatments upon inflammation in the joints, hyperplasia of the synovium, imbalance in the ratios of Th1/Th2 and Th17/Treg cells, intestinal cytokines and the gut microbiota following long-term intervention. In the present study, we carried out a 16-week experiment to investigate changes in the gut microbiota of CIA rats, and evaluated the modulatory effect of total glucosides of paeony (TGP), an immunomodulatory agent widely used in the treatment of RA, after 12 weeks of administration. We found that taxonomic differences developed in the microbial structure between the CIA group and the Control group. Furthermore, the administration of TGP was able to correct 78% of these taxonomic differences, while also increase the relative abundance of certain forms of beneficial symbiotic bacteria. By the end of the experiment, TGP had reduced body weight, thymus index and inflammatory cell infiltration in the ankle joint of CIA rats. Furthermore, the administration of TGP had down-regulated the synovial content of VEGF and the levels of Th1 cells and Th17 cells in CIA rats, and up-regulated the levels of Th2 cells and Treg cells. The administration of TGP also inhibited the levels of intestinal cytokines, secretory immunoglobulin A (SIgA) and Interferon-γ (IFN-γ). In conclusion, the influence of TGP on dynamic changes in gut microbiota, along with the observed improvement of indicators related to CIA symptoms during 12 weeks of administration, supported the hypothesis that the microbiome may play a role in TGP-mediated therapeutic effects in CIA rats. The present study also indicated that the mechanism underlying these effects may be related to the regulation of intestinal mucosal immunity remains unknown and deserves further research attention.

Recognition and Optimization of Ingredients Treating Nitroglycerin-Induced Migraine Rats from Wuzhuyu Decoction.

Wuzhuyu decoction (WZYD) has been clinically used to treat migraine effectively since Eastern Han Dynasty of ancient China. However, its antimigrainic ingredients remain unclear. In present study, the antimigrainic ingredients of WZYD were explored and optimized in nitroglycerin-induced migraine rats through correlation analysis of decoction spectra-pharmacological effects and absorption spectra-pharmacological using entropy-weighted partial least squares regression method. The decoction spectra and absorption spectra were obtained through the determination of nine main ingredients in ten kinds of WZYDs and WZYDs' single-pass intestinal perfusion samples using high performance liquid chromatography-diode array detector. The pharmacodynamics indexes related to migraine model rats were detected using high performance liquid chromatography method and kits after oral administration of WZYDs. Then, the key ingredients influencing indexes were achieved through the correlation analysis. And the optimization of key ingredients was acquired through uniform design experiment. The pharmacodynamic verification test was used to clarify the advantages of the optimized sample. The results showed that the final optimized sample, in which the concentrations of rutaecarpine, evodiamine, ginsendside Rb1, 6-gingerol, ginsendside Rg1, rutaevine, and limonin were 0.081, 0.565, 1.455, 0.159, 0.871, 0.178, and 0.009 mg·mL-1, respectively, provided the best comprehensive effect than another optimized sample and the best uniform design sample. Therefore, a new reliable method for rapidly recognizing and optimizing the effective constituents of WZYD treating migraine was established.

Comparative Pharmacokinetic Study of Taxifolin after Oral Administration of Fructus Polygoni Orientalis Extract in Normal and Fibrotic Rats by UPLC-MS/MS.

Fructus polygoni orientalis (FPO) is widely used in clinical practice in China, especially in treatment of liver diseases including viral hepatitis, liver fibrosis, and liver cirrhosis. However, its pharmacokinetic (PK) alterations in liver fibrotic rats have rarely been reported. To study whether taxifolin, one of the main flavonoids in FPO can be absorbed into blood after oral administration of FPO extract and to compare the differences in pharmacokinetic parameters of taxifolin to normal and liver fibrotic rats induced by porcine serum (PS), a UPLC-MS/MS method was developed and validated for determination of taxifolin in rat plasma using puerarin as the internal standard (IS). All validation parameters met the acceptance criteria according to regulatory guidelines. The results indicated that after treatment of rats with PS alone for 12 weeks, the liver fibrotic model group was built successfully. The taxifolin can be absorbed into the blood after oral administration of the FPO extract. The C max of taxifolin was 1940 ± 502.2 ng/mL and 2648 ± 208.5 ng/mL (p < 0.05), the AUC0∼t of taxifolin was 4949.7 ± 764.89 h·ng/mL and 6679.9 ± 734.26 h·ng/mL (p < 0.05), the AUC0∼∞ of taxifolin was 5049.4 ± 760.7 and 7095.2 ± 962.3 h·ng/mL (p < 0.05), and the mean residence time (MRT) of taxifolin was 2.46 ± 0.412 h and 3.17 ± 0.039 h (p < 0.05) in the normal and fibrotic model groups, respectively. These results confirmed that the pharmacokinetic parameters of taxifolin are altered in liver fibrosis, manifested as C max, AUC0∼t , AUC0∼∞, and the mean residence time (MRT). It suggested that it is essential to consider the characteristics of pharmacokinetics after oral administration of FPO in liver disease patients.

[Difference in intestinal absorption of Wuzhuyu decoction between normal and migraine model rats].

To compare the intestinal absorption of Wuzhuyu decoction(WZYD) between normal rats and migraine model rats, and investigate the optimized WZYD from aspect of absorption. The rat single pass intestinal perfusion test(SPIP) was adopted for optimized sample and un-optimized sample in normal and migraine model rats induced by nitroglycerin and reserpine. The contents of 8 ingredients were determined by high performance liquid chromatography(HPLC), and 4 absorption parameters for each ingredient were calculated and compared: unit area absorption(Mper area), absorption rate constant(Ka), apparent coefficient(Papp) and relative absorption rate(RA). The results showed that there was a great difference between normal rats and model rats in the intestinal absorption of the same WZYD. As compared with normal rats, the absorption parameters of most ingredients in optimized sample were increased in migraine model rats induced by nitroglycerin; Similar phenomena were also found in migraine model rats induced by reserpine. However, the absorption parameters of most ingredients were decreased in un-optimized sample. Therefore, pathological model rats shall be used for effective ingredient recognition based on the correlation between intestinal absorption spectra and pharmacological effects. As compared with the un-optimized samples, the absorption of effective ingredients was faster, easier and more adequate in the optimized samples, revealing their mechanism on better efficacy from the aspect of absorption.

[Diagnostic value of dynamic-extended focused assessment with sonography for trauma in patients with multiple trauma].

OBJECTIVE: To investigate the diagnostic value of dynamic-extended focused assessment with sonography for trauma (D-EFAST) in patients with multiple trauma in intensive care unit (ICU). METHODS: A prospective clinical study was conducted. Eighty patients with multiple trauma admitted to ICU of Anhui Provincial Hospital from September 1st, 2014 to December 31st, 2016 were enrolled. Extended focused assessment with sonography for trauma (E-FAST) check was conducted at first, for those who had positive findings diagnosis was confirmed by immediately CT examination or surgical exploration. If it was negative, the patients received E-FAST every morning for 7 days (defined as D-EFAST), for those with positive findings, immediately CT or surgery was performed to clarify the diagnosis. The final clinical diagnosis was used as the "gold standard" to calculate the diagnostic accordance rate of EFAST and D-EFAST examination technique for pneumothorax, pleural effusion, spleen injury, kidney damage, liver damage, gastrointestinal injury, pericardial effusion, bladder rupture, and pancreatic injury, as well as their sensitivity, specificity, positive predictive value, negative predictive value, accuracy rate, and missed diagnosis rate, and the difference between EFAST and D-EFAST was compared. RESULTS: There were 4 patients excluded because of death and abandoning treatment, and finally 76 patients were included in the study. The total sensitivity of E-FAST examination technique for pneumothorax, pleural effusion, spleen injury, liver damage, gastrointestinal injury, pericardial effusion, and bladder rupture was 75.9% (66/87), and the specificity was 98.3% (587/597), the positive predictive value was 86.8% (66/76), and the negative predictive value was 96.5% (587/608), the accuracy rate was 95.5% (653/684), and the rate of missed diagnosis was 24.1% (21/87). The most of the delayed injury in patients with multiple trauma occurred at 2-7 days after injury with incidence of 4.8% (33/684). The diagnostic sensitivity of D-EFAST for delayed injury was 98.3% (118/120), the specificity was 99.8% (563/564), the positive predictive value was 99.2% (118/119), the negative predictive value was 99.6% (563/565), the diagnostic accuracy rate was 99.6% (681/684), and rate of missed diagnosis was 1.7% (2/120). When the final clinical diagnosis was set as the "gold standard", D-EFAST technology for the detection rate was 98.3% (118/120) for patients with multiple trauma on organ injury while the detection rate of E-FAST was 75.9% (66/87), with statistical significant difference (P < 0.01), indicating that D-EFAST was better than E-FAST in check of multiple trauma patients with organ injury. CONCLUSIONS: Although the E-FAST technology can quickly diagnose the multiple trauma patients and win the rescue time for critical patients, multiple trauma patients injured after 2-7 days prone to delayed damage and are difficult to detect, and D-EFAST can be used to find delayed damage earlier, and reduce the misdiagnosis rate of multiple trauma patients.

[Study on correlation between color and effective components contents of Glycyrrhiza uralensis].

To explore the correlation between color of Glycyrrhiza uralensis and its quality evaluation,the colors of root bark and transverse section were determined by Precision Color Reader and Visual Analyzer,and the contents of six flavonoids and two saponins in G.uralensis were determined by high performance liquid chromatography（HPLC）.The partial least squares regression（PLSR）method was employed to correlate the colors with component contents in G.uralensis. The results showed that there were no significant differences in the colors of root bark but significant or very significant differences（P<0.05,P<0.01）in the colors of transverse section between the wild and cultivated G. uralensis. Compared with those in the cultivated G. uralensis, the contents of liquiritin, isoliquiritin isoliquiritigenin and the contents of ammonium glycyrrhizinate, glycyrrhetinic acid were obviously significant or remarkably significant in the wild G. uralensis.The correlation results showed that there was a significant or very significant correlation between the colors and the effective component contents. This study provides a scientific basis to evaluate the quality of G.uralensis by color and a new reference for the traditional evaluation methods for Chinese drugs.

The effects of 18ß-glycyrrhetinic acid and glycyrrhizin on intestinal absorption of paeoniflorin using the everted rat gut sac model.

Paeoniflorin (PF), the main active component of Shaoyao-Gancao-tang, possesses significantly antinociceptive effects and many other pharmacological activities. However, its poor intestinal absorption results in low bioavailability. Therefore, enhancing PF absorption plays a vital role in exerting its therapeutic effect. Shaoyao combined with Gancao exhibited a synergistic effect. The enhancement of PF absorption through the interaction of its constituents in intestinal absorption would be greatly implicated. The present study aimed at investigating the effects of glycyrrhizin, the main constituent of Gancao, and its main metabolite, 18ß-glycyrrhetinic acid (18ß-GA), on the intestinal absorptive behavior of PF, and the role of P-glycoprotein (P-gp) in PF absorption using the in vitro everted rat gut sac model. The results demonstrated that 1 mM of 18ß-GA significantly increased PF absorption in both the jejunum and the ileum, while 100 µM of 18ß-GA only promoted the ileum absorption and had no obvious effect on the jejunum absorption. The effect of glycyrrhizin on intestinal PF absorption was related to concentrations. One mM of glycyrrhizin significantly increased PF absorption in the jejunum after 45 min and in the ileum after 90 min. But 100 µM of glycyrrhizin had an inhibitory effect in the jejunum and no effect in the ileum before 60 min. Moreover, verapamil, the well-known P-gp inhibitor, could significantly enhance the PF absorption. In conclusion, the influence of 18ß-GA and glycyrrhizin on the PF absorption was related to concentrations and intestinal segments. This might be involved in the intervention of efflux transport of PF mediated by intestinal P-gp.

Liver-targeting self-assembled hyaluronic acid-glycyrrhetinic acid micelles enhance hepato-protective effect of silybin after oral administration.

In order to enhance oral bioavailability and liver targeting delivery of silybin, two amphiphilic hyaluronic acid derivatives, hyaluronic acid-deoxycholic acid (HA-adh-DOCA) and hyaluronic acid-glycyrrhetinic acid (HA-adh-GA) conjugates, were designed and synthesized. Silybin was successfully loaded in HA-adh-DOCA and HA-adh-GA micelles with high drug-loading capacities (20.3% ± 0.5% and 20.6% ± 0.6%, respectively). The silybin-loaded micelles were spherical in shape with the average size around 130 nm. In vitro release study showed that two silybin-loaded micelles displayed similar steady continued-release pattern in simulated gastrointestinal fluids and PBS. Single-pass intestinal perfusion studies indicated that silybin-loaded micelles were absorbed in the whole intestine and transported via a passive diffusion mechanism. Compared with suspension formulation, silybin-loaded HA-adh-DOCA and HA-adh-GA micelles achieved significantly higher AUC and Cmax level. Moreover, liver targeting drug delivery of micelles was confirmed by in vivo imaging analysis. In comparison between the two micellar formulations, HA-adh-GA micelles possessed higher targeting capacity than HA-adh-DOCA micelles, owing to the active hepatic targeting properties of glycyrrhetinic acid. In the treatment of acute liver injury induced by CCl4, silybin-loaded HA-adh-GA micelles displayed better effects over suspension control and silybin-loaded HA-adh-DOCA micelles. Overall, pharmaceutical and pharmacological indicators suggested that the HA-adh-GA conjugates can be successfully utilized for liver targeting of orally administered therapeutics.

Optimization of combinations of ginsenoside-Rg1, ginsenoside-Rb1, evodiamine and rutaecarpine for effective therapy of mouse migraine.

Wuzhuyu decoction (WZYD) is a classic traditional Chinese medicine (TCM) formula. It has been extensively used for treating migraine for thousands of years in TCM. Four potential active ingredients from WZYD, ginsenoside-Rg1 (Rg1), ginsenoside-Rb1 (Rb1), evodiamine (Ev) and rutaecarpine (Ru), were found to have positive correlations with pharmacodynamic indicators involving mouse migraine in our previous study. To find a better therapeutic effect on migraine, this research was carried out to optimize the combinations of Rg1, Rb1, Ev and Ru using the uniform design method. The results showed that Rb1 and Ev played key roles in improving the therapeutic effect on mouse migraine by strongly ameliorating pharmacodynamic indicators associated with migraine. They significantly increased the contents of 5-hydroxytryptamine, noradrenaline and dopamine in brain tissues, and reduced the content of nitric oxide in brain tissues and the activities of nitric oxide synthase in both brain tissues and blood serum. The optimal concentrations of Rb1 and Ev were 1057.4 mg/L and 312.5 mg/L, respectively. Rg1 and Ru contributed less to the overall desirability, suggesting that they had reverse effects on some pharmacodynamic indicators of this type of migraine. The verification test demonstrated by the immunohistochemical method that the optimal combination inhibited the expression of c-fos and c-jun in periaqueductal gray of mice, and strongly ameliorated pharmacodynamic indicators. These results suggested that the therapeutic effect of the optimal combination of the four ingredients was strong, and the optimal results were proven to be reliable and accurate.

Identification of active ingredients in Wuzhuyu decoction improving migraine in mice by spectral efficiency association.

Wuzhuyu decoction is a traditional Chinese medicine used for the effective treatment of migraines, termed 'Jueyin headache', in China. However, there have been few investigations to clarify the composition of Wuzhuyu decoction for the treatment of migraines. In the present study, 10 types of Wuzhuyu decoction were analyzed by chromatograms. 5-hydroxytryptamine (5-HT)-depletion mouse models of migraine were prepared by subcutaneous injection of reserpine and placement of autologous blood clots in the cerebral cortex. The levels of 5-HT, noradrenaline (NE), dopamine (DA), nitric oxide (NO) and nitric oxide synthase (NOS) in the brain tissues and sera of the mice were determined. The ingredients and pharmacodynamic indices of the Wuzhuyu decoctions were analyzed using spectral efficiency association by partial least squares regression. The levels of 5-HT, NE and DA in the mouse brain tissues were reduced to 337.785 ± 84.504, 171.173 ± 65.172 and 242.075 ± 158.621 mg/g brain tissue, respectively. The level of NO in the brain tissues increased to 0.425 ± 0.184 µmol/g protein and the activities of NOS in the brain tissues and sera increased to 0.719 ± 0.477 U/mg and 50.688 ± 8.132 U/ml, respectively. Regarding the ingredients of the Wuzhuyu decoction, those with significant regression coefficients were ginsenoside-Rg1, Re, Rb1, rutaevine (Rv), limonin (Li), evodiamine (Ev), rutaecarpine (Ru) and substance X (awaiting identification). Rg1, Re, Rb1, Rv, Li, Ev, Ru and X in the Wuzhuyu decoction were observed to yield the pharmacological effects, whereas Rb1, Rv and Ev were important in index improvement.