To screen the active ingredients of Gardenia jasminoides and potential targets,and investigate the mechanisms against cholestasis based on network pharmacology technology. Twenty-one active components of G. jasminoides were retrieved and the target sites were screened by using Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform( TCMSP). Cytoscape3. 2. 1 was used to construct the component-target network. Two hundred and eight targets related to cholestasis were searched and screened through Dis Ge NET,KEGG and OMIM databases. The key targets of G. jasminoides components and cholestasis were integrated and screened,and the component-target-disease network was constructed with Cytoscape 3. 2. 1 software to screen out the core network whose freedom degree was greater than the average value. The Clue GO plug-in of Cytoscape 3. 2. 1 software was used to analyze the biological processes and pathway enrichment of G. jasminoides in regulation of cholestasis. GO biological process analysis revealed 17 biological processes,involving 3 signaling biological processes related to cholestasis,i.e. acute inflammatory response,positive regulation of reactive oxygen species metabolic process,and nitric oxide biosynthetic process. KEGG-KEEG-305 terms and REACTOME pathways analysis revealed 17 regulatory pathways,involving 4 signaling pathways related to cholestasis,i.e. metabolism of xenobiotics by cytochrome P450,nuclear receptor transcription pathway,GPVI-mediated activation cascade and platelet activation. It was found that aqueous extract of G. jasminoides could improve serum biochemical abnormalities in ANIT-induced cholestasis rats. Aqueous extract of G. jasminoides could decrease the protein and mRNA expression levels of ESR1 in liver tissues,and increase the protein and mRNA expression levels of PPARG,NOS2,F2 R,NOS3,and NR3 C1. To sum up,the possible mechanisms of G. jasminoides against cholestasis may be related with the above three processes and four pathways.
Animals ; Cholestasis ; drug therapy ; Drugs, Chinese Herbal ; pharmacology ; Gardenia ; chemistry ; Medicine, Chinese Traditional ; Plant Extracts ; pharmacology ; Rats ; Signal Transduction

To investigate the effects of geniposidic acid( GPA) on hepato-enteric circulation in cholestasis rats,and to explore the mechanism based on the sirtuin 1( Sirt1)-farnesol X receptor( FXR) pathway,sixty SD rats were randomly divided into 6 groups:blank control group,ANIT model group,ursodeoxycholic acid group( 100 mg·kg~(-1)·d-1 UDCA),and GPA high,medium and low( 100,50 and 25 mg·kg~(-1)·d-1) dosage groups,10 rats in each group. Corresponding drugs were intragastrically( ig) administered for10 days. After administration on day 8,all rats except blank rats were administered with 65 mg·kg~(-1)α-naphthalene isothiocyanate( ANIT) once. After the last administration,the serum levels of alanine aminotransferase( ALT),glutamine oxalacetate aminotransferase( AST),gamma-glutamyltransferase( γ-GGT),alkaline phosphatase( ALP),total bilirubin( TB) and total bile acid( TBA)were measured,and the mRNA transcription levels of Sirt1,FXR,multidrug resistant associated protein 2( MRP2),bile salt export pump( BSEP),sodium taurocholate contractible polypeptide( NTCP) in liver and apical sodium bile acid transporter( ASBT),ileum bile acid binding protein( IBABP) in ileum were detected by reverse transcription-polymerase chain reaction( RT-PCR). The protein expression levels of Sirt1,FXR and NTCP were detected by Western blot; the expression of MRP2,BSEP in liver and ASBT,IBABP in ileum were determined by immunofluorescence three staining. Primary rat hepatocytes were cultured in vitro to investigate the inhibitory effect of GPA on a potent and selective Sirt1 inhibitor( EX 527),and the mRNA and protein expression levels of Sirt1 and FXR were detected by RT-PCR and Western blot. GPA significantly decreased the levels of ALT,AST,γ-GGT,ALP,TB,TBA in serum( P<0.01) and improved the pathological damage of liver tissues in ANIT-induced cholestasis rats; significantly increased the mRNA and protein expression levels of Sirt1,FXR,MRP2,BSEP,NTCP in liver and ASBT,IBABP in ileum( P< 0.01). In vitro primary hepatocytes experiment indicated that the gene and protein expression levels of FXR and Sirt1 were noticeably improved by GPA in primary hepatocytes inhibited by EX-527( P<0.01). It was found that the improvement of GPA was in a dose-dependent manner. GPA could improve bile acid hepatointestinal circulation and play a liver protection and cholagogu role in cholestasis rats induced by ANIT.The mechanism may be that GPA activated FXR by regulating Sirt1,a key regulator of oxidative stress injury,and then the activated FXR could regulate protein of bile acid hepato-enteric circulation.
Animals ; Cholestasis ; Iridoid Glucosides ; Liver ; Rats ; Rats, Sprague-Dawley ; Receptors, Cytoplasmic and Nuclear ; Signal Transduction ; Sirtuin 1

Objective To explore whether perioperative bright light therapy could inhibit the occurrence of postoperative delirium in tumor patients undergoing open hepatobiliary surgery. Methods Totally120 elderly tumor patients scheduled for open hepatobiliary surgery and postoperative ICU treatment were recruited and randomized into bright light and control group with 60 cases per group in accordance with the random number table.Bright light was delivered to the patients at an intensity of 10 000 lux from 2 days before surgery to postoperative day 7.Each intervention began at 7 am and lasted for 2 hours.Delirium was screened using the Confusion Assessment Method for Intensive Care Unit(CAM-ICU) during the first 7 days after surgery. The differences in the incidence of postoperative delirium, the duration of delirium and the length of ICU as well as postoperative hospital stay were compared. Results Demographic characteristics and surgical outcomes were similar between groups. The incidence of postoperative delirium were 46.67%(28/60)and 23.33%(14/60)for control group and bright light group respectively with a significant difference (χ2=7.179,P=0.007). The difference was seen on postoperative days 3 (χ2=5.187, P = 0.023) and 4 (χ2=8.749,P = 0.003). The incidences of delirium on postoperative days 3 for bright light and control group were 5.08%(3/59)and 18.64%(11/59)respectively.On postoperative day 4, the incidences of delirium for bright light and control group were 0 (0/58) and 14.04% (8/57) respectively.There was no difference in the duration of delirium(χ2=1.248,P=0.264),the length of ICU stay (χ2=0.036,P=0.849)or the hospital stay(χ2=1.706,P=0.192). Conclusion Bright light is useful in preventing postoperative delirium in elderly tumor patients undergoing open hepatobiliary surgery.

OBJECTIVETo investigate the effect of clopidogrel on the binding rate of ginsenosides with rat serum proteins (RSA).

METHODSEquilibrium dialysis and liquid chromatography-mass spectrometry were employed to quantify the concentration of ginsenoside Rg1 and Rb1. The protein-binding rates of Rg1 and Rb1 in the presence or absence of clopidogrel (1.0 mg/L) were determined. A molecular simulation model (consisting of homology modeling and molecular docking interaction) was used to reveal the target protein-compound interactions.

RESULTSThe binding rates of ginsenosides Rg1 (0.4, 1.0, and 2.0 mg/L) with RSA were (30.16∓2.82)%, (33.42∓4.21)%, and (34.61∓3.42)%, and those of and Rb1 were (50.13∓2.34)%, (51.23∓3.23)%, and (53.11∓3.26)%, respectively. In the presence of clopidogrel, the binding rates of Rg1 decreased to (22.13∓2.72)%, (21.42∓3.22)%, and (25.45∓3.52)%, and those of Rb1 to (40.13∓3.24)%, (41.25∓4.15)%, and (43.11∓3.31)%, receptively. The molecular docking suggested that these compounds competed to bind with RSA.

CONCLUSIONClopidogrel can competitively bind to RSA with ginsenosides to lower the plasma protein binding rates of ginsenosides.

To explore the effect of geniposidic acid (GPA) on the signal pathway of small heterodimer dimer receptor (SHP) and liver receptor homologue 1 (LRH-1) in cholestasis rats induced by alpha-naphthalene isothiocyanate (ANIT).
 Methods: Fifty SD rats were randomly divided into five groups: a blank group, an ANIT group, an ANIT+GPA (100 mg/kg) group, an ANIT+GPA (50 mg/kg) group, and an ANIT+GPA (25 mg/kg) group (n=10 in each group). The GPA were intragastrically given to rats for 10 days, and the control group and the ANIT group were given normal saline. At the eighth day of administration, all rats except the blank group were given 65 mg/kg ANIT once until the tenth day. After the last administration, serum total cholesterol (TC), triglyceride (TG) and total bile acids (TBA) were measured. The primary hepatocytes (RPH) were isolated from normal rats and cultured. The cells were divided into a blank group, an ANIT (40 μmol/L) group, an ANIT (40 μmol/L)+GPA (4.00 mmol/L) group (A4.00G group), an ANIT (40 μmol/L)+GPA (1.00 mmol/L) group (A1.00G group), and an ANIT (40 μmol/L)+GPA (0.25 mmol/L) group (A0.25G group). The mRNA transcription levels of SHP and cholesterol 7 alpha hydroxylase (CYP7A1) in RPH were detected by real-time-PCR, and the protein levels of SHP and CYP7a1 were detected by Western blotting. In the LRH-1 silence experiment, the RPH were divided into a blank group, a negative transfection group, a siRNA-LRH group (ZR group), a siRNA-LRH+GPA (4.00 mmol/L) group (ZR4.00G group), a siRNA-LRH+GPA (1.00 mmol/L) group (ZR1.00G group) and a siRNA-LRH+GPA (0.25 mmol/L) group (ZR0.25G group). The protein and mRNA levels of SHP, CYP7a1, LRH-1 were detected. In the over-expression experiment, the RPH were also divided into a blank group, a negative transfection group, a LRH-1 over-expression plasmid group (OE group), a LRH-1 over-expression plasmid+GPA (4.00 mmol/L) group (OE4.00G group), a LRH-1 over-expression plasmid+GPA (1.00 mmol/L) group (OE1.00G group), and a LRH-1 over-expression plasmid+GPA (0.25 mmol/L) group (OE0.25G group). The protein and mRNA levels of SHP, CYP7a1 and LRH-1 were detected.
 Results: Compared with the blank control group, TC and TBA were significantly increased (both P<0.01) in the ANIT group, but there was no difference in TG; compared with the ANIT group, the contents of TC and TBA in the AG100 and AG50 groups were significantly reduced (all P<0.01). Compared with the blank control group, the proteins and mRNA levels of SHP were significantly decreased (P<0.01), while CYP7a1 were dramatically increased (P<0.01) in the ANIT group; compared with the ANIT group, the proteins and mRNA levels of SHP in the A4.00G group and the A1.00G group were significantly increased (both P<0.01), while the levels of CYP7a1 proteins and mRNA levels were evidently decreased in the A4.00G and A1.00G groups (both P<0.01). Compared with the negative transfection group, the proteins and mRNA levels of CYP7a1 and LRH-1 were dramatically restrained (all P<0.01), while there was no change in SHP in the ZR group; compared with the ZR group, the proteins and mRNA levels of SHP were significantly increased (all P<0.01), while LRH-1 and CYP7a1 were not changed in the ZR4.00G, ZR1.00G and ZR0.25G groups. Compared with the negative transfection group, the protein and mRNA levels of CYP7a1 and LRH-1 were significantly suppressed in the OE group (all P<0.01). Compared with the OE group, the protein and mRNA levels of SHP were evidently increased in the OE4G and OE1G groups (all P<0.01), while LRH-1 and CYP7a1 were not changed in the OE4G, OE1G and OE0.25G groups.
 Conclusion: The over-expression of LRH-1 in RPH can up-regulate the mRNA and protein levels of CYP7a1. GPA can improve the biochemical and liver pathology of ANIT-induced cholestasis rats, which may be related to the decrease of CYP7a1 by activating SHP through LRH-1 in RPH.
Animals ; Cholestasis ; Iridoid Glucosides ; Rats ; Rats, Sprague-Dawley ; Receptors, Cytoplasmic and Nuclear ; Signal Transduction

Zn²⁺ is required for the activity of many mitochondrial proteins, which regulate mitochondrial dynamics, apoptosis and mitophagy. However, it is not understood how the proper mitochondrial Zn²⁺ level is achieved to maintain mitochondrial homeostasis. Using Caenorhabditis elegans, we reveal here that a pair of mitochondrion-localized transporters controls the mitochondrial level of Zn²⁺. We demonstrate that SLC-30A9/ZnT9 is a mitochondrial Zn²⁺ exporter. Loss of SLC-30A9 leads to mitochondrial Zn²⁺ accumulation, which damages mitochondria, impairs animal development and shortens the life span. We further identify SLC-25A25/SCaMC-2 as an important regulator of mitochondrial Zn²⁺ import. Loss of SLC-25A25 suppresses the abnormal mitochondrial Zn²⁺ accumulation and defective mitochondrial structure and functions caused by loss of SLC-30A9. Moreover, we reveal that the endoplasmic reticulum contains the Zn²⁺ pool from which mitochondrial Zn²⁺ is imported. These findings establish the molecular basis for controlling the correct mitochondrial Zn²⁺ levels for normal mitochondrial structure and functions.
Animals ; Caenorhabditis elegans/metabolism* ; Cation Transport Proteins/genetics* ; Homeostasis ; Mitochondria/metabolism* ; Zinc/metabolism*