Puerarin prevents sepsis-associated encephalopathy by regulating the AKT1 pathway in microglia.

BACKGROUND: Previous studies have reported that puerarin possesses cardioprotective, vasodilatory, anti-inflammatory, anti-apoptotic, and hypoglycemic properties. However, the impact of puerarin on sepsis-associated encephalopathy (SAE) remains unexplored. In this study, we explored whether puerarin can modulate microglia-mediated neuroinflammation for the treatment of SAE and delved into the underlying mechanisms. METHODS: We established a murine model of SAE through intraperitoneal injection of lipopolysaccharide (LPS). The puerarin treatment group received pretreatment with puerarin. For in vitro experiments, BV2 cells were pre-incubated with puerarin for 2 h before LPS exposure. We employed network pharmacology, the Morris Water Maze (MWM) test, Novel Object Recognition (NOR) test, immunofluorescence staining, enzyme-linked immunosorbent assay (ELISA), Western blotting, and quantitative real-time PCR (qRT-PCR) to elucidate the molecular mechanism of underlying puerarin's effects in SAE treatment. RESULTS: Our findings demonstrate that puerarin significantly reduced the production of inflammatory cytokines (TNF-α and IL-6) in the peripheral blood of LPS-treated mice. Moreover, puerarin treatment markedly ameliorated sepsis-associated cognitive impairment. Puerarin also exhibited inhibitory effects on the release of TNF-α and IL-6 from microglia, thereby preventing hippocampal neuronal cell death. Network pharmacology analysis identified AKT1 as a potential therapeutic target for puerarin in SAE treatment. Subsequently, we validated these results in both in vitro and in vitro experiments. Our study conclusively demonstrated that puerarin reduced LPS-induced phosphorylation of AKT1, with the AKT activator SC79 reversing puerarin's anti-inflammatory effects through the activation of the AKT1 signaling pathway. CONCLUSION: Puerarin exerts an anti-neuroinflammatory effect against SAE by modulating the AKT1 pathway in microglia.

[Clinical research of Pinggan Jiangya decoction combined with penetrating needling at Baihui (GV20) in a period of day from 7 am to 9 am in the treatment of grade 1 and 2 essential hypertension].

OBJECTIVE: To explore the clinical effect of Pinggan Jiangya decoction combined with penetrating needling at Baihui (GV20) in a period of day from 7 am to 9 am in the treatment of grade 1 and 2 essential hypertension (EH). METHODS: A total of 150 cases of grade 1 and 2 EH patients were randomized into an observation group and a control group, 75 cases in each group. In the control group, Pinggan Jiangya decoction was prescribed for oral administration one dose a day, while in the observation group, on the basis of the treatment as the control group, penetrating needling was exerted at GV20 once daily. The treatment duration was 8 weeks. Before and after treatment, the TCM syndrome score, 24 h average systolic blood pressure (24 h ASBP), 24 h average diastolic blood pressure (24 h ADBP), 24 h average pulse pressure difference (24 h PP), morning blood pressure surge (MBPS), 24 h SBP variability (24 h SBPV), 24 h DBP variability (24 h DBPV), serum levels of 5-hydroxytryptamine (5-HT) and melatonin (MT) were compared in the patients of the two groups. The clinical therapeutic effect was observed in the two groups. RESULTS: After the treatment, in the self-comparison of each group, the scores of headache, vertigo, backache, soft knees, tinnitus, 24 h ASBP， 24 h ADBP, 24 h PP, MBPS, 24 h SBPV and 24 h DBPV in the two groups were lower than those before treatment (P<0.01), and the above indexes in the observation group were lower than those in the control group (P<0.01). The level of serum 5-HT after the treatment was lower than that of before the treatment (P<0.01), while the level of MT was higher than that of before the treatment (P<0.01) in both two groups, and the level of 5-HT in the observation group was lower than that of the control group, while the level of MT was higher than that of the control group (P<0.01). The total effective rate of the observation group was 93.3% (70/75), better than 76.0% (57/75) of the control group (P<0.01). CONCLUSION: Pinggan Jiangya decoction combined with penetrating needling at GV20 in a period of day from 7 am to 9 am can regulate the levels of serum MT and 5-HT, effectively reduce blood pressure, improve blood pressure variability, control morning peak blood pressure, and has a remarkable effect in the treatment of grade 1 and 2 EH.

Paeoniflorin Improved Constipation in the Loperamide-Induced Rat Model via TGR5/TRPA1 Signaling-Mediated 5-Hydroxytryptamine Secretion.

Slow transit constipation (STC) is a common type of constipation with a high incidence rate and a large number of patients. We aimed to investigate the therapeutic effects and potential mechanism of paeoniflorin (PAE) on loperamide-induced Sprague Dawley (SD) rat constipation models. Rats with loperamide-induced constipation were orally administered different concentrations of PAE (10, 20, or 40 mg/kg). In vitro, enterochromaffin (EC)-like RIN-14B cells were treated with 20, 40, or 80 µg/ml PAE. We found that PAE treatment significantly improved the symptoms of constipation and increased the intestinal transit rate. Hematoxylin and eosin (H&E) staining showed that PAE alleviated colonic tissue pathological damage. Besides, our results implied that PAE concentration-dependently promoted the content of 5-hydroxytryptamine (5-HT) catalyzed by tryptophan hydroxylase (Tph)-1 in the serum of loperamide-induced rats and in RIN-14B cells. Western blot and immunofluorescence (IF) stain indicated that PAE also promoted the expression of G protein-coupled BA receptor 1 (TGR5), transient receptor potential ankyrin 1 (TRPA1), phospholipase C (PLC)-γ1, and phosphatidylinositol 4,5-bisphosphate (PIP2) in vivo and in vitro. RIN-14B cells were cotreated with a TGR5 inhibitor (SBI-115) to explore the mechanism of PAE in regulating the 5-HT secretion. We observed inhibition of TGR5 reversed the increase of 5-HT secretion induced by PAE in RIN-14B cells. We provided evidence that PAE could promote 5-HT release from EC cells and improve constipation by activating the TRPA1 channel and PLC-γ1/PIP2 signaling. Thus, PAE may provide therapeutic effects for patients with STC.

A Review on Classification and Biological Activities of Alkaloids from the Genus Zanthoxylum Species.

Many plants in the genus Zanthoxylum, belonging to the Rutaceae family, are used as folk medicines for the treatment of various diseases, which have gained much attention for their phytochemical and pharmacological activity investigations. Alkaloids are the largest secondary metabolites with structurally diverse types found in this genus and they demonstrate a wide range of biological activities. The aim of this review is to provide a summary on the isolation, classification, and biological properties of alkaloids from Zanthoxylum species, which also will bring more attention to other researchers for further biological study on alkaloids for the new drug development.

Penicacids E-G, three new mycophenolic acid derivatives from the marine-derived fungus Penicillium parvum HDN17-478.

Three new mycophenolic acid derivatives, penicacids E-G (1-3), together with three known analogues, mycophenolic acid (4), 4'-hydroxy-mycophenolic acid (5) and mycophenolic methyl ester (6), were isolated from a marine-derived fungus Penicillium parvum HDN17-478 from a South China Sea marine sediment sample. The structures of compounds 1-3 were elucidated by HRMS, NMR, and Mosher's method. Among them, compounds 1 and 2 were the first examples of mycophenolic acid analogs with a double bond at C-3'/C-4' position. The cytotoxicity of 1-6 was evaluated against the HCT-116, BEL-7402, MGC-803, SH-SY5Y, HO-8910 and HL-60 cell lines, and compounds 4 and 6 showed potent cytotoxicity with IC50 values ranging from 1.69 to 12.98 µmol·L-1.

[Poor weight gain, recurrent metabolic alkalosis and hypokalemia in a neonate].

The study reports a female neonate with a gestational age of 29+2 weeks and a birth weight of 1 210 g. Ten minutes after birth, the neonate was admitted to the hospital due to shortness of breath. Several days after birth, the neonate presented with hyperglycemia, polyuria, and poor weight gain, accompanied by azotemia, hypochloremic metabolic alkalosis, hypokalemia, and hyponatremia. Laboratory examinations showed elevated levels of aldosterone, renin, and angiotensin II. Gene detection revealed SLC12A1 gene mutation. Neonatal Bartter syndrome was thus confirmed. The neonate was treated with sodium and potassium supplements, and was followed up for 8 months. During the follow-up, the mental and neural development of the neonate was almost normal at the corrected age, and regular reexaminations showed slight metabolic alkalosis and almost normal electrolyte levels. For the neonates who have the symptoms of unexplainable polyurine and electrolyte disorders, it is important to examine the levels of aldosterone, renin and angiotensin. A definite diagnosis of neonatal Bartter syndrome can be made based on the presence of SLC12A1 gene mutation.

[Investigation of Variations of Components in Euphorbia pekinensis Root and Its Processed Products Based on Plant Metabolomics].

Objective: An ultra-performance liquid chromatography coupled with mass spectrometry( UPLC-MS) based on plant metabonomics method was proposed and developed for the investigation of variations of components in Euphorbia pekinensis root after herb-processing procedure. Methods: The samples were separated on an Agilent XDB C8column( 150 mm × 4. 6 mm,5 µm) using a mobile phase composed of 0. 1% formic acid in acetonitrile and 0. 1% formic acid in water under gradient elution. Analysis was performed with electrospray ionization( ESI) interface in negative ion mode within the m / z range of 100 ~ 900. The data were subjected to principal component analysis( PCA). Results: Seven components were screened out which could be considered as potential chemical markers to discriminate Euphorbia pekinensis root from its processed products. Six of the chemical markers were identified as 3,3'-di-O-methylellagic acid-4'-O-ß-D-xylopyranoside,3,3'-di-O-methyl ellagic acid,(-)-( 1S)-15-hydroxy-18-carboxycembrene,ellagic acid,brevifolincarboxylic acid and gallic acid. Conclusion: The method has been successfully applied in distinguishing Euphorbia pekinensis root from its processed products,which predicts the potential substances contributed to the toxicity reducing effect of traditional processing procedure on the herb.

Assessment of Cuscuta chinensis seeds׳ effect on melanogenesis: comparison of water and ethanol fractions in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Cuscuta chinensis seeds have traditionally been used to treat freckles and melasma in Asia, although recent reports have revealed that Semen cuscutae is a promoter of melanogenesis. The present study aims to investigate the mechanism of this opposite effect of Semen cuscutae on melanogenesis. MATERIALS AND METHODS: In accordance with traditional usage, the water fraction and the ethanol fraction from Semen cuscutae (WFSC/EFSC) were extracted to determine the herbal effects by examining the activity of mushroom tyrosinase, cellular melanin contents, tyrosinase activity assay, quantitative-reverse transcription polymerase chain reaction (qRT-PCR), and Western blot analysis for tyrosinase in B16F10 mouse melanoma cells. The melanocyte phenotypes of zebrafish larvae were observed while the in vivo melanin contents and tyrosinase activity were determined. RESULTS: The activity of mushroom tyrosinase assay shown that WFSC was an uncompetitive inhibitor of mushroom tyrosinase, while EFSC indicated dose-dependent activation of the mushroom tyrosinase activity. The WFSC markedly inhibited 3-isobutyl-1-methylxanthine (IBMX)-stimulated melanin synthesis and tyrosinase activity in vitro. Howeveran accelerant role in melanin synthesis and tyosinase activity. Neither fraction had any effect on the IBMX-induced expression of tyrosinase protein or mRNA. The WFSC strongly inhibited melanin synthesis and cellular tyrosinase activity in vivo. Furthermore, with the function of WFSC at a higher concentration, a punctate melanocyte pattern appeared that was similar to the pattern induced by arbutin or Mequinol (MQ). The EFSC had no effect on the melanocytes of zebrafish larvae. It was discovered that WFSC did not show a stable inhibitory effect until it was extracted 1 month later. CONCLUSIONS: These results suggest that the opposite effects of Cuscuta chinensis seeds were caused by the extraction methods and that time has an important role on the effect of WFSC. Both WFSC and EFSC significantly influence melanogenesis by regulating enzymatic activity of tyrosinase. In addition, the data indicate that wildtype (WT) zebrafish may be an ideal model for testing inhibitors of melanogenesis from clinically active herbs.

Ligustrazine attenuates elevated levels of indoxyl sulfate, kidney injury molecule-1 and clusterin in rats exposed to cadmium.

In this study, we aimed at evaluating the effect of ligustrazine, a major constituent of Ligusticum wallichii from traditional Chinese medicine, on Cd-induced changes in nephrotoxicity indices. Rats were divided into four experimental groups: control; ligustrazine; Cd and ligustrazine+Cd. Cd treated alone group showed significant decreases (P<0.05) in body weight, renal levels of superoxide dismutase (SOD) and glutathione reductase (GR); and significant increases (P<0.05) in urine volume (24h), pH values, serum blood urea nitrogen (BUN), serum uric acid, kidney malondialdehyde (MDA), urinary total protein, urinary glucose, urinary lactate dehydrogenase (LDH) and urinary alkaline phosphatase (ALP). Apart from indoxyl sulfate (a uremic toxin), two newly accepted nephrotoxicity biomarkers including kidney injury molecule-1 (kim-1) and clusterin were also found to be increased. Nonetheless, all these effects induced by Cd were reversed upon treatment by ligustrazine although it failed in decreasing the concentrations of Cd in kidney and urine. Histopathological studies in Cd-treated rats exhibited renal tubule damage, which was also ameliorated by ligustrazine pretreatment. These results suggest that ligustrazine exhibits protective effects on Cd-induced nephrotoxicity. Additionally, this study also demonstrates Cd exposure induces elevated levels of indoxyl sulfate in serum and kidney, and clusterin in urine.

[ILs-HPLC simultanesous determination of five alkaloids in phellodenddri chinensis cortex].

A RP-HPLC method was established for simultaneous determination of phellodendrine hydrochloride (PH1), magnoflorine hydrochloride (MH), jatrorrhizine hydrochloride (JH), palmatine hydrochloride (PH2) and berberine hydrochloride (BH) in Phellodendri Chinensis Cortex by using ionic liquids as mobile phase additives. The separation was performed on a Kromasil C18 (4.6 mm x 250 mm, 5 µm) coupled with ultraviolet (UV) detection. The effect of extraction solvent, detection wavelength, length of alkyl chain on different imidazolium ionic liquids and concentration of ionic liquids on the separation and determination of alkaloids were investigated. Ionic liquid, [BMIm] BF4, can obviously improve the resolution and peak shape. This ILs-HPLC method is simple, rapid, and reliable, which can be used for determination of alkaloids in Phellodenddri Chinensis Cortex.

[Homoharringtonine combined arsenic trioxide induced apoptosis in human multiple myeloma cell line RPMI 8226: an experimental research].

OBJECTIVE: To clarify the effects and mechanisms of homoharringtonine (HHT) monomer therapy or combination therapy with arsenic trioxide (ATO) on human multiple myeloma (MM) cell line RPMI 8226 in in vitro researches. METHODS: Effects of HHT, ATO, and HHT combined ATO on the growth of MM cell line RPMI 8226 were detected using MTT assay. The morphological changes of cell apoptosis were detected by Hoechst staining. The early apoptosis rate was detected using flow cytometry. Expressions of Caspase-3, Caspase-9, poly-ADP-ribose polymerase (PARP), Bcl-2, Mcl-1, Bcl-xl, and AKT protein were detected by Western blot. RESULTS: HHT and ATO inhibited the proliferation of RPM1 8226 cell line in a time- and dose-dependent manner (P < 0.05). Synergistic effects was shown in the combination group (Cl < 1). HHT and ATO induced the apoptosis of RPMI 8226 in a dose-dependent manner with typical morphological changes of apoptosis and higher early stage apoptosis rate. The enhancement in apoptotic induction was seen when two agents were combined. HHT activated expressions of Caspase-3 and PARP in a dose dependent manner at 24 h. HHT at 40 ng/mL and ATO at 8.5 micromol/L could significantly activate expressions of Caspase-3 and Caspase-9, and down-regulate expressions of anti-apoptotic proteins Bcl-xl and Mcl-1. In addition, the combination therapy of HHT at 40 ng/mL and ATO at 8.5 micromol/L inhibited phosphorylation of AKT in a time-dependent manner. CONCLUSION: HTT, ATO, and combination therapy of HHT and ATO induced the apoptosis of RPMI 8226 cell line possibly through activating Caspase pathways, regulating expressions of Bcl-2 families, and inhibiting phosphorylation of AKT.

Elution-extrusion counter-current chromatography separation of two new benzyl ester glucosides and three other high-polarity compounds from the tubers of Pleione bulbocodioides.

INTRODUCTION: The tubers of Pleione bulbocodioides (Franch.) Rolfe, with gastrodin and benzyl ester glucosides as main components, have been used in traditional Chinese medicine for the treatment of various cancers and bacterial infections. Up to now, their official quality control method is still inadequate, and the difficulty of obtaining these high-polarity compounds is one of the major reasons. OBJECTIVE: To develop a rapid and efficient method for preparative separation of the high-polarity compounds gastrodin and benzyl ester glucosides. METHODS: An optimised solvent system composed of n-butanol:ethanol:water (20:1:20, v/v/v) was applied for the elution-extrusion counter-current chromatography (EECCC) separation. The upper phase was used as the stationary phase, and the lower phase was used as the mobile phase at a flow rate of 1.5 mL/min, a rotation speed of 850 rpm and a temperature of 35°C. RESULTS: Five high-polarity glucosides, including two new compounds, (E)-4-ß-D-glucopyranosyloxycinnamic acid 9-(4-ß-D-glucopyranosyloxybenzyl) ester (4 mg) and (Z)-2-(2-methylpropyl)butenedioic acid bis(4-ß-D-glucopyranosyloxybenzyl) ester (9 mg), and three main components, gastrodin (87 mg), dactylorhin A (60 mg) and militarine (15 mg), with HPLC purities of 95.4%, 96.4%, 91.1%, 97.2% and 95.5% respectively, were yielded from 400 mg of the prepared sample. CONCLUSION: Elution-extrusion counter-current chromatography could be used as a useful tool for the separation of high-polarity compounds such as gastrodin and benzyl ester glucosides and the enrichment of the minor ones.

[Comparative study on pharmacokinetics of senkyunolide I after administration of simple recipe and compound recipe in rats].

OBJECTIVE: To develop a LC-MS method for the determination of senkyunolide I (SI) in rat plasma, in order to observe whether there is significant change in the pharmacokinetics parameters of complex prescriptions of Huoluoxiaolingdan (HLXL) and single herbal extracts from Ligusticum chuanxiong Hort. in rats, and assess the effect of other components in HLXL on the pharmacokinetics of SI. METHOD: Twelve male Sprague-Dawley (SD) rats were randomly divided into two groups, and orally administered with extract from HLXL and L. chuanxiong (both equal to SI 4.53 mg x kg(-1)). Their blood was collected at different time points for LC-MS, in order to detect the plasma concentration of SI. The pharmacokinetic parameters of SI were calculated by DAS 2.0 software. SPSS 16.0 software was used for independent-sample T-test and Nonparametric T-test. RESULT: A linear relationship of SI ranged from 6.750 to 675.0 microg x L(-1), and with the lowest limit of detection being 6.750 microg L(-1). Both of the plasma concentration-time curves of SI were fitted with the two-compartment model for extract of HLXL and L. chuanxiong. The detected AUC and Cmax of SI showed significant difference, with no significant difference in other parameters. CONCLUSION: The LC-MS determination method established in this experiment was so exclusive, accurate and sensitive that it is suitable for pharmacokinetic studies on extracts of HLXL and SI from L. chuanxion. The experiment results show that other ingredients of HLXL have noticeable effect on the absorption of SI in rat plasma.

[Screening of hepatotoxicity fraction of Genkwa Flos and study on UPLC fingerprint of hepatotoxicity fraction].

OBJECTIVE: To look for the active fraction of ethanol extract of Genkwa Flos (EGF) induced hepatotoxicity and develop an UPLC fingerprint of the active fraction. METHOD: Target fraction of EGF induced hepatotoxicity was guided by the serum biochemical and histopathology methods. The UPLC method was applied to establish the chromatographic fingerprint. The separation was achieved on a BEH C18 column (2.1 mm x 50 mm, 1.7 microm) with a mobile phase consisting of acetonitrile and water containing 0.05% phosphate acid running gradient elution. The detection was carried out at 210 nm and the analysis was finished within 10 min. RESULT: The chloroform phase of EGF could be responsible for the hepatotoxicity of this herb. The common mode of the UPLC fingerprint was set up under the established condition. There were 17 common peaks in fourteen batches of herbs, eight of which were identified, and the similar degrees of the fourteen batches to the common mode were between 0.890-0.999. CONCLUSION: It is easy to locate the chloroform extraction of EGF with hepatotoxicity. And the UPLC fingerprint was developed for the above fraction, which could provide valuable references for safe and effective clinical use of EGF.

Neolignanamides, lignanamides, and other phenolic compounds from the root bark of Lycium chinense.

Seven new neolignanamides (1-7), including two pairs of cis- and trans-isomers, and a new lignanamide (8) were isolated from the EtOAc-soluble fraction of an EtOH extract of the root bark of Lycium chinense, together with 22 known phenolic compounds (9-30), four of which were obtained from the genus Lycium for the first time. Compounds 5, 6, and 7 are unusual dimers having a rare connection mode between the two cinnamic acid amide units, while compounds 6, 7, and 8 are the first naturally occurring dimers derived from two dissimilar cinnamic acid amides. The cinnamic acid amides, neolignanamides, and lignanamides possess moderate radical-scavenging activity against the DPPH (2,2-diphenyl-1-picrylhydrazyl) and superoxide radicals.

Simultaneous quantification of three iridoids in rat plasma after oral administration of Zhi-zi-chi decoction using LC-MS.

A sensitive and efficient liquid chromatography-mass spectrometry (LC-MS) method was developed and validated for the simultaneous determination of geniposide, 6α-hydroxygeniposide, and genipin gentiobioside in rat plasma. After the addition of internal standard (I.S.) salidroside and acidification (formic acid, 0.1%), plasma samples were carried out by protein precipitation with acetonitrile and separated on a Kromasil C(18) column (200 mm × 4.6 mm, 5 µm) within a runtime of 15.0 min. The linear ranges were 2-250 ng/mL for both 6α-hydroxygeniposide and genipin gentiobioside and 2-2000 ng/mL for geniposide, respectively. The lower limit of quantification (LLOQ) was 2 ng/mL for all the analytes. The validated method was successfully applied to the pharmacokinetics study of geniposide, 6α-hydroxygeniposide, and genipin gentiobioside in rats after oral administration of Zhi-zi-chi decoction.

A metabonomic analysis of urine from rats treated with rhizoma alismatis using ultra-performance liquid chromatography/mass spectrometry.

A metabonomic approach based on ultra-performance liquid chromatography coupled to mass spectrometry (UPLC/MS) was used to study the nephrotoxicity of rhizoma alismatis (RA) in rats. Potential biomarkers of RA toxicity were identified and the toxicological mechanism is discussed. Urine samples were collected from control and treated rats at various stages and analyzed by UPLC/MS in positive ionization mode. Histopathological analysis was used to evaluate renal function. The differences in the metabolic profiles of the control and treated rats were clearly distinguishable with principal components analysis (PCA) of the chromatographic data, and significant changes in 13 metabolite biomarkers were detected in the urine. This metabonomic method combined with PCA could discriminate the treated rats from the control rats on days 60, 120, and 180 after treatment, before serious organic renal damage was apparent on day 180 with histopathology. This research indicates that UPLC/MS-based metabonomic analysis of urine samples can be used to predict the chronic nephrotoxicity induced by rhizoma alismatis.

LC-MS determination and pharmacokinetic study of salidroside in rat plasma after oral administration of suspensions of traditional Chinese medicine Erzhi Wan and Fructus Ligustri lucidi.

A simple, rapid and sensitive liquid chromatography-mass spectrometry (LC-MS) method was developed for the determination of salidroside in rat plasma and study of its pharmacokinetics after oral administration of suspension of Erzhi Wan and Fructus Ligustri lucidi into Wistar rats. Plasma sample of 200 µL was extracted with acetic ether-isopropanol (2:1) and the extraction was performed on a Kromasil C18 column (150 mm × 4. 6 mm, 5 µm) with the mobile phase of methanol-water (41:59, v/v) within a run time of 6.0 min. The analyte was monitored with positive electrospray ionization (ESI) by selected ion monitoring (SIM) mode. The target ions were m/z 323.05 for salidroside and m/z 411.05 for internal Standard (IS) geniposide. A good linear relationship was obtained over the range of 5.0-500.0 ng/mL and the lower limit of quantification was 5.0 ng/mL. The validated method was successfully applied to the pharmacokinetic study of salidroside in rat plasma after oral administration of suspension of Erzhi Wan and Fructus Ligustri lucidi.

TNF-alpha induces mitochondrial dysfunction in 3T3-L1 adipocytes.

TNF-alpha was the first proinflammatory cytokine identified linking obesity, insulin resistance and chronic inflammation. However, the mechanism of TNF-alpha in the etiology of insulin resistance is still far from clear. Because the mitochondria play an important role in energy metabolism, we investigated whether mitochondrial dysfunction is involved in pathogenesis of TNF-alpha-mediated insulin resistance. First, a fully differentiated insulin-resistant 3T3-L1 adipocyte model was established by incubating with 4 ng/ml TNF-alpha for 4 d, and then the mitochondrial morphology and functions were observed. TNF-alpha treatment induced pronounced morphological changes in the mitochondria, which became smaller and condensed, and some appeared hollow and absent of cristae. Mitochondrial dynamics changes were observed as increased mitofusion protein mfn1 and mitofission protein Drp1 levels compared with controls. No obvious effects on mitochondrial biogenesis were found. PGC-1alpha levels decreased, but no significant changes were found in mtTFA mRNA expression, NRF1mRNA expression and mitochondrial DNA (mtDNA). TNFalpha treatment also led to decreased mitochondrial membrane potential and reduced production of intracellular ATP, as well as accumulation of significant amounts of reactive oxygen species (ROS). Further research is required to determine if mitochondrial dysfunction is involved in the inflammatory mechanism of insulin resistance and may be a potential target for the treatment of insulin resistance.

[Determination of the binding rate of rat plasma protein with salvianolic acid B].

This paper is aimed to report the development of a method for the determination of the binding rate of plasma protein with salvianolic acid B. In vitro, equilibrium dialysis method was used to imitate the binding process between salvianolic acid B and plasma protein, in vivo, ultrafiltration method was used and the binding rate with HPLC was determined. Plasma samples were treated with methanol to precipitate the protein, and the buffer solution was directly determined after filtering. The calibration curve of the buffer solution was linear in the range of 0.5-20 microg mL(-1). The calibration curve of the plasma was linear in the range of 2-200 microg mL(-1). The extract recovery was 68.6%-81.9%. RSDs of intra- and inter-day precisions were all less than 8.5%. The binding rates of plasma protein with salvianolic acid B in vitro was 75.2% and in vivo was 92.1%. This paper shows the high binding power of salvianolic acid B to plasma protein with high sensitivity, good reproduction, simple management and fulfilling the requirement.