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Histidine phosphorylation (pHis) was first reported in 1962. There are few studies on pHis because of the thermal and acidic instability of pHis and the lack of specific methods to detect it. pHis has two isomers of 1-phosphate histidine (1-pHis) and 3-phosphate histidine (3-pHis). pHis antibodies have been developed recently and have promoted research in this field. In this study, we established a CCl4-induced liver fibrosis model in C57 mice and a TGF-β1-induced HSC activation model in LX-2 cells, to study the role of histidine phosphorylation. The expression of histidine kinases NME1 and NME2 was increased, histidine phosphatase PGAM5 and PHPT1 was unchanged, and 1-pHis and 3-pHis were increased in the in vivo and in vitro models. The expression of LHPP was decreased in the in vivo model but not in the in vitro model. To further study the role of NME1, NME2, and histidine phosphorylation in HSC activation, we silenced NME1 or NME2 and administered TGF-β1 in LX-2 cells. The results showed silencing NME1 or NME2 decreased TGF-β1-induced pHis levels and the expression of α-SMA and COL1A1, indicating the activation of HSC was suppressed. Then, we found the inhibitory effect on HSC activation is due to reduced phosphorylation of Smad2 and Smad3. In summary, our studies indicate that NME1 and NME2 are involved in TGF-β1-induced HSC activation and CCl4-induced liver fibrosis, which may be mediated by histidine phosphorylation.

Purpose: SET and MYND domain-containing protein2 (SMYD2), a histone lysine methyltransferases, is a candidate human oncogene in multiple tumors. However, the expression dynamics of SMYD2 in hepatocellular carcinoma (HCC) and its clinical/prognostic significance are unclear. Methods: The SMYD2 expression profile was examined by quantitative real-time polymerase chain reaction (qRT-PCR), and immunohistochemistry (IHC) in HCC tissues and matched adjacent non-tumorous tissues. SMYD2 was silenced in HCC cell lines to determine its role in tumor proliferation and cell cycle progression, and the possible mechanism. Spearman's rank correlation, Kaplan-Meier plots and Cox proportional hazards regression model were used to analyze the data. Results: The SMYD2 expression in HCC tissues were significantly up-regulated at both mRNA and protein levels as compared with the matched adjacent non-tumorous tissues. By IHC, positive expression of SMYD2 was examined in 122/163 (74.85%) of HCC and in 10/59 (16.95%) of tumor-adjacent tissues. Positive expression of SMYD2 was correlated with tumor size, vascular invasion, differentiation and TNM stage (P < 0.05). In univariate survival analysis, a significant association between positive expression of SMYD2 and shortened patients' survival was found (P < 0.05). Importantly, SMYD2 expression together with vascular invasion (P < 0.05) provided significant independent prognostic parameters in multivariate analysis. Functionally, SMYD2 silenced markedly inhibited cell proliferation and cell cycle progression in SMMC-7721 cell. Conclusions: Our findings provide evidences that positive expression of SMYD2 in HCC may be important in the acquisition of an aggressive phenotype, and it is an independent biomarker for poor prognosis of patients with HCC.

Recent evidence shows that resveratrol (RSV) may ameliorate high-glucose-induced cardiac oxidative stress, mitochondrial dysfunction and myocardial fibrosis in diabetes. However, the mechanisms by which RSV regulates mitochondrial function in diabetic cardiomyopathy have not been fully elucidated. Mitochondrial dysfunction contributes to cardiac dysfunction in diabetic patients, which is associated with dysregulation of peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α). In this study we examined whether resveratrol alleviated cardiac dysfunction in diabetes by improving mitochondrial function via SIRT1-mediated PGC-1α deacetylation. T2DM was induced in rats by a high-fat diet combined with STZ injection. Diabetic rats were orally administered RSV (50 mg·kg–1·d–1) for 16 weeks. RSV administration significantly attenuated diabetes-induced cardiac dysfunction and hypertrophy evidenced by increasing ejection fraction (EF%), fraction shortening (FS%), ratio of early diastolic peak velocity (E velocity) and late diastolic peak velocity (A velocity) of the LV inflow (E/A ratio) and reducing expression levels of pro-hypertrophic markers ANP, BNP and β-MHC. Furthermore, manganese superoxide dismutase (SOD) activity, ATP content, mitochondrial DNA copy number, mitochondrial membrane potential and the expression of nuclear respiration factor (NRF) were all significantly increased in diabetic hearts by RSV administration, whereas the levels of malondialdehvde (MDA) and uncoupling protein 2 (UCP2) were significantly decreased. Moreover, RSV administration significantly activated SIRT1 expression and increased PGC-1α deacetylation. H9c2 cells cultured in a high glucose (HG, 30 mmol/L) condition were used for further analyzing the role of SIRT1/PGC-1α pathway in RSV regulation of mitochondrial function. RSV (20 μmol/L) caused similar beneficial effects in HG-treated H9c2 cells in vitro as in diabetic rats, but these protective effects were abolished by addition of a SIRT1 inhibitor sirtinol (25 μmol/L) or by SIRT1 siRNA transfection. In H9c2 cells, RSV-induced PGC-1α deacetylation was dependent on SIRT1, which was also abolished by a SIRT1 inhibitor and SIRT1 siRNA transfection. Our results demonstrate that resveratrol attenuates cardiac injury in diabetic rats through regulation of mitochondrial function, which is mediated partly through SIRT1 activation and increased PGC-1α deacetylation.

Background Oxymatrine (OM), a quinolizidine alkaloid extracted from a herb Sophorae Flavescentis Radix, has been used to treat liver fibrotic diseases. However, the mechanism of its anti-fibrosis effects is still unclear. TGF-β/Smad signaling and miR-195 have been proved to paly an important role in hepatic stellate cells (HSCs) activation and liver fibrosis. In this study, we investigated whether OM could inhibit HSCs activation through TGF-β1/miR-195/Smads signaling or not. Methods First, the effects of OM on HSC-T6 in different concentrations and time points were tested by MTT assay. We choose three appropriate concentrations of OM as treatment concentrations in following experiment. By Quantitative Real-time PCR and Western Blot, then we investigated the effect of OM on miR-195, Smad7 and α-SMA’s expressions to prove the correlation between OM and the TGF-β1/miR-195/Smads signaling. Last, miR-195 mimic and INF-γ were used to investigate the relation between miR-195 and OM in HSC activation. Results Our results showed that the proliferation of HSC was significantly inhibited when OM concentration was higher than 200 μg/mL after 24 h, 100 μg/mL after 48 h and 10 μg/mL after 72 h. The IC50 of OM after 24, 48 and 72 h were 539, 454, 387 μg/mL respectively. OM could down-regulate miR-195 and α-SMA (P

Aim: Iloperidone is an atypical antipsychotic drug that is mainly metabolized by CYP2D6, CYP3A4, and cytosolic enzymes. Previous studies show that extensive and poor metabolizers of CYP2D6 exhibit different plasma concentrations of iloperidone and its metabolites. The aim of this study was to develop a parent-metabolite population pharmacokinetic (PPK) model to quantify the effects of CYP2D6*10 allele on the pharmacokinetics of iloperidone and its metabolites in Chinese schizophrenia patients. Methods: Seventy Chinese schizophrenia patients were enrolled, from whom limited blood samples were collected on d 15 (0 h) and d 28 (0, 4 and 12 h after drug administration). The plasma concentrations of iloperidone and its metabolites M1 (P-88) and M2 (P-95) were simultaneously detected using a validated HPLC-MS assay. CYP2D6*10 (rs1065852) genotyping was performed. A PPK model was developed based on data from the patients using the NONMEM software (version 7.2). A one-compartment model with first-order absorption and elimination was used to describe the pharmacokinetic data related to iloperidone and its metabolites. Results: Patients with the CYP2D6*10 T/T genotype had significantly higher concentrations of iloperidone and M1, and lower concentrations of M2 than the patients with C/C or C/T genotypes. The CYP2D6*10 genotype affected the elimination constants for transformation of iloperidone to the metabolites M1 (K23) and M2 (K24). The K23 value of the patients with T/T genotype was 1.34-fold as great as that of the patients with C/C or C/T genotype. The K24 value of the patients with C/T and T/T genotypes was 0.693- and 0.492-fold, respectively, as low as that of the patients with C/C genotype. Conclusion: CYP2D6*10 mutations affect the pharmacokinetics of iloperidone and its metabolites in Chinese schizophrenia patients, suggesting that the clinical doses of iloperidone for patients with CYP2D6*10 mutations need to be optimized.

MicroRNAs (miRNAs) are a family of non-coding RNA that are able to adjust the expression of many proteins, including ATP-binding cassette transporter and organic cation transporter. We sought to evaluate the effect of miR-511 on the regulation of OATP1B1 expression by free fatty acids. When using free fatty acids to stimulate Chang liver cells, we found that the expression of miR-511 increased significantly while the expression of OATP1B1 decreased. We also proved that SLCO1B1 is the target gene of miR-511 with a bioinformatics analysis and using the dual luciferase reporter assay. Furthermore, the expressions of SLCO1B1 and OATP1B1 decreased if transfecting Chang liver cells with miR-511, but did not increase when transfecting the inhibitors of miR-511 into steatosis cells. Our study indicates that miR-511 may play an important role in the regulation of OATP1B1 expression by free fatty acids.

New Onset Diabetes after Transplantation (NODAT) is defined as sustained hyperglycemia developing in patients without diabetes history before transplantation. A cohort study was performed to access the effects of tacrolimus on insulin secretion and insulin sensitivity and consequently in the development of NODAT in kidney transplant recipients. Then, we further investigated the association between NODAT and single-nucleotide polymorphisms of IRS-1 and IRS-2 in renal allograft recipients. One hundred and fifty-eight kidney transplant patients, receiving tacrolimus as the base immunosuppressant, were divided into two groups: with or without NODAT. Plasma levels of fasting insulin concentration (FINS) and C-peptide were determined by enhanced chemiluminescence immunoassay and ADVIA Centaur C peptide assay, respectively. The genotypes of Gly1057Asp in IRS-2 and Gly972Arg in IRS-1 were detected through polymerase chain reaction fragment length polymorphism in NODAT and non-NODAT patients. It was found that the concentrations of fasting plasma insulin and C-peptide in NODAT and non-NODAT patients treated with tacrolimus were higher than that in healthy volunteers (p 0.05). Fasting plasma insulin concentration in NODAT was significantly elevated compared with than that in non-NODAT group (p 0.05). But there are no statistical differences in fasting plasma C-peptide concentrations between NODAT and non-NODAT groups. The allele and genotype frequencies of IRS-2 Gly1057Asp and IRS-1 Gly972Arg in NODAT patients were not significantly different from non-NODAT patients (p 0.05). In conclusion, insulin resistance is the primary cause of tacrolimus-induced NODAT. The IRS-2 Gly1057Asp and IRS-1 Gly972Arg genotypes are not related to NODAT.

Background and Aim. Aberrant activation of the TGF-β1/Smad pathway contributes to the activation of hepatic stellate cells (HSCs). MicroRNA-195 has been shown to regulate the activation of HSCs. The aim of this study was to investigate the role of miRNA-195 in HSCs activation. Methods. A liver fibrotic rat model induced by diethylnitrosamine was established. Dual luciferase reporter assays were performed to verify that Smad7 was the target of miRNA-195. The expression levels of miR-195, Smad7, and α-SMA in HSC-T6 transfected, respectively, with miR-195 mimic, inhibitor, or control were measured by qRT-PCR. The protein expression of Smad7 was detected by Western blot analysis. Results. Enhanced miR-195 and decreased Smad7 were observed in diethylnitrosamine-induced liver fibrotic rats (P<0.05). Dual luciferase reporter assays showed that the miR-195 mimic significantly suppressed the luciferase activity of a reporter plasmid carrying the binding site of miR-195 on the 3′UTR of Smad7 (P<0.05). The miR-195 mimics activated HSCs, further elevated miR-195 and α-SMA (P<0.01), and reduced the Smad7 level (P<0.05). The miR-195 inhibitors blocked the activation of HSCs, reduced the expression of miR-195 and α-SMA (P<0.01), and upregulated the expression of Smad7 (P<0.05). Conclusion. Collectively, we demonstrated that miRNA-195 activated HSCs by targeting Smad7.

The objective of this study was to evaluate the effect of the CYP3A5*3 allele on the pharmacokinetics of tacrolimus and amlodipine, and drug-drug interactions between them in healthy subjects. Pharmacokinetic drug interactions between tacrolimus and amlodipine were evaluated in a randomized, 3-period, 6-sequence crossover study in healthy Chinese volunteers according to CYP3A5 genotype. A single-dose and multiple-dose study were designed. A 96-h pharmacokinetic study followed either tacrolimus or amlodipine dose, and the washout periods between the study phases were 14 days. In the single-dose study, apparent oral clearance (CL/F) of tacrolimus (5 mg) in CYP3A5 expressers was 3.8-fold (p = 0.008) higher than that in CYP3A5 non-expressers. Amlodipine decreased mean tacrolimus CL/F in CYP3A5 expressers by 2.2-fold (p = 0.005), while it had no effect on that in CYP3A5 non-expressers. The CL/F of amlodipine in CYP3A5 non-expressers was 2.0-fold (p = 0.001) higher than that in CYP3A5 expressers. Tacrolimus increased mean amlodipine CL/F in CYP3A5 expressers by 1.4-fold (p = 0.016) while it had no effect on that in CYP3A5 non-expressers. Tacrolimus slightly reduced the AUC₀-∞ of amlodipine in both CYP3A5 expressers and non-expressers. Dose adjustment of tacrolimus should be considered according to CYP3A5*3 genetic polymorphism when tacrolimus is coadministered with amlodipine.

Background There is no single standard chemotherapy regimen for elderly patients with advanced gastric cancer (AGC). A phase III trial has confirmed that both capecitabine monotherapy and capecitabine plus oxaliplatin are well tolerated for elderly patients with AGC, but their economic influence in China is unknown. Objective The purpose of this cost-effectiveness analysis was to estimate the effects of capecitabine monotherapy and capecitabine plus oxaliplatin in elderly patients with AGC on health and economic outcomes in China. Methods We created a Markov model based on data from a Korean clinical phase III trial to analyze the cost-effectiveness of the treatment of elderly patients in the capecitabine monotherapy (X) group and capecitabine plus oxaliplatin (XELOX) group. The costs were obtained from published reports and the local health system. The utilities were assumed on the basis of the published literature. Costs, quality-adjusted life years (QALYs), and incremental cost-effectiveness ratios (ICER) were estimated. One-way and probabilistic sensitivity analyses (Monte Carlo simulations) were performed. Results In the cost-effectiveness analysis, X had a lower total cost ($45,731.68) and cost-effectiveness ratio ($65,918.93/QALY). The one-way sensitivity analysis suggested that the most influential parameter was the risk of requiring second-line chemotherapy in XELOX group. The probabilistic sensitivity analysis predicted that the X regimen was cost-effective 100% of the time, given a willingness-to-pay threshold of $26,598. Conclusions Our findings show that the XELOX regimen is less cost-effective compared to the X regimen for elderly patients with AGC in China from a Chinese healthcare perspective.

OBJECTIVES: We aim to obtain the intra-subject coefficient of variability of a highly variable antidepressant agomelatine in humans, and propose an adjusted bioequivalence assessment strategy. METHODS: A single-dose, randomized crossover design was conducted in four periods (reference administered thrice, placebo administered once) separated by seven days. A validated LC-MS/MS assay was used to measure drug concentrations in serial blood samples. RESULTS: The intra-subject coefficient of variability was calculated using the residual variance of ANOVA analysis, and the results for Cmax and AUC0-t was 78.34% and 43.52%, respectively, in Chinese healthy subjects. The sample size required for standard BE study were 124(192, 340) if the expected deviation between the reference and generic products was set to 0 (5%, 10%). CONCLUSIONS: Agomelatine meets the criteria for highly variable drug in Chinese healthy male subjects, and the traditional BE criteria for agomelatine needs to be adjusted to alleviate the resource and ethical burden of using a large numbers of subjects in clinical trials. Our clinical data on the intra-subject variability of agomelatine PK in Chinese healthy population enables to adjust bioequivalence (BE) assessment approach for agomelatine based on the RSABE approaches recommended by regulatory agencies. TRIAL REGISTRATION: ChiCTR.org ChiCTR-TTRCC-13003835.

Chlorogenic acid (ChA) is proposed as the major bioactive compounds of Lonicerae Japonicae Flos (LJF). Forty-two Wistar rats were randomly divided into seven groups to investigate the pharmacokinetics and tissue distribution of ChA, via oral administration of LJF extract, using ibuprofen as internal standard, employing a high performance liquid chromatography in conjunction with tandem mass spectrometry. Analytes were extracted from plasma samples and tissue homogenate by liquid–liquid extraction with acetonitrile, separated on a C18 column by linear gradient elution, and detected by electrospray ionization mass spectrometry in negative selected multiple reaction monitoring mode. Our results successfully demonstrate that the method has satisfactory selectivity, linearity, extraction recovery, matrix effect, precision, accuracy, and stability. Using noncompartment model to study pharmacokinetics, profile revealed that ChA was rapidly absorbed and eliminated. Tissue study indicated that the highest level was observed in liver, followed by kidney, lung, heart, and spleen. In conclusion, this method was suitable for the study on pharmacokinetics and tissue distribution of ChA after oral administration.

Objectives:The aims of this study were to investigate the pharmacokinetic (PK) properties of aripiprazole in the steady state in Han Chinese adults with schizophrenia and to compare them between Han Chinese and white populations described in the literature.Methods:This prospective, open4abel, pilot study was conducted at the Mental Health Institute, Xiang-ya Second Hospital, Central South University, Changsha, China. Male and female hospitalized patients aged 18 to 45 years diagnosed with Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV)-defined schizophrenia, with a Positive and Negative Syndrome Scale (PANSS) total score ⩾60 (indicating schizophrenia of at least mild severity) were eligible. On study days 1 and 2, patients were pretreated with aripiprazole 10 mg PO QD, followed by 10 mg g12h on days 3 to 21. Blood samples were drawn for analysis on day 21 before dosing and 1, 3, 4, 5, 12, 24, 48, 72, 96, 144, and 192 hours after the morning dosing of aripiprazole on day 21. Patients received low-dose (25–100 mg/d) clozapine on day 25 until day 28. The samples were assessed using high-performance liquid chromatography-mass spectrometry and compartment model analysis for aripiprazole. PK properties included mean residence time (MRT) steady-state Cmax (Css max), time to Css max (Tmax), elimination t/12, apparent oral clearance (CL/F), and apparent volume of distribution (V/F). Adverse effects were monitored using physical examination (including vital sign measurements), electrocardiography, electroencephalography, and clinical laboratory testing (including biochemistry, hematology, and urinalysis) at baseline and at the end of the study. Patients were asked about adverse events on days 1 to 7 and at random intervals thereafter. Patients were also instructed to report any spontaneous symptoms they experienced.Results:Twelve patients were enrolled (6 men, 6 women; mean [SD] age, 26.1 [7.0] years; mean [SD] weight, 56.6 [9.0] kg; mean [SD] PANSS score, 116.8 [12.2]). Aripiprazole exhibited linear kinetic characteristics on a 2-compartment model. After multiple oral doses (10 mg g12h), the mean (SD) t1/2, Css max, Tmax, MRT, V/F, and CL/F were 62.2 (9.0) hours, 557.3 (135.5) ng/mL, 2.6 (1.1) hours, 84.5 (11.2) hours, 173 (48) L, and 1.9 (0.5) L/h, respectively. In Chinese patients, the t/12 values were numerically similar (62.2 [9.0] vs 68.1 [22.9] hours); Css max values were numerically higher (557.3 [135.5] vs 393 [181 ] ng/mL); and V/F and CL/F values were numerically lower (V/F: 173 [48] vs 196 [66] L; CL/F: 1.9 [0.5] vs 3.4 [1.6] L/h) compared with healthy white male volunteers. Adverse effects were mild to moderate: lightheadedness (5 of 12 patients), somnolence (3), tachycardia (3), hypodynamia (2), and extrapyramidal symptoms (EPS) (1). The EPS (convulsive movement of the muscles related to the larynx) led to one patient's discontinuation of the study.Conclusions:In this small pilot study of the PK properties of aripiprazole 10 mg PO g12h in Han Chinese patients with schizophrenia, the mean t1/2 value was numerically similar to that previously reported in a population of healthy white male volunteers. However, the mean Css max value was numerically higher, and V/F and CL/F values were numerically lower, compared with those in healthy white male volunteers.