Knockdown of HE4 suppresses tumor growth and invasiveness in lung adenocarcinoma through regulation of EGFR signaling

It has been shown that the high expression of human epididymis protein 4(HE4)in most lung cancers is related to the poor prognosis of patients,but the mechanism of pathological transformation of HE4 in lung cancer is still unclear.The current study is expected to clarify the function and mechanism of HE4 in the occurrence and metastasis of lung adenocarcinoma(LUAD).Immunoblotting evaluated HE4 expression in lung cancer cell lines and biopsies,and through analysis of The Cancer Genome Atlas(TCGA)dataset.Frequent HE4 overexpression was demonstrated in LUAD,but not in lung squamous cell carcinoma(LUSC),indicating that HE4 can serve as a biomarker to distinguish between LUAD and LUSC.HE4 knockdown significantly inhibited cell growth,colony formation,wound healing,and invasion,and blocked the G1-phase of the cell cycle in LUAD cell lines through inactivation of the EGFR signaling downstream including PI3K/AKT/mTOR and RAF/MAPK pathways.The first-line EGFR inhibitor gefitinib and HE4 shRNA had no synergistic inhibitory effect on the growth of lung adenocarcinoma cells,while the third-line EGFR inhibitor osimertinib showed additive anti-proliferative effects.Moreover,we provided evidence that HE4 regulated EGFR expression by transcription regulation and protein interaction in LUAD.Our findings suggest that HE4 positively modulates the EGFR signaling pathway to promote growth and invasiveness in LUAD and highlight that targeting HE4 could be a novel strategy for LUAD treatment.

Impact of release characteristics of sinomenine hydrochloride dosage forms on its pharmacokinetics in beagle dogs

AIM: To investigate the effect of release behavior of sustained-release dosage forms of sinomenine hydrochloride dosage forms, including commercial 12-h sustained-release tablets and 24-h sustained-release pellets prepared in our laboratory, was examined. The two dosage forms were orally administrated to beagle dogs, and then the in vivo formulation was achieved by mixing slow- and rapid-of the sustained-release pellets were scarcely influenced by the pH of the dissolution medium. Release from the 12-h sustained-release tablets was markedly quicker than that from the 24-h sustained-release pellets, the cumulative release up to 12-h was 99.9% vs68.7%. From release pellets had longer tmax and lower Cmax compared to the 12-h sustained-release tablets, the tmax being 2.67±0.52 h vs9.83±0.98 h and the Cmax being 1 334.45±368.76 ng/mL vs 893.12±292.55 ng/mL, respectively. However, the AUC0-tn preparations were statistically bioequivalent. Furthermore,percentage absorption in vivo and the cumulative percentage release in vitro.CONCLUSION: The in vitro release properties of the dosage forms strongly affect their pharmacokinetic behavior in vivo. Therefore, managing the in vitro release behavior of dosage forms is a promising strategy for obtaining the optimal in vivo pharmacokinetic characteristics and safe therapeutic drug concentration-time curves.

Permeabilities of rebamipide via rat intestinal membranes and its colon specific delivery using chitosan capsule as a carrier

AIM: To investigate the permeability characteristics of rebamipide across intestinal mucosa, and examine the effects of some absorption enhancers on the permeability across the colonic tissue. Another purpose is to demonstrate the colon-specific delivery of rebamipide with or without absorption enhancers using chitosan capsule as a carrier. METHODS: The permeability of rebamipide was evaluated using an in vitro diffusion chamber system, and the effects of some absorption enhancers on the permeability via colon were further investigated. The release of rebamipide from chitosan or gelatin capsule was studied by Japan Pharmacopoeia rotating basket method. The colonic and plasma concentrations were analyzed by high performance liquid chromatography (HPLC) to evaluate colon-targeting action after oral administration of various dosage forms, and rebamipide with absorption enhancers in chitosan dosage forms. RESULTS: The permeability of rebamipide across the jejunal or ileal membranes was higher than the colonic membranes. Both sodium laurate (C12) and labrasol signifi cantly increased permeability across the colon membranes. On the other hand, the release of rebamipide from chitosan capsule was less than 10%totally within 6 h. The area under concentration-time profile of drug in the colon mucosa using chitosan capsules (AUCLI, 1 6011.2 ng·h/g) was 2.5 times and 4.4 times greater than using gelatin capsules and CMC suspension, respectively. Meanwhile, the area under concentration-time profile of drug in the plasma (AUCPL) was 1016.0 ng·h/mL for chitosan capsule, 1887.9 ng·h/mL for CMC suspension p and 2163.5 ng·h/mL for gelatin capsule. Overall, both AUCLI and AUCPL were increased when C12 was co-administrated, but the increase of AUCLI was much greater; the drug delivery index (DDI) was more than 1 compared with simple chitosan capsule group. CONCLUSION: There was a regional difference in the permeability of Rabamipide across the jejunum, ileum and the colon, and passive diffusion seems to be one of the major transport mechanisms of rebamipide. Absorption enhancers can increase the permeability of rebamipide across the colon tissue signifi cantly. In addition, chitosan capsule may be a useful carrier to deliver rebamipide to the colon specifi cally and the co-administration of C12 with rebamipide may also be very useful in local treatment.

A rapid and sensitive determination of paclitaxel in rat plasma by UPLC-MS/MS method: Application to a pharmacokinetic study

A rapid and sensitive method for quantitative determination of paclitaxel in rat plasmawas developed and validated by using ultra-performance liquid chromatography-tandemmass spectrometry (UPLC-MS/MS). Docetaxel was used as an internal standard anddiethyl ether was the liquideliquid extraction agent. Multiple reaction monitoring (MRM)mode via positive electrospray ionization (ESI) was applied to detect paclitaxel and IS at thetransitions m/z 854 / 286 and m/z 808.48 / 527.3, respectively. This method covered alinearity range from 5 to 5000 ng/ml, with the total run time of 3.0 min. In summary, a highthroughout UPLC-MS/MS method was successfully developed to measure paclitaxel in ratplasma and was applied to pharmacokinetic study after intravenous administration ofpaclitaxel.

Sensitive detection of alkaline phosphatase based on terminal deoxynucleotidyl transferase and endonuclease Ⅳ-assisted exponential signal amplification

Alkaline phosphatase(ALP)is widely expressed in human tissues.ALP plays an important role in the dephosphorylation of proteins and nucleic acids.Therefore,quantitative analysis of ALP plays a vital role in disease diagnosis and the development of biological detection methods.Terminal deoxynucleotidyl transferase(TdT)catalyzes continuous polymerization of deoxynucleotide triphosphates at the 30-OH end of single-stranded DNA in the absence of a template.In this study,we developed a highly sensitive and selective method based on TdT and endonuclease Ⅳ(Endo Ⅳ)to quantify ALP activity.After ALP hydrolyzes the 30-PO_(4) end of the substrate and generates 30-OH,TdT can effectively elongate the 30-OH end with deoxynucleotide adenine triphosphate(dATP)and produce a poly A tail,which can be detected by the poly T probes.Endo Ⅳ digests the AP site in poly T probes to generate a fluorescent signal and a new 30-OH end,leading to the generation of exponential fluorescence signal amplification.The substrate for TdT elongation was optimized,and a limit of detection of 4.3×10^(-3) U/L was achieved for ALP by the optimized substrate structure.This method can also detect ALP in the cell lysate of a single cell.This work has potential applications in disease diagnosis and biomedical detection.

Genomic Diversity and Evolution of Bacillus subtilis

Bacillus subtilis is the focus of both academic and industrial research.Previous studies have reported a number of sequence variations in different B.subtilis strains.To uncover the genetic variation and evolutionary pressure in B.subtilis strains,we performed whole genome sequencing of two B.subtilis isolates,KM and CGMCC63528.Comparative genomic analyses of these two strains with other B.subtilis strains identified high

Biological activities of the hydro-alcoholic and aqueous extracts of <i>Achillea fragrantissima</i>(Forssk.) grown in Jordan

Aqueous and hydro-alcoholic extracts of Achillea fragrantissima L. (Asteraceae) grown in Jordan were screened for their antioxidant, antimicrobial, antiplatelet, anti-proliferative and acetylcholinesterase (AChE) inhibition efficacy. Total phenols and flavonoids were determined colorimetrically. The radical scavenging activities were evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2’-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) radical scavenging activity assays. High performance liquid chromatography-mass spectrometry (HPLCMS) analysis resulted in the identification of 7 phenolic compounds in the hydro-alcoholic extract and 4 compounds in the aqueous extract;quercetin 3-β-D-glucoside was the main component for both extracts. Antimicrobial activities were determined by antimicrobial susceptibility testing such as agar well-diffusion method, minimum inhibition concentration and minimum bactericidal concentration. Gram positive bacteria showed sensitivity to hydro-alcoholic extract in the agar-well diffusion test. No significant activity was observed against gram negative bacteria and Candida albicans. Hydro-alcoholic extract had a bactericidal activity against Streptococcus pneumoniae and Bacillus cereus at high concentrations (MIC 12.5 mg/ml) rather than inhibitory effect. In vitro antiplatelet activity was tested on human whole blood using an electrical impedance method. At concentrations (50, 100, and 200 μg/ml), no effect on platelet aggregation was noticed. Anti-proliferative activity was investigated using the MTT assay. At concentrations up to 200 μg/ml, extracts did not possess cytotoxic activity against the MCF-7 cells. Acetylcholinesterase (AChE) inhibitory capacity of A. fragrantissima extracts was tested using TLC assay method, and neither aqueous, nor hydroalcoholic extracts showed AChE inhibition. The present investigation supported the traditional use of A. fragrantissima in the Jordanian folk medicine as an antimicrobial active representative of the genus Achillea. A. fragrantissima extracts should be further studied for their potential use in preventing/treating diseases in which oxidative stress is a part of the pathophysiology.

Induction of Cytochrome P450 2A6 by Bilirubin in Human Hepatocytes

The influence of bilirubin on mRNA expression of cytochrome P450 (CYP), UDP-glucuronosyltransferase (UGT) and nuclear receptors in human hepatocytes was investigated. The treatment of the hepatocytes with 40 μg/mL bilirubin, which corresponds to hyperbilirubinemia, resulted in 1.7-fold increase of CYP2A6 mRNA compared to the vehicle control while CYP2A6 mRNA did not change after treatment with 1 μg/mL bilirubin, corresponding to physiologically normal level. No significant change of mRNA expression by 40 μg/mL bilirubin treatment was observed for CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP3A4 and CYP3A5, UGT1A1, UGT1A3, UGT1A6, UGT2B4, UGT2B7, UGT2B10 and UGT2B15, constitutive androstane receptor (CAR), pregnane X receptor (PXR), retinoid X receptor α (RXRα) and hepatocyte nuclear factor-4α (HNF-4α). The induction profile of bilirubin was different from that of rifampicin, a typical PXR activator. This study demonstrated that CYP2A6 can be induced by bilirubin in a concentration dependent manner.

Patient Perspectives of the Role of the Community Pharmacist in the Middle East: Jordan, United Arab Emirates and Iraq

Objectives: To explore patient’s perspectives of the role of the community pharmacist in view of other health care professionals (specialist physician, general practitioner, nurse and others (e.g. herbalist)) in Jordan, UAE and Iraq. The study also investigated patient’s willingness to pay for specialized pharmaceutical care services. Methods: This study followed a single phase cross sectional survey methodology, conducted in three Middle Eastern countries. Data were collected over a period of 6 months on two consecutive years (March to May 2009 and 2010). A questionnaire was designed and validated, then completed by patients walking into community pharmacies. The source of advice (specialist physician, general practitioner, pharmacist, nurse, or herbalist) regarding patient’s medication use and medical management were investigated. Key findings: Patients visiting community pharmacies in Jordan (n = 1000), UAE (n = 1000) and Iraq (n = 968) were recruited into the study (mean age 35.9 ± 13.1, 50.6% males). Significant difference between the three countries was shown, as more patients chose the pharmacist as their primary source of advice on medication use vs. the specialist physician in Jordan (50.8% vs. 37.3%) and Iraq (41.9% vs. 36.7%) compared to UAE (38.0% vs. 40.1%), P < 0.001, Chi-square test. Few agreed to receive paid pharmaceutical care services (Jordan 19.5%;UAE 24.7%;Iraq 2.3%). Conclusion: Current situation resulting in the lower socioeconomic status in Jordan and Iraq seems to lead to patients’ higher reliance on the pharmacist, as compared to the UAE. Majority of patients, regardless of the country of origin, are not willing to pay for specialized pharmaceutical care services. These findings are important for future social pharmacy research in the area.

pH-sensitive polymeric micelles triggered drug release for extracellular and intracellular drug targeting delivery

Most of the conventional chemotherapeutic agents used for cancer chemotherapy suffer from multidrug resistance of tumor cells and poor antitumor efficacy.Based on physiological differences between the normal tissue and the tumor tissue,one effective approach to improve the efficacy of cancer chemotherapy is to develop pH-sensitive polymeric micellar delivery systems.The copolymers with reversible protonationedeprotonation core units or acid-liable bonds between the therapeutic agents and the micelle-forming copolymers can be used to form pH-sensitive polymeric micelles for extracellular and intracellular drug smart release.These systems can be triggered to release drug in response to the slightly acidic extracellular fluids of tumor tissue after accumulation in tumor tissues via the enhanced permeability and retention effect,or they can be triggered to release drug in endosomes or lysosomes by pH-controlled micelle hydrolysis or dissociation after uptake by cells via the endocytic pathway.The pH-sensitive micelles have been proved the specific tumor cell targeting,enhanced cellular internalization,rapid drug release,and multidrug resistance reversal.The multifunctional polymeric micelles combining extracellular pH-sensitivity with receptor-mediated active targeting strategies are of great interest for enhanced tumor targeting.The micelles with receptor-mediated and intracellular pH targeting functions are internalized via receptor-mediated endocytosis followed by endosomal-pH triggered drug release inside the cells,which reverses multidrug resistance.The pH sensitivity strategy of the polymeric micelles facilitates the specific drug delivery with reduced systemic side effects and improved chemotherapeutical efficacy,and is a novel promising platform for tumor-targeting drug delivery.

Physiological Correlation of Airway Pressure and Transpulmonary Pressure Stress Index on Respiratory Mechanics in Acute Respiratory Failure

Background： Stress index at post-recruitment maneuvers could be a method of positive end-expiratory pressure （PEEP） titration in acute respiratory distress syndrome （ARDS） patients. However, airway pressure （P_aw） stress index may not reflect lung mechanics in the patients with high chest wall elastance. This study was to evaluate the P stress index on lung mechanics and the correlation between P stress index and transpulmonary pressure （P_L） stress index in acute respiratory failure （ARF） patients. Methods： Twenty-four ARF patients with mechanical ventilation （MV） were consecutively recruited from July 2011 to April 2013 in Zhongda Hospital, Nanjing, China and Ospedale S. Giovanni Battista-Molinette Hospital, Turin, Italy. All patients underwent MV with volume control （tidal volume 6 ml/kg） for 20 rain. PEEP was set according to the ARDSnet study protocol. The patients were divided into two groups according to the chest wall elastance/respiratory system elastance ratio. The high elastance group （H group, n = 14） had a ratio _〉30%, and the low elastance group （L group, n = 10） had a ratio 〈30%. Respiratory elastance, gas-exchange, P stress index, and Pt stress index were measured. Student＇s t-test, regression analysis, and Bland-Altman analysis were used for statistical analysis. Results： Pneumonia was the major cause of respiratory failure （71.0%）. Compared with the L group, PEEP was lower in the H group （5.7 ± 1.7 cmH,O vs. 9.0 ± 2.3 cmH2O, P 〈 0.01 ）. Compared with the H group, lung elastance was higher （20.0 ± 7.8 cmH20/L vs. 11.6 ± 3.6 cmH,O/L, P 〈 0.01）, and stress was higher in the L group （7.0 ±1.9 vs. 4.9 ± 1.9, P = 0.02）. A linear relationship was observed between the P stress index and the P stress index in H group （x2 = 0.56, P 〈 0.01 ） and L group （x2 = 0.85, P 〈 0.01）. Conclusion： In the ARF patients with MV, P,w stress index can substitute for PL to guide ventilator settings.

Glycine origin of the methyl substituent on C7′-N of octodiose for the biosynthesis of apramycin

Apramycin is unique in the aminoglycoside family due to its octodiose moiety. However, either the biosynthesis process or the precursors involved are largely unknown. Addition of glycine, as well as serine or threonine, to the Streptomyces tenebrabrius UD2 fermentation medium substantially increases the production of apramycin with little effect on the growth of mycelia, indicating that glycine and/or serine might be involved in the biosynthesis of apramycin. The 13C-NMR analysis of [2-13C] glycine-fed (25% enrichment) apramycin showed that glycine specifically and efficiently incorporated into the only N-CH3 substituent of apramycin on the C7′ of the octodiose moiety. We noticed that the in vivo concentration of S-adenosyl methionine increased in parallel with the addition of glycine, while the addition of methione in the fermentation medium significantly decreased the productivity of apra-mycin. Therefore, the methyl donor function of glycine is proposed to be involved in the methionine cycle but methionine itself was proposed to inhibit the methylation and methyl transfer processes as previously reported for the case of rapamycin. The 15N NMR spectra of [2-13C,15N]serine labeled apramycin indicated that serine may also act as a limiting precursor contributing to the ―NH2 substituents of apramycin.

Spontaneous modulation of a dynamic balance between bacterial genomic stability and mutability: roles and molecular mechanisms of the genetic switch

Bacteria need a high degree of genetic stability to maintain their species identities over long evolutionary times while retaining some mutability to adapt to the changing environment.It is a long unanswered question that how bacteria reconcile these seemingly contradictory biological properties.We hypothesized that certain mechanisms must maintain a dynamic balance between genetic stability and mutability for the survival and evolution of bacterial species.To identify such mechanisms,we analyzed bacterial genomes,focusing on the Salmonella mismatch repair(MMR)system.We found that the MMR gene mutL functions as a genetic switch through a slipped-strand mispairing mechanism,modulating and maintaining a dynamic balance between genetic stability and mutability during bacterial evolution.This mechanism allows bacteria to maintain their phylogenetic status,while also adapting to changing environments by acquiring novel traits.In this review,we outline the history of research into this genetic switch,from its discovery to the latest findings,and discuss its potential roles in the genomic evolution of bacteria.

Atomic force microscopy analysis of orientation and bending of oligodeoxynucleotides in polypod-like structured DNA

我们以前证明象多足一样组织了 DNA，或 polypodna，与三或更构造了 oligodeoxynucleotides (ODN ) ，被象树枝状的房间和巨噬细胞那样的有免疫力的房间高效地收起，取决于它的结构的复杂性。包括 polypodna 的 ODN 应该弯曲形成象多足一样结构，并且可以那么由采用 bendtype 符合构造或跨类型的符合构造做。这里，我们试着阐明取向并且用原子力量显微镜学(AFM ) 在 polypodnas ODN 弯曲。我们设计了 pentapodnas 的二种类型(即，有五个豆荚的 polypodna ) 使用 60- 到 88 底 ODN，它然后在 DNA origami 上被使不能调动装裱。AFM 成像证明在 pentapodna 的 ODN 采用了拐弯类型符合构造。当他们在非强迫的条件下面在框架上被使不能调动时， Tetrapodna 和 hexapodna 也采用了拐弯类型符合构造。这些调查结果提供向协调设计的有用信息，和 structureactivity 关系为，许多 DNA nanostructures。

Pullulan Acetate Coated Magnetite Nanoparticles for Hyper-Thermia:Preparation,Characterization and In Vitro Experiments

Amphipathic polymer pullulan acetate (PA)-coated magnetic nanoparticles were prepared and characterized by various physicochemical means. The cytotoxicity and cellular uptake of the magnetic nanoparticles were examined. The hyperthermic effect of the magnetic nanoparticles on tumor cells was evaluated. Transmission electron microscopy (TEM) showed that the PA coated magnetic nanoparticles (PAMNs) had spherical morphology. Dynamic light scattering (DLS) showed that the size distribution of PAMNs was unimodal,with an average diameter of 25.8 nm ± 6.1 nm. The presence of the adsorbed layer of PA on the magnetite surface was confirmed by Fourier transform infrared (FTIR) spectroscopy. Magnetic measurements revealed that the saturation magnetization of the PAMNs reached 51.9 emu/g and the nanoparticles were superparamagnetic. Thermogravimetric analysis (TGA) showed that the Fe_(3)O_(4) particles constituted 75 wt% of the PAMNs. The PAMNs had good heating properties in an alternating magnetic field. Cytotoxicity assay showed that PAMNs exhibited no significant cytotoxicity against L929 cells. TEM results showed that a large number of PAMNs were internalized into KB cells. PAMNs have good hyperthermia effect on KB cells in vitro by magnetic field induced hyperthermia. These novel magnetic nanoparticles have great potential as magnetic hyperthermia mediators.