Activated hepatic stellate cells promote progression of post-heat residual hepatocellular carcinoma from autophagic survival to proliferation

Background: Microscopic residual tumor often occurs after thermal ablation for medium-large hepatocellular carcinoma (HCC), leading to early aggressive recurrence or late relapse during follow-up. The mechanism how microscopic residual HCC cells survive sublethal heat stress and develop rapid outgrowth remains poorly understood.

Methods: HCC cells were exposed to sublethal heat treatment and co-cultured with conditioned media from activated HSCs (HSC-CM). Changes of cell proliferation, parameters of cell autophagy and activation of signaling pathways in heat-treated residual HCC cells were analyzed. An HCC orthotopic model was subjected to partial thermal ablation and antitumor effects of a combined treatment regimen were studied.

Results: HCC cells survived sublethal heat stress via activation of autophagy. HSC-CM enhanced autophagic survival within 24?h and then promoted proliferation of heat-treated residual HCC cells through HGF/c-Met signaling. Inhibition of autophagy or c-Met increased apoptosis of heat-treated residual HCC cells and reversed the protective effect of HSC-CM. HGF modulated biological status in autophagic survival or proliferation of heat-treated residual HCC through HGF/c-Met/ERK signaling and downstream components of ATG5/Beclin1 or cyclinD1. In an animal model, inhibiting autophagy in combination with c-Met inhibitor significantly thwarted tumor progression of residual HCC after incomplete thermal ablation via the suppressed autophagy, the decreased proliferation and the increased apoptosis.

Conclusions: Activated HSCs promote progression of residual HCC cells after sublethal heat treatment from autophagic survival to proliferation via HGF/c-Met signaling. A combined treatment regimen of inhibiting autophagy and c-Met signaling could be used to suppress tumor progression of residual HCC after incomplete thermal ablation.     

Investigation of non-linear Mate1-mediated efflux of trimethoprim in the mouse kidney as the mechanism underlying drug-drug interactions between trimethoprim and organic cations in the kidney

The purpose of this study was to elucidate the involvement of Mate1 in the tubular secretion of trimethoprim and saturation of Mate1-mediated efflux to address the mechanisms underlying the pharmacokinetic drug interactions with trimethoprim. Trimethoprim is a more potent inhibitor of MATE2-K than MATE1 with K_i values (μM) of 0.030–0.28 and 2.4–5.9, respectively. Trimethoprim is a substrate of human MATE1 and MATE2-K with K_m values of 2.3 ± 0.9 and 0.018 ± 0.004 μM, and mouse Mate1, but not human OCT2, mouse Oct1 and Oct2. Pyrimethamine significantly reduced the renal clearance (CL_R) of trimethoprim (mL/min/kg) from 40.0 ± 5.1 to 20.1 ± 3.7 (p < 0.05). Trimethoprim was given to mice at three infusion rates (150, 500, and 1500 nmol/min/kg). Together with an increase in the plasma concentrations of trimethoprim, the CL_R (mL/min/kg) of trimethoprim decreased to 25.9 ± 3.2, 13.5 ± 5.7, and 8.92 ± 1.50 at the respective rates. Trimethoprim decreased the CL_R of rhodamine 123 in an infusion rate-dependent manner: 11.5 ± 1.3 (control), 5.17 ± 1.55, 1.31 ± 0.50, and 0.532 ± 0.180. These results suggest that Mate1 mediates the tubular secretion of trimethoprim, and at therapeutic doses, MATEs-mediated efflux can be saturated, and thereby, cause drug interactions with other MATE substrates.     

何首乌“九蒸九晒”炮制工艺优选及对L02肝细胞生长的影响

目的探索何首乌"九蒸九晒"最佳炮制工艺及不同炮制品对正常人L02肝细胞生长的影响。方法选择蒸制时间、炮制次数、晒制时间3个因素,采用L9(34)正交试验,以二苯乙烯苷、游离蒽醌、结合蒽醌含量为考察指标,优选炮制工艺;以细胞抑制率为评价指标,考察何首乌不同炮制品对L02肝细胞生长的影响。结果优选的何首乌"九蒸九晒"最佳炮制工艺为:蒸制4 h,晒制4 h,蒸晒11次。优选炮制品的肝细胞抑制率为2.86%,药典法炮制品的肝细胞抑制率为23.67%。结论本研究优选的何首乌"九蒸九晒"炮制工艺结果可靠,评价指标可控,所得炮制品的肝细胞抑制率低于药典法炮制品。     

Nobiletin and its derivatives overcome multidrug resistance (MDR) in cancer: total synthesis and discovery of potent MDR reversal agents

Our recent studies demonstrated that the natural product nobiletin (NOB) served as a promising multidrug resistance (MDR) reversal agent and improved the effectiveness of cancer chemotherapy in vitro. However, low aqueous solubility and difficulty in total synthesis limited its application as a therapeutic agent. To tackle these challenges, NOB was synthesized in a high yield by a concise route of six steps and fourteen derivatives were synthesized with remarkable solubility and efficacy. All the compounds showed improved sensitivity to paclitaxel (PTX) in P-glycoprotein (P-gp) overexpressing MDR cancer cells. Among them, compound 29d exhibited water solubility 280-fold higher than NOB. A drug-resistance A549/T xenograft model showed that 29d, at a dose of 50 mg/kg co-administered with PTX (15 mg/kg), inhibited tumor growth more effective than NOB and remarkably increased PTX concentration in the tumors via P-gp inhibition. Moreover, Western blot experiments revealed that 29d inhibited expression of NRF2, phosphorylated ERK and AKT in MDR cancer cells, thus implying 29d of multiple mechanisms to reverse MDR in lung cancer.     

Comparison of urinary amino acid excretory pattern in patients with type 2 diabetes mellitus and non-diabetic healthy controls at a tertiary referral hospital in India

AimSpecific amino acids have been linked to regulation of insulin secretion from pancreatic β-cells; on the contrary, increased concentration of certain amino acids is associated with insulin resistance (IR) and development of type 2 diabetes mellitus (T2DM). Nowadays, urine as a biological sample has attracted more attention for diagnosis of disease for its special superiority; insufficient research in the study of urinary amino acid (UAA) pattern in patients with T2DM has led to the present study with the aim to determine the levels of UAAs, their excretory patterns and the association of UAA to plasma glucose and IR in patients with T2DM.MethodsQuantification of total urinary amino acids was done spectrophotometrically and the patterns of amino acid excretion was elucidated by thin layer chromatography technique. Fasting blood samples were used for plasma glucose and insulin estimation by fully automated analyzer.ResultThe levels of UAA in patients with T2DM in comparison to healthy controls were higher (p < 0.0001). The frequency of urinary phenylalanine, arginine, tryptophan, tyrosine and cysteine were significantly higher in patients with T2DM than controls. There was also a strong positive correlation of UAA levels with blood glucose levels and HOMA–IR in patients with T2DM.ConclusionOur study has shown subtle abnormalities in UAA patterns in patients with T2DM. The measurement of UAA levels and excretory pattern can be used as an index of hyperglycemia and IR which could serve as an inexpensive and non-invasive marker for T2DM. More studies are required to confirm this association.     

Rapid fabrication of zwitterionic sulfobetaine vinylimidazole-based monoliths via photoinitiated copolymerization for hydrophilic interaction chromatography

Zwitterionic sulfobetaine-based monolithic stationary phases have attracted increasing attention for their use in hydrophilic interaction chromatography. In this study, a novel hydrophilic polymeric monolith was fabricated through photo-initiated copolymerization of 3-(3-vinyl-1-imidazolio)-1-propanesulfonate (SBVI) with pentaerythritol triacrylate using methanol and tetrahydrofuran as the porogenic system. Notably, the duration for the preparation of this novel monolith was as little as 5 min, which was significantly shorter than that required for previously reported sulfobetaine-based monoliths prepared via conventional thermally initiated copolymerization. Moreover, these monoliths showed good morphology, permeability, porosity (62.4%), mechanical strength (over 15 MPa), column efficiency (51,230 plates/m), and reproducibility (relative standard deviations for all analytes were lower than 4.6%). Mechanistic studies indicated that strong hydrophilic and negative electrostatic interactions might be responsible for the retention of polar analytes on the zwitterionic SBVI-based monolith. In particular, the resulting monolith exhibited good anti-protein adhesion ability and low nonspecific protein adsorption. These excellent features seem to favor its application in bioanalysis. Therefore, the novel zwitterionic sulfobetaine-based monolith was successfully employed for the highly selective separation of small bioactive compounds and the efficient enrichment of N-glycopeptides from complex samples. In this study, we prepared a novel zwitterionic sulfobetaine-based monolith with good performance and developed a simpler and faster method for preparation of zwitterionic monoliths.     

3,5-O-二咖啡酰基奎宁酸的制备及其对HeLa细胞的抗增殖活性研究

目的 制备高纯度3，5-O-二咖啡酰基奎宁酸，并评价其对人宫颈癌HeLa细胞的抗增殖活性。方法 采用柱色谱提取法和中压液相色谱法从奇蒿花中分离、纯化得到高纯度的3，5-O-二咖啡酰基奎宁酸。采用MTT法评价该化合物对人宫颈癌HeLa细胞的体外抗增殖活性。结果 柱色谱提取的提取率和中压液相色谱法的回收率分别为99.0%，61.2%，总回收率为54.0%。随着3，5-O-二咖啡酰基奎宁酸浓度升高，HeLa细胞存活率下降，细胞形态损伤增加，受试药物的IC₅₀值为26.5µg·mL^-1。结论 本研究提供了一种简单、高效、节能的3，5-O-二咖啡酰基奎宁酸制备方法。3，5-O-二咖啡酰基奎宁酸对HeLa细胞具有一定的体外抗增殖活性。