Synthesis, characterization, and in vitro antitumor properties of gold(III) compounds with the traditional Chinese medicine (TCM) active ingredient liriodenine

Abstract  

Liriodenine, an oxoaporphine alkaloid with anticancer activity isolated from Zanthoxylum nitidum (rutaceous anticancer traditional Chinese medicine), was selected as a bioactive ligand to react with HAuCl₄ and NaAuCl₄ to afford [LH][AuCl₄] (1) and [AuCl₃L] (2), respectively (where L is liriodenine). The structures of 1 and 2 were characterized by IR spectroscopy, electrospray ionization mass spectrometry, ¹H-NMR spectroscopy, and elemental analysis. The single-crystal X-ray diffraction analysis of 1 revealed that it is an ionic compound consisting of protonated liriodenine cation [LH]⁺ and [AuCl₄]⁻ anion. The spectroscopic analysis showed that 2 is a coordination compound, in which one liriodenine coordinates to gold via its 7-N donor. In aqueous solution, 1 is relatively stable, but 2 undergoes rapid hydrolysis. The in vitro cytotoxicity towards five human tumor cell lines shows that 1 and 2 manifest roughly similar biological behavior and appreciable antiproliferative properties, with IC₅₀ values falling in the 2–16 μM range. The flow-cytometric analysis of 1 and 2 suggests that both compounds induced an S-phase arrest. Compounds 1 and 2 significantly poison topoisomerase I in vitro at low concentration (25 μM or less). DNA binding studies indicate that both 1 and 2 interact with DNA mainly via intercalation between the neighboring base pairs of the DNA double helix. Electrostatic interactions of 1 and 2 with the polyanionic DNA phosphate backbone may reinforce the intercalation because both 1 and 2 are composed of planar cationic species.     

Progression of Cartilage Degradation, Bone Resorption and Pain in Rat Temporomandibular Joint Osteoarthritis Induced by Injection of Iodoacetate

Background

Osteoarthritis (OA) is an important subtype of temporomandibular disorders. A simple and reproducible animal model that mimics the histopathologic changes, both in the cartilage and subchondral bone, and clinical symptoms of temporomandibular joint osteoarthritis (TMJOA) would help in our understanding of its process and underlying mechanism.

Objective

To explore whether injection of monosodium iodoacetate (MIA) into the upper compartment of rat TMJ could induce OA-like lesions.

Methods

Female rats were injected with varied doses of MIA into the upper compartment and observed for up to 12 weeks. Histologic, radiographic, behavioral, and molecular changes in the TMJ were evaluated by light and electron microscopy, MicroCT scanning, head withdrawal threshold test, real-time PCR, immunohistochemistry, and TUNEL assay.

Results

The intermediate zone of the disc loosened by 1 day post-MIA injection and thinned thereafter. Injection of an MIA dose of 0.5 mg or higher induced typical OA-like lesions in the TMJ within 4 weeks. Condylar destruction presented in a time-dependent manner, including chondrocyte apoptosis in the early stages, subsequent cartilage matrix disorganization and subchondral bone erosion, fibrosis, subchondral bone sclerosis, and osteophyte formation in the late stages. Nociceptive responses increased in the early stages, corresponding to severe synovitis. Furthermore, chondrocyte apoptosis and an imbalance between anabolism and catabolism of cartilage and subchondral bone might account for the condylar destruction.

Conclusions

Multi-level data demonstrated a reliable and convenient rat model of TMJOA could be induced by MIA injection into the upper compartment. The model might facilitate TMJOA related researches.     

Wnt/beta-catenin signaling directs multiple stages of tooth morphogenesis

Wnt/beta-catenin signaling plays key roles in tooth development, but how this pathway intersects with the complex interplay of signaling factors regulating dental morphogenesis has been unclear. We demonstrate that Wnt/beta-catenin signaling is active at multiple stages of tooth development. Mutation of beta-catenin to a constitutively active form in oral epithelium causes formation of large, misshapen tooth buds and ectopic teeth, and expanded expression of signaling molecules important for tooth development. Conversely, expression of key morphogenetic regulators including Bmp4, Msx1, and Msx2 is downregulated in embryos expressing the secreted Wnt inhibitor Dkk1 which blocks signaling in epithelial and underlying mesenchymal cells. Similar phenotypes are observed in embryos lacking epithelial beta-catenin, demonstrating a requirement for Wnt signaling within the epithelium. Inducible Dkk1 expression after the bud stage causes formation of blunted molar cusps, downregulation of the enamel knot marker p21, and loss of restricted ectodin expression, revealing requirements for Wnt activity in maintaining secondary enamel knots. These data place Wnt/beta-catenin signaling upstream of key morphogenetic signaling pathways at multiple stages of tooth development and indicate that tight regulation of this pathway is essential both for patterning tooth development in the dental lamina, and for controlling the shape of individual teeth.     

腹泻仔猪源致病性大肠杆菌生物膜与耐药性及毒力的相关性

【目的】探讨腹泻仔猪源致病性大肠杆菌生物膜形成能力及其与耐药性、毒力之间的相关性。【方法】收集临床分离鉴定的129株致病性大肠杆菌,采用96孔微量板法、K-B法、微量稀释法、寇氏改良法分别测定体外生物膜形成能力、耐药表型、生物膜菌与浮游菌的最小抑菌浓度(MIC)、对小鼠的半数致死量(LD_(50))。【结果】129株致病性大肠杆菌生物膜阳性率为96.1%,以弱阳性(1+)为主;分离株对四环素、氨苄青霉素、阿莫西林的耐药率分别为92.2%、92.2%、93%,对亚胺培南的耐药率最低为1.6%,共呈现94种多重耐药谱,其中以阿莫西林-氨苄青霉素-四环素-强力霉素-复方新诺明-甲氧苄啶构成比最大,为86.0%,且其与环丙沙星、氟苯尼考、左氧氟沙星、诺氧沙星、头孢拉定及头孢哌酮的耐药性有相关性(P0.05),环丙沙星和氟苯尼考对生物膜形成菌的MIC较对应浮游菌分别提高2-16倍和8-16倍;生物膜形成能力3+的菌株LD_(50)最大。【结论】腹泻仔猪源致病性大肠杆菌普遍具有生物膜形成能力,呈现多重耐药,生物膜形成菌对环丙沙星及氟苯尼考的耐药性与生物膜形成能力呈正相关,但随着生物膜形成能力的增强,LD_(50)值则相应增大。     

糖-蛋白质之间CH-π作用研究进展

糖-蛋白质相互作用是多种生命活动的核心,其本质是协同作用的多重非共价键弱相互作用.其中,糖CH基团与蛋白质中芳环基团之间发生的CH-π作用在糖-蛋白质识别与结合中扮演了重要的角色.随着现代测试分析技术的进步,人们对CH-π作用的起源、结构特点及其能量贡献等方面的认识和理解均得到了飞速的发展.因此,进一步从仿生学角度探索CH-π作用在药物设计与研发、智能材料和智能器件、组织工程材料等方面的应用是一系列新兴研究方向,并将促进糖和蛋白质相互作用研究的发展.在本文中,首先对糖-蛋白质之间CH-π作用的各种特点做了系统性的梳理,然后从抑制淀粉样蛋白聚集、水相糖受体设计以及驱动材料宏观性质变化三个方面,讨论了现阶段人们对CH-π作用的宏观效应及潜在应用的探索,最后对研究方向做了展望.     

天鹅洲迁地保护江豚种群近亲繁殖状况评估

长江江豚(Neophocaena asiaeorientalis asiaeorientalis)已处于极度濒危状况,迁地保护被认为是避免其灭绝最有希望的保护措施。本文选用21个多态性微卫星标记对2010年10月天鹅洲迁地保护江豚种群进行了亲子鉴定和亲缘关系分析,以检测该种群的近亲繁殖状况,为种群管理提供参考信息。本研究从18个体中检测到3个父-母-子家庭,以及母子和父子各1对。由于检测到的亲子关系较少,单从亲子鉴定结果不能判断该种群是否存在近亲繁殖。然而,亲缘关系分析结果表明,该迁地保护江豚种群的平均亲缘系数r为0.118 2,候选亲本间亲缘系数r为0.115 2,均显著高于长江江豚自然种群。而且,天鹅洲迁地保护江豚种群中具有亲缘关系的个体对达26.14%,高于自然种群6倍以上。此外,该种群的近交系数(Fis)为0.046。基于亲缘系数和近交系数的分析结果均表明,该种群存在较高的近交风险或者可能已经发生近交。本研究建议将种群中亲缘关系最多的雌性F34和雄性M45移出,并以每代(约5年)按雌雄1∶1的比例引进2头可繁殖个体,以降低近亲繁殖风险。此外,建议尽快为该迁地保护江豚种群构建遗传谱系,以便今后开展种群遗传管理。     

Variations in the antioxidant system and circadian rhythms of goldfish,Carassius auratus,exposed to ammonia: profile of the effects of green LED spectra

The present study evaluated effects of green light emitting diode (LED) spectra on oxidative stress and circadian rhythms in goldfish exposed to various concentrations (0.25 and 0.5 mg/L) of NH₃, under a white fluorescent bulb (control; simulated natural period) and green LED light. We measured mRNA expression and activity of antioxidant enzymes (superoxide dismutase and catalase) and mRNA expression of circadian rhythms (period 2), in addition to levels of plasma hydrogen peroxide, cortisol and melatonin. Damage to nuclear DNA was assessed using the comet assay. All stress indicators and melatonin were significantly lower in the green LED group than in the control group. With an increase in the concentration of ammonia, the observed effects became even more significant and generally increased with time. Comparatively, damage to the nuclear DNA was greater in the 0.5 mg/L NH₃ group, and lower in the green LED group. The Period 2 mRNA expression reduced as increasing ammonia treatment but increased as green LED exposed. We have suggested that Green LED reduced levels of oxidative stress, which suggests an antioxidant effect against NH₃ toxicity. Additionally, ammonia is affected the circadian rhythms and the green LED wavelength is able to regulate effectively the circadian rhythm.     

Structure, dynamics, and selectivity of the sodium channel blocker mu-conotoxin SIIIA

mu-SIIIA, a novel mu-conotoxin from Conus striatus, appeared to be a selective blocker of tetrodotoxin-resistant sodium channels in frog preparations. It also exhibited potent analgesic activity in mice, although its selectivity profile against mammalian sodium channels remains unknown. We have determined the structure of mu-SIIIA in aqueous solution and characterized its backbone dynamics by NMR and its functional properties electrophysiologically. Consistent with the absence of hydroxyprolines, mu-SIIIA adopts a single conformation with all peptide bonds in the trans conformation. The C-terminal region contains a well-defined helix encompassing residues 11-16, while residues 3-5 in the N-terminal region form a helix-like turn resembling 3 10-helix. The Trp12 and His16 side chains are close together, as in the related conotoxin mu-SmIIIA, but Asn2 is more distant. Dynamics measurements show that the N-terminus and Ser9 have larger-magnitude motions on the subnanosecond time scale, while the C-terminus is more rigid. Cys4, Trp12, and Cys13 undergo significant conformational exchange on microsecond to millisecond time scales. mu-SIIIA is a potent, nearly irreversible blocker of Na V1.2 but also blocks Na V1.4 and Na V1.6 with submicromolar potency. The selectivity profile of mu-SIIIA, including poor activity against the cardiac sodium channel, Na V1.5, is similar to that of the closely related mu-KIIIA, suggesting that the C-terminal regions of both are critical for blocking neuronal Na V1.2. The structural and functional characterization described in this paper of an analgesic mu-conotoxin that targets neuronal subtypes of mammalian sodium channels provides a basis for the design of novel analogues with an improved selectivity profile.     

Activated Effects of Parathyroid Hormone-Related Protein on Human Hepatic Stellate Cells

Background & Aims

After years of experiments and clinical studies, parathyroid hormone-related protein(PTHrP) has been shown to be a bone formation promoter that elicits rapid effects with limited adverse reaction. Recently, PTHrP was reported to promote fibrosis in rat kidney in conjunction with transforming growth factor-beta1 (TGF-β1), which is also a fibrosis promoter in liver. However, the effect of PTHrP in liver has not been determined. In this study, the promoting actions of PTHrP were first investigated in human normal hepatic stellate cells (HSC) and LX-2 cell lines.

Methods

TGF-β1, alpha-smooth muscle actin (α-SMA), matrix metalloproteinase 2 (MMP-2), and collagen I mRNA were quantified by real-time polymerase chain reaction (PCR) after HSCs or LX-2 cells were treated with PTHrP(1–36) or TGF-β1. Protein levels were also assessed by western-blot analysis. Alpha-SMA were also detected by immunofluorescence, and TGF-β1 secretion was measured with enzyme-linked immunosorbent assay (ELISA) of HSC cell culture media.

Results

In cultured human HSCs, mRNA and protein levels of α-SMA, collagen I, MMP-2, and TGF-β1 were increased by PTHrP treatment. A similar increasing pattern was also observed in LX-2 cells. Moreover, PTHrP significantly increased TGF-β1 secretion in cultured media from HSCs.

Conclusions

PTHrP activated HSCs and promoted the fibrosis process in LX-2 cells. These procedures were probably mediated via TGF-β1, highlighting the potential effects of PTHrP in the liver.