中药化学生物学——“中药化学”与“生物学”交叉形成的新兴学科

中医药学是我国最具原创特色的科学领域,是中国走向世界的一张名片。但是由于中医药学自身遵循一套区别于西方现代医学体系的独特的科学思想和理论体系,长期以来难以被现代社会广泛接受与认可,也严重影响了中医药的国际化。因此,如何创建一套既体现中医药自身特色,又能够桥接现代科学理论的研究策略,全面系统地揭示中医药的独特科学内涵,一直是人们关注的热点问题。课题组经过在中药学领域多年的实践探索与科学思想凝练,首次提出了中药化学生物学(traditional Chinese medicine chemical biology,TCM chemical biology,TCMCB)的概念。其核心思想就是以中医药理论为指导,将中药药效成分作为化学工具,探讨中药调控生命过程的科学本质,架起中医与现代医学之间的桥梁。特别是中药化学生物学聚焦于阐释中药作用靶点和分子机制,发现中药调控疾病进程的基本规律,最终科学诠释中医药理论。该研究思路不仅科学诠释中医药防治疾病的分子机制,而且发现活性分子及其作用靶点,促进创新药物的发展;探讨生命过程的科学本质和疾病发生发展的规律,促进生命科学和现代医学的发展,具有重要意义。该文详细介绍了中药化学生物学的定义、学术思想、研究方法及其技术体系、科学意义,以期为中医药学发展提供新的研究思路和学术思想。

荒漠肉苁蓉总苷对酒精性肝损伤小鼠的保护作用研究

现代药理研究表明荒漠肉苁蓉具有肝保护作用,但其活性部位及作用机制并不清楚。为了明确荒漠肉苁蓉发挥肝保护作用的活性部位,本研究建立了急性酒精性肝损伤小鼠模型,并在此基础上对荒漠肉苁蓉的不同提取物部位(荒漠肉苁蓉总苷、荒漠肉苁蓉多糖和荒漠肉苁蓉寡糖)进行肝保护活性筛选。连续灌胃给药14天后,检测小鼠的肝脏病理结构及脂质沉积,并采用免疫荧光检测核因子E2相关因子(nuclear factor E2 related factors, Nrf-2)、胞质蛋白伴侣分子(kelch-like ECH-associated protein-1, Keap-1)及质膜囊泡相关蛋白1 (plasmalemma vesicle-associated protein-1, PV1)的蛋白表达,采用ELISA方法检测血清中谷丙转氨酶(alanine aminotransferase, ALT)、谷草转氨酶(aspartate aminotransferase, AST)、内毒素(endotoxin, ET)、二胺氧化酶(diamine oxidase, DAO)和D-乳酸(Dlactic acid, D-LA)含量,及肝脏中超氧化物歧化酶(superoxide dismutase, SOD)、谷胱甘肽过氧化物酶(glutathione peroxidase, GSH-Px)及过氧化氢酶(catalase, CAT)活性和丙二醛(malondialdehyde, MDA)含量。动物实验操作和福利均遵循北京大学医学部实验动物伦理与动物福利委员会的规定。结果显示,与模型组相比,荒漠肉苁蓉总苷能够改善肝脏病理结构及降低肝脏中脂质沉积和血清中ET、DAO和D-LA含量。同时,荒漠肉苁蓉总苷能够促进Nrf-2入核并降低Keap-1和PV1表达。由此可见,荒漠肉苁蓉总苷在400 mg·kg^(-1)剂量下对急性酒精性肝损伤具有保护作用,其作用机制可能与激活Nrf-2/Keap-1通路和改善肠壁完整性有关。

Ginsenoside Rk1 suppresses pro-inflammatory responses in lipopolysaccharide-stimulated RAW264.7 cells by inhibiting the Jak2/Stat3 pathway

The saponin ginsenoside Rk1 is a major compound isolated from ginseng.Ginsenoside Rk1 has been reported to have anti-inflammatory and anti-tumor properties and to be involved in the regulation of metabolism.However,the effect and mechanism of anti-inflammatory action of ginsenoside Rk1 has not been fully clarified.We investigated whether ginsenoside Rk1 could suppress the inflammatory response in lipopolysaccharide-stimulated RAW264.7 macrophages and to explore its mechanism of the action.RAW264.7 cells were treated with LPS(1 μg×mL^(–1))in the absence or the presence of Ginsenoside Rk1(10,20,and 40 μmol×L^(–1)).Then the inflammatory factors were tested with Griess reagents,ELISA,and RT-PCR.The proteins were analyzed by Western blotting.Ginsenoside Rk1 inhibited lipopolysaccharide-induced expression of nitric oxide(NO),interleukin(IL)-6,IL-1β,tumor necrosis factor(TNF)-α,and monocyte chemotactic protein(MCP)-1.Ginsenoside Rk1 inhibited the lipopolysaccharide-stimulated phosphorylation of NF-κB and janus kinase(Jak)2 and signal transducer and activator of transcription(Stat)3 at Ser727 and Tyr705.These data suggested that ginsenoside Rk1 could inhibit expression of inflammatory mediators and suppress inflammation further by blocking activation of NF-κB and the Jak2/Stat3 pathway in LPS-stimulated RAW264.7 cells.

Anti-inflammatory iridoids from the stems of Cistanche deserticola cultured in Tarim Desert

In order to determine the chemical constituents of Cistanche deserticola cultured in Tarim desert,a systematically phytochemical investigation was carried out.The constituents were isolated by silica gel,Sephadex LH-20,MCI gel,ODS column chromatography,and semi-preparative HPLC.Their structures were determined on the basis of MS and NMR spectroscopic analyses,by chemical methods,and/or comparison with literature data.The anti-inflammatory activities of the isolates were evaluated for their inhibitory effects on the lipopolysaccharide(LPS)-induced nitric oxide(NO)production in BV-2 mouse microglial cells.Nine iridoids were isolated and identified as cistadesertoside A(1),cistanin(2),cistachlorin(3),6-deoxycatalpol(4),gluroside(5),kankanoside A(6),ajugol(7),bartsioside(8),and 8-epi-loganic acid(9).Compound 9 exhibited potent inhibition on the NO production with an IC_(50) value being 5.2 μmol·L^(-1),comparable to the positive control quercetin(4.3 μmol·L^(-1)).Compound 1 was a new iridoid,and compounds 5,6,and 8 were isolated from this species for the first time.

Protosappanin A exerts anti-neuroinflammatory effect by inhibiting JAK2-STAT3 pathway in lipopolysaccharide-induced BV2 microglia

Microglial activation and resultant neuroinflammatory response are implicated in various brain diseases including Alzheimer's disease and Parkinson's disease. Treatment with anti-neuroinflammatory agents could provide therapeutic benefits for such disorders. Protosappanin A(PTA) is a major bioactive ingredient isolated from Caesalpinia sappan L.. In this work, the anti-neuroinflammatory effects of PTA on LPS-stimulated BV2 cells were investigated and the underlying mechanisms were explored. Results showed that PTA significantly inhibited the production of TNF-α and IL-1β in LPS-activated BV2 microglia. Moreover, the mR NA expressions of IL-6, IL-1β, and MCP-1 were reduced by PTA in a dose-dependent manner. Furthermore, PTA suppressed JAK2/STAT3-dependent inflammation pathway through down-regulating the phosphorylation of JAK2 and STAT3, as well as STAT3 nuclear translocation against LPS treatment. These observations suggested a novel role for PTA in regulating LPS-induced neuroinflammatory injuries.

Banxia Xiexin Decoction(半夏泻心汤)Treats Diabetic Gastroparesis through PLC-IP3-Ca^2+/NO-cGMP-PKG Signal Pathway

Objective To test the effect of Banxia Xiexin Decoction(半夏泻心汤,BXD)on the contraction and relaxation of gastric smooth muscle(SM)in diabetic gastroparesis(DGP)model rats,and to explore the mechanism of BXD in the prevention and treatment of DGP through experiments of signal pathway both in vivo and in vitro.Methods Sixty Sprague-Dawley rats were divided into 6 groups according to a random number table:control group,model group,high-,medium-and low-dose BXD groups(9.2,4.6 and 1.8 g/(kg·d),respectively),and domperidone group(10 mg/(kg·d)),10 rats per group.DGP model was established initially by a single intraperitoneal injection of streptozotocin(STZ),and was confirmed by recording gastric emptying,intestinal transport velocity and gastric myoelectric activity of rats after 2 months.Each group was treated with a corresponding drug for 4 weeks.The mRNA and protein expressions of phospholipase C(PLC),inositol triphosphate(IP3),neuronal nitric oxide synthase(nNOS),and cyclic guanosine monophosphate(cGMP)dependent protein kinase G(PKG)were detected by reverse transcription-polymerase chain reaction and Western blot,respectively,while nitric oxide(NO)and cGMP expressions were detected by enzyme-linked immunosorbent assay.Gastric tissues were obtained from rats for primary cell culture preparation.Gastric SM cells were treated with 0.8µmol/L of STZ or STZ plus 1,000,500 and 200µg/mL of BXD or STZ plus 2.5µmol/mL of domperidone for 24,48,72 or 96 h,respectively.The length of gastric SM cells and intracellular Ca^2+concentration([Ca^2+]i)before and after BXD treatment was measured.Results Compared with the model group,high-and medium-dose BXD and domperidone significantly increased the expressions of PLC,IP3,NO,nNOS,cGMP and PKG in rat’s gastric tissue(P<0.01).Gastric SM cells treated with BXD showed a time-and dose-dependent increase in cell viability(P<0.01).The treatment with high-and medium-dose BXD and domperidone inhibited the increase in gastric SM cells length and increased[Ca^2+]i compared with the model cells(P<0.01).Conclusions Treatment with high-and medium-dose BXD significantly attenuated STZ-induced experimental DGP in rats.The therapeutic effect of BXD on DGP rats might be associated with the PLC-IP3-Ca^2+/NO-cGMP-PKG signal pathway.

Extract of Fructus Schisandrae chinensis Inhibits Neuroinflammation Mediator Production from Microglia via NF-κB and MAPK Pathways

Objective: To investigate the anti-neuroinflammation effect of extract of Fructus Schisandrae chinensis(EFSC) on lipopolysaccharide(LPS)-induced BV-2 cells and the possible involved mechanisms. Methods: Primary cortical neurons were isolated from embryonic(E17-18) cortices of Institute of Cancer Research(ICR) mouse fetuses. Primary microglia and astroglia were isolated from the frontal cortices of newborn ICR mouse. Different cells were cultured in specific culture medium. Cells were divided into 5 groups: control group, LPS group(treated with 1 μg/mL LPS only) and EFSC groups(treated with 1 μg/mL LPS and 100, 200 or 400 mg/mL EFSC, respectively). The effect of EFSC on cells viability was tested by methylthiazolyldiphenyltetrazolium bromide(MTT) colorimetric assay. EFSC-mediated inhibition of LPS-induced production of pro-inflammatory mediators, such as nitrite oxide(NO) and interleukin-6(IL-6) were quantified and neuron-protection effect against microglia-mediated inflammation injury was tested by hoechst 33258 apoptosis assay and crystal violet staining assay. The expression of pro-inflammatory marker proteins was evaluated by Western blot analysis or immunofluorescence. Results: EFSC(200 and 400 mg/mL) reduced NO, IL-6, inducible nitric oxide synthase(iN OS) and cyclooxygenase 2(COX-2) expression in LPS-induced BV-2 cel s(P<0.01 or P<0.05). EFSC(200 and 400 mg/mL) reduced the expression of NO in LPS-induced primary microglia and astroglia(P<0.01). In addition, EFSC al eviated cel apoptosis and inflammation injury in neurons exposed to microglia-conditioned medium(P<0.01). The mechanistic studies indicated EFSC could suppress nuclear factor(NF)-κB phosphorylation and its nuclear translocation(P<0.01). The anti-inflammatory effect of EFSC occurred through suppressed activation of mitogen-activated protein kinase(MAPK) pathway(P<0.01 or P<0.05). Conclusion: EFSC acted as an anti-inflammatory agent in LPS-induced glia cel s. These effects might be realized through blocking of NF-κB activity and inhibition of MAPK signaling pathways.

Three new coumarins and a new coumarin glycoside from Micromelum integerrimum

Three new coumarins,integmarins A–C(1–3),and a new coumarin glycoside,integmaside A(4)were isolated from the leaves and stems of Micromelum integerrimum.Their structures were elucidated on the basis of 1 D and 2 D NMR and MS data,and their absolute configurations were assigned according to the ECD data of the in situ formed transition metal complexes and comparison of experimental and calculated ECD data.Compounds 1 and 2 are two rare coumarins with butyl and propyl moieties at the C-6 position;compound 3 is a novel coumarin with a highly oxidized prenyl group,and compound 4 is a rare bisdihydrofuranocoumarin glycoside.