Mechanism of action of Orthosiphon stamineus against non-alcoholic fatty liver disease: Insights from systems pharmacology and molecular docking approaches

Non-alcoholic fatty liver disease (NAFLD) is one of the most common complications of a metabolic syndrome caused by excessive accumulation of fat in the liver. Orthosiphon stamineus also known as Orthosiphon aristatus is a medicinal plant with possible potential beneficial effects on various metabolic disorders. This study aims to investigate the in vitro inhibitory effects of O. stamineus on hepatic fat accumulation and to further use the computational systems pharmacology approach to identify the pharmacokinetic properties of the bioactive compounds of O. stamineus and to predict their molecular mechanisms against NAFLD. Methods: The effects of an ethanolic extract of O. stamineus leaves on cytotoxicity, fat accumulation and antioxidant activity were assessed using HepG2 cells. The bioactive compounds of O. stamineus were identified using LC/MS and two bioinformatics databases, namely the Traditional Chinese Medicine Integrated Database (TCMID) and the Bioinformatics Analysis Tool for the Molecular Mechanism of Traditional Chinese Medicine (BATMAN-TCM). Pathway enrichment analysis was performed on the predicted targets of the bioactive compounds to provide a systematic overview of the molecular mechanism of action, while molecular docking was used to validate the predicted targets. Results: A total of 27 bioactive compounds corresponding to 50 potential NAFLD-related targets were identified. O. stamineus exerts its anti-NAFLD effects by modulating a variety of cellular processes, including oxidative stress, mitochondrial β-oxidation, inflammatory signalling pathways, insulin signalling, and fatty acid homeostasis pathways. O. stamineus is significantly targeting many oxidative stress regulators, including JNK, mammalian target of rapamycin (mTOR), NFKB1, PPAR, and AKT1. Molecular docking analysis confirmed the expected high affinity for the potential targets, while the in vitro assay indicates the ability of O. stamineus to inhibit hepatic fat accumulation. Conclusion: Using the computational systems pharmacology approach, the potentially beneficial effect of O. stamineus in NAFLD was indicated through the combination of multiple compounds, multiple targets, and multicellular components.     

Development of an IgG-Fc fusion COVID-19 subunit vaccine,AKS-452

AKS-452 is a biologically-engineered vaccine comprising an Fc fusion protein of the SARS-CoV-2 viral spike protein receptor binding domain antigen (Ag) and human IgG1 Fc (SP/RBD-Fc) in clinical development for the induction and augmentation of neutralizing IgG titers against SARS-CoV-2 viral infection to address the COVID-19 pandemic. The Fc moiety is designed to enhance immunogenicity by increasing uptake via Fc-receptors (FcγR) on Ag-presenting cells (APCs) and prolonging exposure due to neonatal Fc receptor (FcRn) recycling. AKS-452 induced approximately 20-fold greater neutralizing IgG titers in mice relative to those induced by SP/RBD without the Fc moiety and induced comparable long-term neutralizing titers with a single dose vs. two doses. To further enhance immunogenicity, AKS-452 was evaluated in formulations containing a panel of adjuvants in which the water-in-oil adjuvant, Montanide™ ISA 720, enhanced neutralizing IgG titers by approximately 7-fold after one and two doses in mice, including the neutralization of live SARS-CoV-2 virus infection of VERO-E6 cells. Furthermore, ISA 720-adjuvanted AKS-452 was immunogenic in rabbits and non-human primates (NHPs) and protected from infection and clinical symptoms with live SARS-CoV-2 virus in NHPs (USA-WA1/2020 viral strain) and the K18 human ACE2-trangenic (K18-huACE2-Tg) mouse (South African B.1.351 viral variant). These preclinical studies support the initiation of Phase I clinical studies with adjuvanted AKS-452 with the expectation that this room-temperature stable, Fc-fusion subunit vaccine can be rapidly and inexpensively manufactured to provide billions of doses per year especially in regions where the cold-chain is difficult to maintain.     

丹参二萜醌类活性成分通过PKCδ/PKD1μ信号通路抗胰腺癌的作用研究

摘要 目的 验证丹参二萜醌类活性成分对胰腺癌和多发性骨髓瘤的抑制效应，阐明其诱导胰腺癌和多发性骨髓瘤凋亡的作用机制。方法 胰腺癌细胞AsPC-1、BxPC-3用含10% gibco胎牛血清的RPMI1640培养液培养，按隐丹参酮组（30μM）、丹参新酮组（15μM）、去氢丹参新酮组（15μM）分别加药处理。cell-counting-kit-8（CCK8）法检测细胞存活率；AnnexinⅤ-FITC/PI流式细胞术检测细胞凋亡；Western Blot检测相关PKC同工酶磷酸化水平；对AsPC-1和BxPC-3细胞进行siRNA转染，Western Blot检测相关PKC同工酶磷酸化水平。结果 隐丹参酮组AsPC-1、BxPC-3细胞存活率分别为40.1%±5.0%、36.2%±5.4%；丹参新酮组AsPC-1、BxPC-3细胞存活率分别为52.1%±5.1%、47.2%±5.7%；去氢丹参新组AsPC-1、BxPC-3细胞存活率分别为46.1%±5.0%、42.2%±5.4%(P<0.01)。流式细胞术结果显示：AsPC-1组内空白对照组、隐丹参酮组、丹参新酮组、去氢丹参新酮组细胞的凋亡率分别为4.71%、30.10%、52.26%、42.30%；BxPC-3组内空白对照组、隐丹参酮组、丹参新酮组、去氢丹参新酮组细胞的凋亡率分别为5.10%、30.66%、33.76%、51.76%（P<0.01）。Western Blot检测显示隐丹参酮组、丹参新酮组、去氢丹参新酮组较空白对照组，胰腺癌AsPC-1、BxPC-3细胞p-PKD/PKCμ ser916、p-PKCδ thr505、p-PKD/PKCμ ser744/748的水平降低。Western Blot检测显示，siRNA沉默胰腺癌AsPC-1、BxPC-3细胞PKCδ，胰腺癌AsPC-1、BxPC-3细胞PKD/PKCμ ser744/748的磷酸化水平下调。结论 隐丹参酮、丹参新酮、去氢丹参新酮通过抑制PKCδthr505的磷酸化水平，继而PKD1μser744/748磷酸化水平下调，从而显著促进胰腺癌AsPC-1和BxPC-3细胞凋亡发生。     

Evaluation of protection induced by a dengue virus serotype 2 envelope domain III protein scaffold/DNA vaccine in non-human primates

We describe the preclinical development of a dengue virus vaccine targeting the dengue virus serotype 2 (DENV2) envelope domain III (EDIII). This study provides proof-of-principle that a dengue EDIII protein scaffold/DNA vaccine can protect against dengue challenge. The dengue vaccine (EDIII-E2) is composed of both a protein particle and a DNA expression plasmid delivered simultaneously via intramuscular injection (protein) and gene gun (DNA) into rhesus macaques. The protein component can contain a maximum of 60 copies of EDIII presented on a multimeric scaffold of Geobacillus stearothermophilus E2 proteins. The DNA component is composed of the EDIII portion of the envelope gene cloned into an expression plasmid. The EDIII-E2 vaccine elicited robust antibody responses to DENV2, with neutralizing antibody responses detectable following the first boost and reaching titers of greater than 1:100,000 following the second and final boost. Vaccinated and naïve groups of macaques were challenged with DENV2. All vaccinated macaques were protected from detectable viremia by infectious assay, while naïve animals had detectable viremia for 2–7 days post-challenge. All naïve macaques had detectable viral RNA from day 2–10 post-challenge. In the EDIII-E2 group, three macaques were negative for viral RNA and three were found to have detectable viral RNA post challenge. Viremia onset was delayed and the duration was shortened relative to naïve controls. The presence of viral RNA post-challenge corresponded to a 10–30-fold boost in neutralization titers 28 days post challenge, whereas no boost was observed in the fully protected animals. Based on these results, we determine that pre-challenge 50% neutralization titers of >1:6000 correlated with sterilizing protection against DENV2 challenge in EDIII-E2 vaccinated macaques. Identification of the critical correlate of protection for the EDIII-E2 platform in the robust non-human primate model lays the groundwork for further development of a tetravalent EDIII-E2 dengue vaccine.     

急性脑梗死血清C-反应蛋白的临床分析

目的观察急性脑梗死患者血清CRP在发病中的作用及临床意义。方法采用免疫比浊法对60例急性脑梗死患者的血清CRP水平进行动态观察（病程第1．7及14天），并与45例健康人进行比较。结果急性脑梗死患者血清CRP从病程第1天即升高（P〈0．01），至第7天达高峰，以后降低；重型患者明显高于轻、中型患者。结论血清CRP可能在急性脑梗死的发生、发展中起一定作用，并与病情严重程度呈正相关。     

检测子宫内膜异位症患者血清中差异表达蛋白的研究

目的：检测子宫内膜异位症（内异症）患者血清中差异表达的蛋白。方法：采用表面增强激光解吸／离子化飞行时间质谱（SELDI—TOF—MS）技术，选用WCX2蛋白质芯片对50例内异症及48例对照组血清标本进行检测以筛选内异症血清中差异表达的蛋白。结果：在Mr 0～50000范围内，检测出106个蛋白峰。内异症患者血清中差异表达的蛋白峰有4个。将发现的差异蛋白峰在Swiss蛋白数据库中搜索，发现Mr 9280蛋白峰与玻璃粘连蛋白Vitronectin相符。Vitronectin属于整合素家族，在内异症的粘附、侵袭、血管形成过程中起重要作用。其他的蛋白峰没有发现与之相匹配的蛋白，提示可能为新的蛋白质。结论：内异症患者血清中存在差异表达的蛋白，其对内异症的早期诊断具有一定的临床意义。SELDI蛋白芯片技术是一种快速、简单易行、样本用量少、高通量、重复性好的分析方法，具有广阔的临床应用前景。     

C反应蛋白测定在小儿败血症中的应用

目的　本研究的目的在了解C反应蛋白的 (CRP)测定对于小儿败血症的诊断价值。方法　连续选取2 0例白细胞总数不高的小儿败血症患者 ,作回顾性研究对象。比较其治疗前和治疗后 7～ 1 0d后疾病的痊愈或明显好转时CRP水平 ,并与体温、白细胞分类结果进行比较。结果　治疗后CRP水平 (0 57± 0 68μg/ml)显著低于治疗前水平 (5 90± 3 57μg/ml) ,P <0 0 0 1 ;治疗前CRP的阳性率为 1 0 0 % ,显著高于嗜中性粒细胞比率 >75 %的阳性率 (30 % )和体温高于 37 5℃的阳性率 (30 % ) ,P <0 0 0 5。结论　CRP可以作为一种反映细菌感染的敏感指标 ,有助于不典型小儿败血症的诊断和疗效观察。     

PTEN基因蛋白在胆囊癌组织中表达的研究

目的 :探讨抑癌基因PTEN蛋白在胆囊癌组织中的表达情况及其临床病理意义。方法 :应用免疫组化对 2 8例胆囊腺癌 ,4例胆囊鳞癌及其相应癌旁组织中PTEN蛋白进行检测 ,并结合患者的临床病理资料进行分析。结果 :PTEN蛋白在 32例癌旁组织中全都呈阳性表达 ,而癌组织中PTEN阳性表达率仅 34.4 %。胆囊癌组织中PTEN蛋白阳性表达与患者性别、年龄及组织类型无明显相关性 ,但与组织分化程度明显相关。结论 :PTEN蛋白在胆囊癌组织中阳性表达率较低 ,说明该基因失活可能在胆囊癌发生发展中起重要作用。PTEN阳性表达对胆囊癌患者的预后有一定意义