Innate immune detection of the type III secretion apparatus through the NLRC4 inflammasome

The mammalian innate immune system uses Toll-like receptors (TLRs) and Nod-LRRs (NLRs) to detect microbial components during infection. Often these molecules work in concert; for example, the TLRs can stimulate the production of the proforms of the cytokines IL-1β and IL-18, whereas certain NLRs trigger their subsequent proteolytic processing via caspase 1. Gram-negative bacteria use type III secretion systems (T3SS) to deliver virulence factors to the cytosol of host cells, where they modulate cell physiology to favor the pathogen. We show here that NLRC4/Ipaf detects the basal body rod component of the T3SS apparatus (rod protein) from S. typhimurium (PrgJ), Burkholderia pseudomallei (BsaK), Escherichia coli (EprJ and EscI), Shigella flexneri (MxiI), and Pseudomonas aeruginosa (PscI). These rod proteins share a sequence motif that is essential for detection by NLRC4; a similar motif is found in flagellin that is also detected by NLRC4. S. typhimurium has two T3SS: Salmonella pathogenicity island-1 (SPI1), which encodes the rod protein PrgJ, and SPI2, which encodes the rod protein SsaI. Although PrgJ is detected by NLRC4, SsaI is not, and this evasion is required for virulence in mice. The detection of a conserved component of the T3SS apparatus enables innate immune responses to virulent bacteria through a single pathway, a strategy that is divergent from that used by plants in which multiple NB-LRR proteins are used to detect T3SS effectors or their effects on cells. Furthermore, the specific detection of the virulence machinery permits the discrimination between pathogenic and nonpathogenic bacteria.     

The targeted intracellular delivery of cytochrome C protein to tumors using lipid-apolipoprotein nanoparticles

Intracellular-acting therapeutic proteins offer a promising clinical alternative to extracellular-acting agents, but are limited in efficacy by their low permeability into the cell cytoplasm. We have developed a nanoparticle (NP) composed of lipid (DOTAP/DOPE) and apolipoprotein (APOA-I) to mediate the targeted delivery of intracellular-acting protein drugs to non-small cell lung tumors. NPs were produced with either GFP, a fluorescent model protein, or cytochrome C (cytC), an inducer of apoptosis in cancer cells. GFP and cytC were separately conjugated with a membrane permeable sequence (MPS) peptide and were admixed with DOPE/DOTAP nanoparticle formulations to enable successful protein loading. Protein-loaded NPs were modified with DSPE-PEG-Anisamide to enable specific NP targeting to the tumor site in a xenograft model. The resulting particle was 20-30 nm in size and exhibited a 64-75% loading efficiency. H460 cells treated with the PEGylated MPS-cytC-NPs exhibited massive apoptosis. When MPS-GFP-NPs or MPS-cytC-NPs were intravenously administered in H460 tumor bearing mice, a specific tumor targeting effect with low NP accumulation in the liver was observed. In addition, MPS-cytC-NP treatment provoked a tumor growth retardation effect in H460 xenograft mice. We conclude that our NP enables targeted, efficacious therapeutic protein delivery for the treatment of lung cancer.     

Association of home visitors' and mothers' attachment style with family engagement

Family engagement in home visiting (HV) is challenging. This study related attachment security of home visitors (n=48) and mothers (n=328) to family engagement in an HV program to prevent child maltreatment. Attachment security was assessed by using the Attachment Style Questionnaire to measure attachment anxiety and attachment avoidance. Family engagement was defined as receipt of a high dose of visits and home visitor response to intimate partner violence (IPV) and poor maternal mental health as measured via record review and by maternal report of trust in the home visitor. Home visitors had lower attachment anxiety and avoidance than mothers. Families were likely to receive a high dose of services if either the mother or home visitor was high on attachment anxiety. Home visitors high on attachment anxiety were less likely to respond to IPV and poor maternal mental health; home visitors high on attachment avoidance were more likely to respond to these issues. Mothers with high attachment anxiety were less trusting of highly anxious home visitors and more trusting of avoidant home visitors. HV programs should consider the influence of both home visitor and maternal attachment style on family engagement. © 2010 Wiley Periodicals, Inc.     

The effects of salt on the physicochemical properties and immunogenicity of protein based vaccine formulated in cationic liposome

Recently, we have developed a simple and potent therapeutic cancer vaccine consisting of a cationic lipid and a peptide antigen. In this report, we expanded the utility of this formulation to protein based vaccines. First, we formulated the human papillomavirus (HPV) 16 E7 protein (E7) in different doses of DOTAP liposome. The results showed that these formulations failed to regress an established tumor. However, when sodium chloride (30 mM) was added to the DOTAP (100 nmol)/E7 (20 microg) formulation, anti-tumor activity was generated in the immunized mice. Correlatively, 30 mM NaCl in the DOTAP/E7 protein formulation increased the particle size from approximately 350 to 550 nm, decreased the protein loading capacity (from 95 to 90%), and finally increased the zeta potential (from 29 to 38 mV). Next, a model protein antigen ovalbumin (OVA) was formulated in different doses of DOTAP liposomes. Similarly, the results showed that 20 microg OVA formulated in 200 nmol DOTAP with 30 mM NaCl had the best OVA-specific antibody response, including both IgG(1) and IgG(2a), suggesting both Th1 and Th2 immune responses were generated by this formulation. In conclusion, we have expanded the application of cationic DOTAP liposome formulation to protein based vaccines and also identified that small amounts of salt could change the physicochemical properties of the vaccine formulation and enhance the activity of the DOTAP/protein based vaccine. The enhancement of immune responses by salt is possibly due to its interference of the electrostatic interaction between the cationic lipid and the protein antigen to facilitate the antigen release from the carrier and at the same time activate the antigen presenting cells.     

Obesity and coronary artery disease risk in Native Americans

With the aging of the US population and the increase in hypertension, diabetes mellitus, and obesity, the prevalence of chronic kidney disease (CKD) is increasing in the United States. Its prevalence rate has risen to 13.1% of the US population. Patients with CKD experience poor outcomes and have high health care costs. Chronic kidney disease is also a major cardiovascular disease risk factor. In fact, most people with CKD die of heart disease before they progress to end-stage renal disease. The National Kidney Foundation has produced evidencebased guidelines known as the Kidney Disease Outcomes Quality Initiative (KDOQI). These guidelines outline many things that the primary care physician can do to delay the progression of CKD, and to arrange for early referral for the prevention of future complications. However, there is limited knowledge and uptake of these guidelines because of their length and and complexity. Patients with CKD risk factors, hypertension, diabetes mellitus, cardiovascular disease, a family history of CKD, and those older than 60 years should be screened using 2 tests: 1) the estimated glomerular filtration rate and 2) the urinary albumin-creatinine ratio. These tests allow the diagnosis and stratification of CKD into 5 stages. This article synthesizes the key evidence-based behaviors and clinical action plan that primary care physicians can implement to treat CKD and its complications.     

胎盘早剥产妇、新生儿血及脐血中组织因子和组织因子途径抑制物的变化

目的:检测胎盘早剥产妇血、新生儿血及脐血组织因子及组织因子途径抑制物蛋白的变化,以及这些变化对其凝血功能的影响。方法:收集2006-10/2007-04在中山大学附属第一医院产科的胎盘早剥产妇血(足月或早产不限)、分娩胎儿的脐带血及新生儿血标本各11例,4例为足月产,其余均为胎龄29～36周早产标本,男婴5份,女婴6份。正常对照15份来自同期广州市妇婴医院产妇及分娩胎儿。标本收集时间母亲血为分娩前24h内、脐血为分娩即刻、新生儿血为出生后2h内抽取,排除标准为产妇本次分娩前无血液系统疾病史,未使用过对凝血功能产生影响的药物,采用酶联免疫吸附方法测定血浆组织因子及组织因子途径抑制物抗原水平,并与正常分娩产妇对照组进行比较。病理及正常标本均经产妇及家属知情同意后获得。结果:①胎盘早剥产妇血、新生儿血及脐血血浆中组织因子测定值明显高于正常对照组(P<0.05)。②胎盘早剥产妇血、新生儿血浆及脐血中组织因子途径抑制物测定值较正常分娩产妇降低,差异具有显著性意义(P<0.01)。③胎盘早剥产妇血、新生儿和脐血组织因子途径抑制物/组织因子比值均较正常分娩产妇明显降低(P<0.01)。结论:胎盘早剥时产妇血、脐血及新生儿血促凝血活性增强,抗凝血活性降低,与其高凝状态相关。     

膳食补充剂黑醋与红曲单用及联用对中老年人血脂的影响

目的:观察并比较黑醋与红曲单用及联用对中国中老年人降低血脂的效能。方法:试验于2003-10/11在大连医科大学中日合作医药科学研究所进行。选择30名年龄为45￣65岁的中老年人。30名受试者被随机分为3组,每组10人,分别服用黑醋丸6粒,红曲丸6粒或黑醋 红曲丸6粒,1次/d,连续4周;3种膳食补充剂黑醋丸、红曲丸及黑醋 红曲丸由含有不同剂量的黑醋固形物、红曲粉末K-F、柠檬酸钠、大豆油、蜂蜡、脂肪酸甘油脂组成(由日本国第一药品工业株式会社提供)。各组服用剂量分别为每天黑醋固形物360mg,红曲粉72mg及黑醋固形物360mg 红曲粉72mg。服用2和(或)4周后,检测血脂和血糖,同时用血清样品检测肝功与肾功能。实验结果用SPSS统计软件进行方差分析。结果:30名中老年人全部进入结果分析。①受试者连续3d每日营养素摄入量:3组之间热量、胆固醇、膳食纤维、宏量或微量营养素摄入量差距均无显著性。②血清脂质浓度和计算的脂质比:2,4周血清三酰甘油含量,黑醋 红曲组低于基础值(分别下降(24.6±4.9)%,(27.6±3.4)%,P<0.05);红曲组与基础值比较,差异无显著性[分别下降(6.5±3.4)%,(11.6±4.5)%];黑醋组与基础值比较,差异无显著性[分别下降(10.2±7.3)%,(12.4±8.2)%]。2,4周血清胆固醇、低密度脂蛋白胆固醇含量,黑醋 红曲丸、红曲丸和黑醋丸组均比基础值降低,差异无统计学意义。3种膳食补充剂对血清高密度脂蛋白胆固醇及计算的脂质比值无明显改变。血糖含量无变化。未发现对受试对象产生明显不良反应。结论:服用黑醋 红曲丸能明显降低受试者血清三酰甘油含量。黑醋与红曲合用对高脂血症者可能有健康效益。     

人参皂甙Rb_1和Rg_1对小鼠中枢神经递质受体和脑内蛋白质合成的影响

应用放射配基结合法测定了人参皂甙Rb_1和Rg_1对α_1,α_2,β肾上腺素能受体、M胆碱能受体、5-羟色胺、多巴胺及GABA等七种受体的作用。结果均不能证明它们对这七种受体有亲和力。但给动物ip药物连续5天,Rb_1和Rg_1均能使中枢M胆碱受体密度显著增高。Rb_1及Rg_1还能显著增加脑内蛋白质的含量。上述实验结果对解释人参的中枢作用提供了重要证据。     

楤木根皮中皂甙化学成分的研究

楤木根皮中的主要成分是皂甙。将总皂甙以柱层析分离,得到三个化合物(Ⅰ～Ⅲ),经光谱分析和化学方法证明,Ⅰ为楤木皂甙A(araloside A);Ⅱ为银莲花甙(narcissiflorine);Ⅲ是3-O-[β-D-吡喃葡萄糖-(1→2)-β-D-吡喃木糖-(1→)2)-α-L-阿拉伯糖]-齐墩果酸。Ⅲ为一新皂甙,命名为楤木皂甙D(araloside D)。