staphylococcus aureus biofilms mediate keratinocyte apoptosis

金黄色葡萄球菌生物膜介导角质形成细胞凋亡

• 批准号: 8496719
• 负责人: KELLY R KIRKER
• 金额: $ 6.84万
• 依托单位: MONTANA STATE UNIVERSITY - BOZEMAN
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8496719
• 关键词: ATM gene; ATM wt Allele; Abbreviations; Affect; Alkanesulfonates; Apoptosis; Apoptosis Inhibitor; Apoptotic; Biological Assay; Biology; Caring; Caspase; Cell Culture Techniques; Cell Death; Cells; Characteristics; Chronic; Coculture Techniques; Conditioned Culture Media; Culture Media; DNA Fragmentation; DNA Nucleotidylexotransferase; Data; Development; Diabetes Mellitus; Diagnostic; Fermentation; Fluorescein; Fluorescein-5-isothiocyanate; Future; Glyceraldehyde-3-Phosphate Dehydrogenases; Goals; Growth; Health; Health Care Costs; High Pressure Liquid Chromatography; Human; In Situ Nick-End Labeling; Incidence; Induction of Apoptosis; Infection; Investigation; Isothiocyanates; Label; Lead; Mammalian Cell; Mass Spectrum Analysis; Mediating; Metabolic; Metabolism; Methods; Microbial Biofilms; Microscopic; Molecular; Molecular Analysis; Molecular Biology; Montana; Morbidity - disease rate; NOESY; Necrosis; Nuclear Magnetic Resonance; Obesity; Outcome; Pathogenesis; Pathway interactions; Patients; Penicillins; Phosphate Buffer; Phosphatidylserines; Polymerase Chain Reaction; Population; Propidium Diiodide; Research; Research Personnel; Resources; Reverse Transcription; Saline; Signal Pathway; Skin; Sodium; Sodium Chloride; Spectrum Analysis; Staining method; Stains; Staphylococcus aureus; Streptomycin; TNF Receptor-Associated Factors; TNF gene; Techniques; Testing; Tetrazolium; Therapeutic Intervention; Time; Tumor Necrosis Factor-alpha; United States; Universities; Work; Wound Healing; aging population; apoptosis inducing factor; aspartyl-glutamyl-valyl-aspartyl-p-nitroanilide; aspartylglutamate; career; experience; gel electrophoresis; keratinocyte; killings; liquid chromatography mass spectrometry; metabolomics; multidisciplinary; nuclear Overhauser enhancement; prevent; response; small molecule; social; soy; stem; wound

DESCRIPTION (provided by applicant): PROJECT SUMMARY/ABSTRACT Chronic wounds affect approximately 6.5 million patients in the United States and represent $25 billion in annual health care costs. This burden is rapidly increasing due to escalating health care costs, an aging population, and the rising incidence of diabetes and obesity. Studies demonstrate that biofilms contribute to wound chronicity, and that Staphylococcus aureus is a prevalent species found in chronic wound biofilms; however, how biofilms contribute to the delayed re-epithelialization remains unclear. It was recently demonstrated that planktonic S. aureus and S. aureus biofilms have differential lethal effects on keratinocytes (KC). Therefore, it is hypothesized that planktonic S. aureus produce soluble factors that induce KC necrosis; whereas S. aureus biofilms produce soluble factors that mediate specific apoptotic pathways in KC. This hypothesis will be tested by comprehensively investigating KC cultures exposed to either biofilm-conditioned medium or planktonic-conditioned medium for apoptotic characteristics, including phosphatidylserine translocation and DNA fragmentation. Next, KC cultures will also be analyzed for the induction of the intrinsic and/or extrinsic apoptotic pathways. More specific molecular analysis of KC apoptosis will be investigated using a large format quantitative polymerase chain reaction array. Finally, bacterial products produced by S. aureus biofilms will be identified using nuclear magnetic resonance and mass spectrometry analysis, and the identified products will be screened to determine if they are apoptosis-inducing factors. The goals of this investigation include identifying the molecular pathways mediated by S. aureus biofilms and the specific bacterial products that induce KC apoptosis. Indentifying clearly defined signaling pathways and apoptosis- inducing bacterial products may provide a molecular rational for future chronic wound therapies and diagnostics, which may have a significant impact on patient health.

在美国，慢性伤口影响了大约650万患者，每年的医疗费用为250亿美元。由于医疗保健费用不断上升、人口老龄化以及糖尿病和肥胖症发病率不断上升，这一负担正在迅速增加。研究表明，生物膜有助于伤口慢性，金黄色葡萄球菌是慢性伤口生物膜中常见的物种；然而，生物膜如何促进延迟的再上皮化仍不清楚。最近的研究表明，浮游金黄色葡萄球菌和金黄色葡萄球菌生物膜对角质形成细胞（KC）具有不同的致死作用。因此，假设浮游金黄色葡萄球菌产生可溶因子诱导KC坏死；而金黄色葡萄球菌生物膜产生可溶性因子，介导KC中特定的凋亡途径。这一假设将通过全面研究暴露于生物膜条件培养基或浮游条件培养基中的KC培养物的凋亡特征（包括磷脂酰丝氨酸易位和DNA片段化）来验证。接下来，还将分析KC培养物对内在和/或外在凋亡途径的诱导作用。更具体的分子分析KC细胞凋亡将研究使用大格式定量聚合酶链反应阵列。最后，利用核磁共振和质谱分析对金黄色葡萄球菌生物膜产生的细菌产物进行鉴定，并对鉴定的产物进行筛选，以确定其是否为凋亡诱导因子。本研究的目的包括确定金黄色葡萄球菌生物膜介导的分子途径和诱导KC凋亡的特定细菌产物。明确定义的信号通路和诱导细胞凋亡的细菌产物可能为未来的慢性伤口治疗和诊断提供分子理性，这可能对患者健康产生重大影响。