THROMBIN IN WOUND HEALING--MULTIPLE FUNCTIONS

点击翻译按钮获取中文摘要

Periodontal diseases are the pathologic outcome of infection with specific pathogens that possess virulence factors that contribute either directly or indirectly to the destruction of the supporting structures of teeth. There is overwhelming evidence that host response especially to the lipopolysaccharide of these pathogens makes a primary contribution to the bone and connective tissue destruction that define periodontal diseases. Most research clearly indicates that the polymorphonuclear leukocyte is central to the defense against periodontopathogens and that the bacterial evasion and/or neutralization of PMN anti-bacterial function are common virulence traits of the successful pathogen. Most forms of periodontal disease are characteristically episodic and represent the result of an iterative process of tissue destruction and wound healing in response to chronic exposure to noxious bacterial products. It is the overall objective of the proposed studies to begin to understand the interactions between pathogenic bacteria, and/or their metabolic products, inflammatory mediators and proinflammatory factors, both acute and chronic inflammatory cells, hemostatic and thrombic components and the initiating factors of wound healing in periodontal diseases. Specifically the subcellular localization of kallikrein in neutrophils will be determined. The signal transduction and second messengers involved in thrombin stimulation of PMN and the up regulation of kallikrein synthesis and release will be determined. The granule content of human neutrophils will be fractionated and the distribution of kallikrein determined relative to that of characterized neutrophil granule markers. Kallikrein synthesis, compartmentalization and release will be characterized following the stimulation of neutrophils with thrombin, as well as, with other well characterized agonists. Release of granule markers, respiratory burst, chemotaxis, change in cell morphology and adherence will be characterized following thrombin stimulation of neutrophils and the promyelocytic cell line, HL60 undifferentiated and differentiated to either neutrophilic or monocytic cells. Pulse-chase experiments will be used to monitor kallikrein synthesis by both human neutrophils and HL6O cells. The influence of thrombin on the phagocytosis and killing of Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis will be determined using a dual fluorescent microscopy assay. Influence of proinflammatory and prothombic components on neutrophil function will be examined in admix experiments using purified cytokines and inflammatory mediators, as well as platelets, monocytes, and extracellular matrix surfaces. Parallel studies will examine the kinetics of appearance of cells types and the appearance and influence of selected biologically relevant molecules in fluid collected from a murine subcutaneous chamber model for the evaluation of host/periodontopathogen interaction in vivo. These studies will provide insight into the complex interactions that occur between host cells, pathogenic bacteria, and biologically active molecules. Such information is essential to the development of interventive clinical approaches for bacterial mediated tissue destructive processes such as periodontal diseases.