Clinical Investigations In Infectious And Autoimmune Dis

该计划的目标集中在传染病和自身免疫性疾病的潜在病理事件的细胞、分子和生化特征上，长期意图是开发新的治疗干预方法。由于感染、炎症或肿瘤造成的无法愈合的粘膜和皮肤损伤可能会变得慢性和虚弱。尽管慢性创面愈合问题和过度疤痕对健康造成了深远的影响，但正常和病理性愈合的机制尚不清楚。此外，粘膜上皮愈合通常更快，几乎没有疤痕，这促使对皮肤和口腔粘膜伤口修复进行比较，以确定可能对临床延迟愈合反应有利的控制因素。在最近的研究中，不仅发现皮肤和口腔创伤的愈合反应的动力学、细胞组成和结果有很大的变化，而且分泌性白细胞蛋白酶抑制物(SLPI)被确定为口腔创伤的关键调节因子。年龄、激素和免疫缺陷在炎性损伤修复中起着重要作用，无论是在愈合率方面，还是在瘢痕形成的程度上，部分归因于老年人炎症反应的改变，中性粒细胞占主导地位，弹性蛋白酶过多，单核细胞流入延迟。延迟的皮肤愈合反应可以在动物模型中模拟，包括去卵巢的啮齿动物和SLPI基因敲除小鼠。使用这些临床前模型，局部应用肿瘤坏死因子拮抗剂(TNFpha抗体、沙利度胺)、重组SLPI和合成纤维连接蛋白多肽可以逆转延迟愈合，这些多肽可以阻断下游的炎症事件，扰乱信号转导途径，以改善过度热情的白细胞募集和促进基质降解。基于这些药物和其他药物在逆转未愈合伤口方面的成功局部应用，正在评估其他新的方法来抑制炎症部位持续的白细胞募集和过多的弹性蛋白酶释放，以评估它们在促进愈合过程中的有效性。 人类口腔粘膜中的组织损伤也激活了一系列在愈合过程中重要的转录事件，这些事件尚未明确定义。为了确定这些事件的特征并确定未来研究的潜在基因靶点，我们在组织损伤的临床模型中使用了cdna表达阵列。分别于拔除第三磨牙前、术后2-4小时或48小时取材。用实时荧光定量聚合酶链式反应对杂交模式进行分析和验证。在提取之前，活检的粘膜组织的特征是组成表达的全套基因。损伤后，分析发现参与转录、炎症和重塑的基因表达存在差异。伤后2~4h，Fos、Jun、早期生长反应蛋白等基因表达上调，细胞间黏附相关基因表达下调。在损伤后48小时，基因图谱已经转向组织重塑。在这些新的研究中，我们确定了在正常口腔粘膜中结构性表达的基因以及粘膜组织损伤后的转录事件，这可能有助于识别新的治疗靶点。 在由细菌和/或其产物(牙周组织等)、病毒(HIV-1)、其他病原体或自身免疫反应(干燥综合征)引起的临床衍生的慢性炎症病变中，使用激光捕获显微切割技术表征系统免疫反应以及病变细胞成分，以及蛋白质组和转录组反应(cDNA表达阵列)，为改善组织退化和延迟修复的靶点提供了新的见解。

The objectives of this program focus on the cellular, molecular and biochemical characterization of the underlying pathologic events in infectious and autoimmune diseases, with the long-term intent to develop novel approaches to therapeutic intervention. Nonhealing mucosal and cutaneous lesions due to infection, inflammation or neoplasia can become chronic and debilitating. Despite the profound health effects that chronic wound healing problems and excessive scarring pose, the mechanisms underlying normal and pathologic healing are little understood. Moreover, mucosal epithelial healing is typically more rapid and nearly scarless, prompting a comparison of cutaneous and oral mucosal wound repair to identify controlling factors which may be of benefit in clinical delayed healing responses. In recent studies, not only were the kinetics, cellular composition, and outcome of the healing response found to be substantially altered between cutaneous and oral wounds, but secretory leukocyte protease inhibitor (SLPI) was identified as a pivotal regulatory factor in oral wounds. Age, hormones and immune deficiency play a significant role in inflammatory lesion repair, both in terms of rate of healing and the degree of scarring incurred, attributed in part, to altered inflammatory responses in the aged, with a predominance of neutrophils, excess elastase and a delay in monocyte influx. The delayed cutaneous healing response can be mimicked in animal models including ovariectomized rodents and SLPI knockout mice. Using these pre-clinical models, delayed healing can be reversed by the topical application of TNF antagonists (TNFalpha antibody, thalidomide), recombinant SLPI and synthetic fibronectin peptides, which block downstream inflammatory events and disrupt signal transduction pathways to ameliorate the overzealous leukocyte recruitment and promote matrix degradation. Based on the successful topical delivery of these and other agents in reversing nonhealing wounds, additional novel approaches for inhibiting the persistent leukocyte recruitment and overabundant elastase released at the inflammatory site are being evaluated for their efficacy in facilitating the healing process. Tissue injury in the oral mucosa of humans also activates a cascade of transcriptional events important during the healing process that are not yet clearly defined. To characterize these events and identify potential gene targets for future studies, we used cDNA expression arrays in a clinical model of tissue injury. Mucosal biopsies were taken before 3rd molar extraction, 2-4 hours post-operatively, or at 48 hours. Hybridization patterns were analyzed and validated using real-time PCR. Prior to extraction, the biopsied mucosal tissues were characterized by a panoply of genes that were constitutively expressed. After injury, analysis revealed differential expression of genes involved in transcription, inflammation and remodeling. At 2-4 hours after injury, genes such as Fos, Jun and early growth response protein were upregulated, while genes responsible for inter-cellular adhesion were downregulated. At 48 hours after injury, the gene profile had shifted towards tissue remodeling. In these new studies, we identify genes constitutively expressed in normal oral mucosa and transcriptional events following mucosal tissue injury, which may be useful in identifying new therapeutic targets. In clinically-derived chronic inflammatory lesions, perpetuated by bacteria and/or their products (periodontal tissues, etc.), viruses (HIV-1), other pathogens, or autoimmune responses (Sjogren's Syndrome), characterization of the systemic immune response, as well as the lesion cellular constituents using laser capture microdissection, and proteome and transcriptome responses (cDNA expression array), offers new insight into targets for amelioration of tissue degradation and delayed repair.