关键词: 单细胞测序, 脓毒症, 血小板, 标志分子, 信号通路, 免疫功能

Abstract:

Objective We systematically analyze the changes in the count and function of platelet at the early sepsis based on clinical study and single cell sequencing. Methods Clinical data of sepsis patients at the early stage were collected and had been compared between different prognostic groups in the prospective case?control study. The independent risk factors of death were analyzed by logistic regression， and the predictive efficacy of clinical indicators was evaluated by receiver operating characteristic （ROC） curve. The healthy volunteers and sepsis patients were recruited. Clinical researchers collected peripheral venous blood samples for sorting cell samples to carry out single?cell RNA sequencing （sc?RNA seq）. Through bioinformatics techniques， we analyzed the changes in platelet count， the significantly differential?expressed genes and its enriched functional signaling pathways in the early stages of sepsis. Results （1） A total of 224 patients were enrolled， with a 90 day survival rate of 70.5%. Compared with the survival group， the count of platelet and MAP in the death group at the early stage of sepsis were significantly lower， but the plasma lactate content and SOFA score were significantly higher. （2） Based on single cell sequencing technology， cells are annotated as six groups. The proportion of innate immune cells （neutrophils， monocytes， and dendritic cells） was significantly increased in the early stage of sepsis compared to the healthy volunteers （2.15∶1）， while platelets significantly decreased （0.31∶1）. （3） Through bioinformatics technology， CD41/CD42a/CD61 was identified as platelet specific molecules， with significantly increased expression levels in sepsis. Three molecules can distinguish platelets together. （4） 771 genes were significantly upregulated and 1101 genes were significantly downregulated in platelets of patients with sepsis， including core molecules involved in physiological functions such as cell adhesion， chemotaxis， and immune response. Functional analysis suggests that differentially expressed genes are enriched in coagulation， immune functions and cell death， participating in oxidative phosphorylation， leukocyte chemotaxis， iron death， and NOD like receptor signaling pathways. Conclusion Reduced platelet count is associated with poor prognosis in the early stage of sepsis. The specific high expression molecules CD41/CD42a/CD61 that are significantly upregulated in platelets can serve as biomarkers for platelets. Platelets not only mediate cell adhesion and coagulation cascade， but also participate in functional changes such as immune cell chemotaxis， inflammatory response， and the pathological death of inflammatory cells.

Key words: single?cell RNA sequencing, sepsis, platelets, marker molecules, signal pathway, immunologic function