Thrombocytopenia In Intra-abdominal Infection Patients

The incidence of intra-abdominal infection which caused by trauma, surgical disease and surgical complications increased year by year. Increasing stab wounds, abdominal traffic accident injuries, gunshot wounds and a variety of complex surgical procedures made accompanied intra-abdominal infection more serious. Meanwhile, it became more challenging in the management of intra-abdominal infection since anabolic aging population and the slow recovery of the merger, diabetes and other metabolic problems.Thrombocytopenia is a very common phenomenon in patients with intra-abdominal infection. Thrombocytopenia is not just a prevalent phenomenon for surgeons but also a very important syndrome which is directly associated with mortality. Recently, the role of platelet as an immune cells is gradually clear, which makes it important to demonstrate the course of surgical patients with intra-abdominal infection and investigate appropriate interventions.In this thesis, we firstly retrospectively enrolled patients with intra-abdominal infection caused by gastrointestinal fistula in our department to assess the incidence of thrombocytopenia in those patients. We also investigated the correlation between thrombocytopenia and outcomes in those patients, as well as the risk factors for thrombocytopenia in hospital-acquired thrombocytopenic patients. In the next, we used flow cytometry method to determine the percentage of reticulated platelet in hospital-acquired thrombocytopenic patients to find out the cause of thrombocytopenia and whether platelet apoptosis is involved in the process or not. The percentage of reticulated platelet has also been used as a biomarker to predict the the prognosis of patients with intra-abdominal infections. In the animal experimental study, we injected anti-mice CD42 monoclonal antibody to C57/BL mice via tail vein to induce a thrombocytopenic mice model. We combined anti-mice CD42 monoclonal antibody and LPS to induce a septic thrombocytopenic model to determine whether platelet associated immune process has been involved in sepsis or not. In addition, we designed a prospective, open-label, multi-center clinical trial to evaluate safety and efficiency of recombinant human thrombopoietin for severe thrombocytopenic and septic patients. Over all, the series of studies in this thesis guide clinical practice and deepened the understanding of platelet-mediated function in sepsis from clinical phenomena to the mechanism, and then back to clinical intervention, namely from bedside to beach to bedside. This thesis is divided into four parts:Part I:Clinical characteristics of thrombocytopenia in intra-abdominal infection patients caused by gastrointestinal fistulaObjective:Intra-abdominal infection is a common complication of gastrointestinal fistula. Thrombocytopenia is very common in intra-abdominal infection patients. Currently, it is still not clear the rate of thrombocytopenia, the relationship between thrombocytopenia and other common clinical parameters and risk factors for hospital acquired thrombocytopenia. This part of the study will identify the incidence of thrombocytopenia in intra-abdominal infection patients caused by gastrointestinal fistula and investigate relation between thrombocytopenic events and other common clinical parameters.Material and Methods:This is a retrospective study. We enrolled patients which discharged from Department of General Surgery in Nanjing Jinling Hospital from the January 1,2013 to December 31.2014. For all patients included in the study, the records of patients including demographic data APACHE score on admission, disease severity score, blood leukocyte count and other data such as procalcitonin were recorded.Results:Median APACHE II score and SOFA score of the whole cohort was 12 and 3 respectively. The overall ICU mortality was 7.87% and the 28-day mortality was 8.98%. The incidence of thrombocytopenia among intra-abdominal infection patients was 21.73%. Regardless of preexisting or hospital-acquired one, thrombocytopenia is associated with an increased ICU mortality and 28-day mortality as well as length of ICU or hospital stay. A higher SOFA and ISTH score at admission were significant hospital-acquired thrombocytopenia risk factors.Conclusion:This is the first study to identify a high incidence of thrombocytopenia in patients with intra-abdominal infections. Our findings suggest that the inflammatory milieu of intra-abdominal infections may uniquely predispose those patients to thrombocytopenia. More effective thrombocytopenia prevention strategies are necessary in intra-abdominal infection patients.Part Ⅱ:Reasons for thrombocytopenia in intra-abdominal infection patients caused by gastrointestinal fistula and related clinical ApplicationObjective:The incidence of thrombocytopenia is relatively high in intra-abdominal infection patients caused by gastrointestinal fistula. However, the reason for thrombocytopenia is still unknown. Reticulated platelets, as reticulocytes similarly, could reflect the state of the bone marrow to generate platelets and help to distinguish increased platelet destruction from decreased platelet production. In this part of the study, the cause of hospital-acquired thrombocytopenia in patients with intra-abdominal infection caused by gastrointestinal fistula would be identified through determining the percentage of reticulated platelets. And we also explored whether platelet apoptosis participated in thrombocytopenia or not and reticulated platelets could predict the outcome of the patients or not.Material and Methods:This is a prospective observational study from June 2013 to June 2014 in intra-abdominal infection patients with gastrointestinal fistula in Department of general surgery in Nanjing jingling Hospital. Blood sample from all enrolled patients after admission would be collected to measure percentage of reticulated platelet, platelet phosphatidylserine exposure and platelet mitochondrial membrane potential.Results:This study included 130 patients and the mean level of all patients in percentage of reticulated platelet is 2.40%. The percentage of reticulated platelet is correlated with platelet count in patients with thrombocytopenia. Compared with patients with normal platelet, thrombocytopenic patients showed an increased platelet mitochondrial membrane potential depolarization significantly. Besides, reticulated platelets is associated with PCT, INR and other coagulation index. The area under the curve for the prediction outcome of enrolled patients is 0.537.Conclusion:Increased platelet destruction is the main reason for thrombocytopenia in intra-abdominal patients. Platelets Apoptosis is involved in the process of platelet destruction. Reticulated platelet could be used as a biomarker to predict outcome of intra-abdominal infection patients.Part Ⅲ:Established and Preliminary explore thrombocytopenic animal model accompanied by sepsisObjective:In the traditional theory, platelet is mainly involved in the process of blood coagulation and thrombosis. In recent years, the process of platelet-mediated immunity has been suggested by several studies. The traditional model of sepsis in mice could not avoid activation of coagulation process to demonstrate platelet induced immune process clearly. In addition, there is no generally accepted simply thrombocytopenic mouse model. In this part, we explore method to establish thrombocytopenic mice model through mouse CD42 monoclonal antibody injection via tail vein and combine it with traditional sepsis model to preliminary explore whether platelets have involved in immune process or not.Material and Methods:In this part, we selected C57/B6 mice and injected intravenously CD42 monoclonal antibody 0.4μg/g, or 2μg/g. and measure peripheral blood platelet count Oh.2h,4h,6h,8h after injection to assess the impact CD42 monoclonal antibody after intravenous injection in mice of blood. We also used LPS 0.5mg/Kg,5mg/Kg and 10mg/Kg to stimulate mice to explore build intra-abdominal infection sepsis model via LPS stimulation.Results:Peripheral blood platelet count decreased rapidly after injected intravenously CD42 monoclonal antibody for 2h, regardless of 0.4μg/g or low-dose 2μg/g. Peripheral mouse blood platelet count can be decreased by about 80%, after both 0.4μg/g or 2μg/g dose group. Unlike platelet count, other blood cell counts in mice peripheral blood remained relatively stable without significant increase or decrease. Compared with the control group, intravenous injection of 0.4μg/g CD42 monoclonal antibody in mice show no significantly difference in morphology of tissues and organs at different time points. The mice showed a mortality of 10% and 40% after intraperitoneal injection of LPS 10mg/Kg and morphology test showed a clear inflammatory change. Animal study and blood biochemistry test showed a significant changes in liver inflammation.Conclusion:Mice injected intravenously 0.4μg/g CD42 monoclonal antibody can effectively induced thrombocytopenia, intraperitoneal injection LPS 10mg/Kg dose may induce a certain degree of mortality in mouse peritoneal sepsis infection with disease models. In the case of thrombocytopenia, mortality was significantly increased in mice, as well as increased inflammation severity, suggesting that platelets directly involved in the pathophysiology of sepsis inflammatory response.Part IV:Evaluating the safety and efficacy of recombinant human thrombopoietin among severe sepsis patients with thrombocytopeniaObjective:Sepsis is still a major health problem that causes high mortality in all populations. Organ dysfunction including sepsis-associated thrombocytopenia is prevalent among sepsis patients, resulting in increasing mortality rates. Considering the clinical role of platelets, thrombocytopenia in sepsis has led to a large spend in research activity and clinical trials in this area, yet there is no consensus upon which treatment should be administered. As a result, platelet transfusion is often indicated to resolve low platelet counts, leading to an increasing risk of the multiple risks transfusion brings, such as infectious or immune system complications. Given the role of thrombopoietin in stimulating proliferation and differentiation of megakaryocytes, our previous study investigated the potential benefits of recombinant human thrombopoietin in severe sepsis patients with thrombocytopenia. However, there are several limitations in the study, which may have led to bias in our conclusion. Thus, we are conducting this study in order to evaluate the safety and efficacy of recombinant human thrombopoietin in a large, varied population.Material and Methods:The study is designed as an open-label, placebo-controlled, multi-center study in tertiary academic centers for evaluating the safety and efficacy of recombinant human thrombopoietin over placebo. An established total of 208 patients with sepsis and thrombocytopenia will undergo prospective random assignment to recombinant human thrombopoietin or placebo (a 1:1 ratio). The primary endpoint is 7-day all-cause mortality and 28-day all-cause mortality.Results:This study started from April 2014 to enter the trial operation stage, from June 2014 to enter the formal research stage, to November 2015 to stop the recruitment of patients. A total of 164 patients were recruited in this study, and 125 patients were included in the final analysis. There was no significant difference in age, sex, underlying disease, operation history, and severity of illness between the two groups. After enrolled in, platelet counts in both group continued to rise, to day 12 platelet count had no significant difference, but platelet transfusion was significantly less in TPO group. No adverse events associated with the use of TPO were observed in this study.Conclusion:The use of recombinant human thrombopoietin in combination with conventional medical therapies could significantly improve the platelet counts in patients with severe sepsis and thrombocytopenia and effectively reduce the platelet transfusion possibility.