Role of HMGB1 in cells of the circulation

The matrix of blood is a liquid plasma that transports molecules and blood cells within vessels lined by endothelial cells. High-mobility group B1 (HMGB1) is a protein expressed in blood cells. Under normal circumstances, HMGB1 is virtually absent from plasma, but during inflammation or trauma its level in plasma is increased. In resting and quiescent cells, HMGB1 is usually localized in the intracellular compartment, with the exception of motile cells that express HMGB1 on their outer surface to mediate cell migration. During cell transformation or immune cell activation HMGB1 can be actively secreted outside of the cell. Further, when a cell is damaged, HMGB1 can passively leak into extracellular environment. Extracellular HMGB1 can then participate in regulation of the immune response and under some conditions it can mediate lethality in systemic inflammatory response. The aim of this study was to evaluate the expression and functions of HMGB1 in cells of the vascular system and to investigate the prognostic value of circulating HMGB1 in severe sepsis and septic shock. HMGB1 was detected in platelets, leukocytes, and endothelial cells. HMGB1 was released from platelets and leukocytes, and it was found to mediate their adhesive and migratory functions. During severe infections the plasma levels of HMGB1 were elevated; however, no direct correlation with lethality was found. Further, the analysis of proinflammatory mechanisms suggested that HMGB1 forms complexes with other molecules to activate the immune system. In conclusion, HMGB1 is expressed in the cells of the vascular system, and it participates in inflammatory mechanisms by activating platelets and leukocytes and by mediating monocyte migration.

The incidence and outcome of severe sepsis in Finland : The Finnsepsis Study

Severe sepsis is associated with common occurrence, high costs of care and significant mortality. The incidence of severe sepsis has been reported to vary between 0.5/1000 and 3/1000 in different studies. The worldwide Severe Sepsis Campaign, guidelines and treatment protocols aim at decreasing severe sepsis associated high morbidity and mortality. Various mediators of inflammation, such as high mobility group box-1 protein (HMGB1) and vascular endothelial growth factor (VEGF), have been tested for severity of illness and outcome in severe sepsis. Long-term survival with quality of life (QOL) assessment is important outcome after severe sepsis. The objective of this study was to evaluate the incidence, severity of organ dysfunction and outcome of severe sepsis in intensive care treated patients in Finland (study I)). HMGB1 and VEGF were studied in predicting severity of illness, development and type of organ dysfunction and hospital mortality (studies II and III). The long-term outcome and quality of life were assessed and quality-adjusted life years and cost per one QALY were estimated (study IV). A total of 470 patients with severe sepsis were included in the Finnsepsis Study. Patients were treated in 24 Finnish intensive care units in a 4-month period from 1 November 2004 to 28 February 2005. The incidence of severe sepsis was 0.38 /1,000 in the adult population (95% confidence interval 0.34-0.41). Septic shock (77%), severe oxygenation impairment (71.4%) and acute renal failure (23.2%) were the most common organ failures. The ICU, hospital, one-year and two-year mortalities were 15.5%, 28.3%, 40.9% and 44.9% respectively. HMGB1 and VEGF were elevated in patients with severe sepsis. VEGF concentrations were lower in non-survivors than in survivors, but HMGB1 levels did not differ between patients. Neither HMGB1 nor VEGF were predictive of hospital mortality. The QOL was measured median 17 months after severe sepsis and QOL was lower than in reference population. The mean QALY was 15.2 years for a surviving patient and the cost for one QALY was 2,139 . The study showed that the incidence of severe sepsis is lower in Finland than in other countries. The short-term outcome is comparable with that in other countries, but long-term outcome is poor. HMGB1 and VEGF are not useful in predicting mortality in severe sepsis. The mean QALY for a surviving patient is 15.2 and as the cost for one QALY is reasonably low, the intensive care is cost-effective in patients with severe sepsis.

Inflammatory and coagulation disturbances in acute pancreatitis

Acute pancreatitis (AP), a common cause of acute abdominal pain, is usually a mild, self-limited disease. However, some 20-30% of patients develop a severe disease manifested by pancreatic necrosis, abscesses or pseudocysts, and/or extrapancreatic complications, such as vital organ failure (OF). Patients with AP develop systemic inflammation, which is considered to play a role in the pathogenesis of multiple organ failure (MOF). OF mimics the condition seen in patients with sepsis, which is characterized by an overwhelming production of inflammatory mediators, activation of the complement system and systemic activation of coagulation, as well as the development of disseminated intravascular coagulation (DIC) syndrome. Vital OF is the major cause of mortality in AP, along with infectious complications. About half of the deaths occur within the first week of hospitalization and thus, early identification of patients likely to develop OF is important. The aim of the present study was to investigate inflammatory and coagulation disturbances in AP and to find inflammatory and coagulation markers for predicting severe AP, and development of OF and fatal outcome. This clinical study consists of four parts. All of patients studied had AP when admitted to Helsinki University Central Hospital. In the first study, 31 patients with severe AP were investigated. Their plasma levels of protein C (PC) and activated protein C (APC), and monocyte HLA-DR expression were studied during the treatment period in the intensive care unit; 13 of these patients developed OF. In the second study, the serum levels of complement regulator protein CD59 were studied in 39 patients during the first week of hospitalization; 12 of them developed OF. In the third study, 165 patients were investigated; their plasma levels of soluble form of the receptor for advanced glycation end products (sRAGE) and high mobility group box 1 (HMGB1) protein were studied during the first 12 days of hos-pitalization; 38 developed OF. In the fourth study, 33 patients were studied on admission to hospital for plasma levels of prothrombin fragment F1+2 and tissue factor pathway inhibitor (TFPI), and thrombin formation capacity by calibrated automated thrombogram (CAT); 9 of them developed OF. Our results showed significant PC deficiency and decreased APC generation in patients with severe AP. The PC pathway defects seemed to be associated with the development of OF. In patients who developed OF, the levels of serum CD59 and plasma sRAGE, but not of HMGB1, were significantly higher than in patients who recovered without OF. The high CD59 levels on admission to the hospital seemed to be predictive for severe AP and OF. The median of the highest sRAGE levels was significantly higher in non-survivors than in survivors. No significant difference between the patient groups was found in the F1+2 levels. The thrombograms of all patients were disturbed in their shape, and in 11 patients the exogenous tissue factor did not trigger thrombin generation at all ( flat curve ). All of the patients that died displayed a flat curve. Free TFPI levels and free/total TFPI ratios were significantly higher in patients with a flat curve than in the others, and these levels were also significantly higher in non-survivors than in survivors. The flat curve in combination with free TFPI seemed to be predictive for a fatal outcome in AP.

Development of atmospheric pressure ionization ion mobility spectrometry and ion mobility spectrometry mass spectrometry

This study is focused on the development and evaluation of ion mobility instrumentation with various atmospheric pressure ionization techniques and includes the following work. First, a high-resolution drift tube ion mobility spectrometer (IMS), coupled with a commercial triple quadrupole mass spectrometer (MS), was developed. This drift tube IMS is compatible with the front-end of commercial Sciex mass spectrometers (e.g., Sciex API-300, 365, and 3000) and also allows easy (only minor modifications are needed) installation between the original atmospheric pressure ion source and the triple quadrupole mass spectrometer. Performance haracteristics (e.g.,resolving power, detection limit, transmission efficiency of ions) of this IMS-MS instrument were evaluated. Development of the IMS-MS instrument also led to a study where a proposal was made that tetraalkylammonium ions can be used as chemical standards for ESI-IMS. Second, the same drift tube design was also used to build a standalone ion mobility spectrometer equipped with a Faraday plate detector. For this highresolution (resolving power about 100 shown) IMS device, a multi-ion source platform was built, which allows the use of a range of atmospheric pressure ionization methods, such as: corona discharge chemical ionization (CD-APCI), atmospheric pressure photoionization (APPI), and radioactive atmospheric pressure chemical ionization (R-APCI). The multi-ion source platform provides easy switching between ionization methods and both positive and negative ionization modes can be used. Third, a simple desorpion/ionization on silicon (DIOS) ion source set-up for use with the developed IMS and IMS-MS instruments was built and its operation demonstrated. Fourth, a prototype of a commercial aspiration-type ion mobility spectrometer was mounted in front of a commercial triple quadrupole mass spectrometer. The set-up, which is simple, easy to install, and requires no major modifications to the MS, provides the possibility of gathering fundamental information about aspiration mobility spectrometry.