The presence of high levels of pro-inflammatory mediators such as TNF-, IL-1 and IFN- is correlated with severe malaria [4]. elusive. In the actual study, using proteomic liquid chromatographic mass spectrometry (LC-MS/MS) and immunochemical approaches, we compared the serum protein profiles of malaria patients and healthy individuals. Particularly, we utilized the malarial HZ itself to capture serum proteins capable to bind to HZ, enabling us to identify several proteins such as apolipoprotein E (ApoE), serum amyloid A (SAA), gelsolin, complement factor H and fibrinogen that were found to differ among healthy and malaria individual. Of particular interest is LPS binding protein (LBP), which is reported herein CHMFL-BTK-01 for the first time in the context of malaria. LBP is usually produced during innate inflammatory response to gram-negative bacterial infections. The exact role of these biomarkers and acute phase responses in malaria in general and HZ in particular remains to be investigated. The identification of these inflammation-related biomarkers in malaria paves the way to potentially utilize them as diagnostic and therapeutic targets. Introduction Malaria is one of the major public health problems worldwide affecting more than 250 million people causing over one million deaths each year. Caused by the parasite and transmitted through the bite of an infected female mosquito, malaria still remains to CHMFL-BTK-01 be a major public health burden particularly to the developing nations [1]. In uncomplicated malaria cases, some of the malaria symptoms include fever, chills, sweats, headache, nausea, vomiting and general malaise. Severe mediated malaria, however, is characterized by cerebral malaria, severe anemia, renal failure, metabolic acidosis, and hyperparasitemia, where more than 5% of the red blood cells (RBC) are infected by the parasites [2]. Severe malaria, which is CHMFL-BTK-01 a complex multi-system disorder, has clinical similarities with sepsis. For example, metabolic acidosis, which is the excessive acidity in the blood and tissue fluids, is also observed in severe malaria patients [3]. During malaria infection, several inflammatory mediators are involved. The presence of high levels of pro-inflammatory mediators such as TNF-, IL-1 and IFN- is correlated with severe malaria [4]. Furthermore, there is a marked increase in plasma concentrations of adhesion receptors such as ICAM-1 and E-selectin [5]. In addition to cytokines, others biomarkers have been used to discriminate cerebral malaria from uncomplicated malaria, such as serum angiopoietin-1 and -2 (ANG-1 and ANG-2). The levels of ANG-1 significantly decrease while ANG-2 increases in cerebral malaria patients [6], [7]. Moreover, ANG-2, which is the angiogenic factor modulating endothelial activation, is significantly elevated in severe malaria patients [8]. Elevated levels of serum and cerebrospinal fluid apoptotic factors such as IP-10 (INF-inducible protein 10 KDa), IL-1ra, sTNF-R1, sTNF-R2 and sFas were also correlated with cerebral malaria-associated mortality in children [9]. Peripheral levels of IL-10, TNF- and ferritin were elevated in inflammatory placental malaria whereas the level of leptin showed a marked decrease [10]. In this line of thought, discovery CHMFL-BTK-01 of biomarkers could help to discriminate malaria severity, as well as their role in the development of malaria-related pathologies in conjunction with parasite bi-product such as hemozoin (HZ). During its intraerythrocytic stage, the obligate intracellular malaria parasite utilizes hemoglobin of the RBCs. This hemoglobin proteolysis results in the production of toxic CHMFL-BTK-01 free heme and the parasite developed a heme detoxification mechanism that results in the formation of heme dimmers, called HZ [11]. Each erythrocytic cycle in a malaria patient with 1C10% parasitemia is believed to produce 0.2C2 g of hemozoin [12]. During malaria infection, HZ is released into the circulation with the merozoites during the rupture of infected RBCs and it induces the production of several proinflammatory molecules and for 5 min. The washing and centrifugation was repeated four times to ensure the maximal removal of unbound proteins. After the last wash, each sample was subject to SDS-PAGE Rabbit Polyclonal to NAB2 followed by silver staining or Western blotting analysis; and mass spectrometry analysis. Silver staining HZ-bound proteins were resolved by standard 12% SDS-polyacrylamide gel and subsequently visualized by silver staining as described by [16] with slight modification. After electrophoresis, the gel slab was first fixed in 40% methanol/5% acetic acid for 20 min followed by washing with 30% methanol for 10 min. The washing was repeated with water for 10 min to remove the remaining acid. Gel sensitization was achieved by 1 min incubation in 0.02% sodium thiosulphate, followed by 2 washes with distilled water for 1 min each. The gel was then submerged in 0.2% silver nitrate solution and incubated for 30 min at RT. After incubation, the silver nitrate was discarded; the gel slab was rinsed twice with water.