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Background: Cerebral malaria (CM) is characterized by the sequestration of Plasmodium-infected erythrocytes (pRBCs) to host brain microvasculature beds via P. falciparum erythrocyte membrane protein 1 (PfEMP1). Under normal conditions, activated protein C (APC) bound to endothelial protein C receptor (EPCR) has cytoprotective properties via the activation of protease-activated receptor 1 (PAR1). During malaria infection, pRBCs transports PfEMP1 to the membranes to bind EPCR in the same region as APC. As a result, APC is less capable of inducing cytoprotective effects via PAR1. Two studies involving adult malaria patients revealed that EPCR rs867186-GG allele is associated with protection against severe malaria, while three other studies involving children malaria patients could not show association between EPCR rs867186-GG allele and severe malaria or increased mortality among children with CM. Methods: We examined whether or not the EPCR rs867186-GG allele protects against cerebral malaria. Peripheral blood samples were collected from 47 malaria patients and four healthy individuals from a study conducted from 2004 to 2007 at the NSCB Medical College Hospital in India. CM and malaria-associated complications were defined based on WHO criteria. Genomic DNA was isolated from the peripheral blood mononuclear cells. Primer sequences were designed to contain rs867186 of the PROCR gene (NM 006404) and were used to amplify a 660 bp product as described before [1]. PCR products were purified, and DNA sequences were determined by Sanger Sequencing (www.Genewiz.com. Genewiz, NJ). Nonparametric tests were used to compare the groups. To analyze differences in allele frequencies, we used chi-squared or Fisher’s exact tests for categorical variables if the expected values were less than 5. P-value

Moringa oleifera trees grow well in Jamaica and their parts are popularly used locally for various purposes and ailments. Antioxidant activities in Moringa oleifera samples from different parts of the world have different ranges. This study was initiated to determine the antioxidant activity of Moringa oleifera grown in Jamaica. Dried and milled Moringa oleifera leaves were extracted with ethanol/water (4:1) followed by a series of liquid–liquid extractions. The antioxidant capacities of all fractions were tested using a 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. IC50 values (the amount of antioxidant needed to reduce 50% of DPPH) were then determined and values for the extracts ranged from 177 to 4458 μg/mL. Extracts prepared using polar solvents had significantly higher antioxidant capacities than others and may have clinical applications in any disease characterized by a chronic state of oxidative stress, such as sickle cell anemia. Further work will involve the assessment of these extracts in a sickle cell model of oxidative stress.

BackgroundChronic inflammation is a characteristic of sickle cell disease (SCD), and is invariably associated with vascular endothelial injury. Hydroxyurea (HU), a naturally cytotoxic chemotherapeutic agent, is the only FDA drug approved for SCD, and is therefore naturally cytotoxic. Quercetin (QCT) is a dietary flavonoid found ubiquitously in plants and foods that have anti-oxidative and anti-inflammatory characteristics. Our hypothesis is that dietary QCT will decrease cytotoxic effects of lipopolysaccharide (LPS) and HU induced vascular cell damage.MethodsLipopolysaccharide (LPS) was used to induce inflammation in immortalized mouse aortic endothelial cells (iMAECs), providing an in vitro model of inflamed endothelial cells. The cells were exposed to LPS throughout the entire experiment. Interventions included treating the LPS exposed cells with QCT, HU, or QCT + HU over 50 hours. The 50-hour period included 24 hours of varying treatments, followed by two hours of hypoxic exposure and then 24 hours under normal aerobic exposure.ResultsLDH level was significantly higher for LPS treated versus untreated cells (P = 0.0004). LPS plus 30 micromole QCT reduced the LDH (p = 0.1, trend), whereas LPS plus 100 micromoles HU, significantly increased LDH (p = 0.0004). However, LPS plus treatment with 30 micromoles QCT/100 micromoles HU, significantly reduced LDH, compared with HU alone (p = 0.0002).DiscussionThese results suggest that quercetin may be effective against vascular endothelial cell damage for iMAECsin vitro. In particular, it shows promise in preventing HU-induced cytotoxicity, surprisingly found from these results. This latter finding is important, and should be given more consideration, since HU is the only FDA-approved drug for treating sickle cell patients, and its use is rapidly increasing.

Finding a widely available cure for sickle cell anemia (HbSS) still remains a challenge one hundred years after its discovery as a genetically inherited disease. However, growing interest in the nutritional problems of the disease has created a body of literature from researchers seeking nutritional alternatives as a means of decreasing morbidity and improving quality of life among HbSS patients. This review demonstrates that over the past 30 years the role of protein/energy deficiency in HbSS has been more clearly defined via direct measurements, leading to the concept of a relative shortage of nutrients for growth and development, despite apparently adequate dietary intakes. Although there is still a paucity of data supporting the efficacy of macronutrient supplementation, it is becoming clearer that recommended dietary allowances (RDAs) for the general population are insufficient for the sickle cell patient. A similar shortage is likely to be true for micronutrient deficiencies, including recent findings of vitamin D deficiency that may be associated with incomplete ossification and bone disease, which are well known complications of HbSS disease. We conclude that there is need for more effort and resources to be dedicated to research (including supplementation studies of larger sample size) aimed at establishing specific RDAs for HbSS patients, much like the specific RDAs developed for pregnancy and growth within the general population.

Previous reports have shown that a high protein diet improves weight gain and decreases expression of inflammatory markers in weanling Berkeley transgenic sickle cell mice. The effect of this diet on the underlying histopathology, however, has not been studied. Age-matched, male C57BL/6 controls (n = 24), Berkley sickle mice (n = 31) and Townes sickle mice (n = 14) were randomized in a terminal experiment at weaning to isoenergetic diets, with either normal (20%) or high (35%) amount of energy from protein, by replacing dextrin. Tissue sampling for blinded histologic study and scoring of changes at baseline and after 3 months of feedings showed progressive siderosis and infarcts in spleen, kidney, and liver in all sickle groups, and no significant changes in age- and sex-matched normal controls. High-protein (35%) fed Berkeley sickle mice had significantly fewer (p

Background We have previously reported that free Heme generated during experimental cerebral malaria (ECM) in mice, is central to the pathogenesis of fatal ECM. Heme-induced up-regulation of STAT3 and CXCL10 promotes whereas up-regulation of HO-1 prevents brain tissue damage in ECM. We have previously demonstrated that Heme is involved in the induction of apoptosis in vascular endothelial cells. In the present study, we further tested the hypothesis that Heme reduces blood-brain barrier integrity during ECM by induction of apoptosis of brain vascular endothelial cells through STAT3 and its target gene matrix metalloproteinase three (MMP3) signaling. Methods Genes associated with the JAK/STAT3 signaling pathway induced upon stimulation by Heme treatment, were assessed using real time RT2 Profile PCR arrays. A human MMP3 promoter was cloned into a luciferase reporter plasmid, pMMP3, and its activity was examined following exposure to Heme treatment by a luciferase reporter gene assay. In order to determine whether activated nuclear protein STAT3 binds to the MMP3 promoter and regulates MMP3 gene, we conducted a ChIP analysis using Heme-treated and untreated human brain microvascular endothelial cells (HBVEC), and determined mRNA and protein expression levels of MMP3 using qRT-PCR and Western blot. Apoptosis in HBVEC treated with Heme was evaluated by MTT and TUNEL assay. Results The results show that (1) Heme activates a variety of JAK/STAT3 downstream pathways in HBVEC. STAT3 targeted genes such as MMP3 and C/EBPb (Apoptosis-related genes), are up regulated in HBVEC treated with Heme. (2) Heme-induced HBVEC apoptosis via activation of STAT3 as well as its downstream signaling molecule MMP3 and upregulation of CXCL10 and HO-1 expressions. (3) Phosphorylated STAT3 binds to the MMP3 promoter in HBVEC cells, STAT3 transcribed MMP3 and induced MMP3 protein expression in HBVEC cells. Conclusions Activated STAT3 binds to the MMP3 promoter region and regulates MMP3 in Heme-induced endothelial cell apoptosis.

Pregnancy in sickle cell disease (SCD) patients is associated with increased risk of maternal and fetal mortality. This study determines pregnancy outcomes among women with SCD delivering at Korle-Bu Teaching Hospital, Accra, Ghana. Nine hundred sixty (960) medical records of pregnant women (131 HbSS, 112 HbSC, and 717 comparison group) from 2007 to 2008 were reviewed. The HbSS women were at increased risk of eclampsia (adjusted odds ratio [AOR] = 10.56, 95% confidence interval [CI] = 3.60-30.96, P < 0.001), intrauterine growth restriction (AOR = 4.00, 95% CI = 1.38-11.64, P = 0.011), and placenta previa (AOR = 22.03, 95% CI = 9.87-49.14, P < 0.001) compared with the comparison group. The HbSC women had increased risk for intrauterine fetal death (AOR = 3.38, 95% CI = 1.15-9.96, P = 0.027) and decreased risk of delivering low birth weight babies (AOR = 0.21, 95% CI = 0.06-0.73, P = 0.014). Women with SCD in Ghana are at a greater risk of morbidity and mortality in pregnancy compared with women without hemoglobinopathies. Improved maternal and fetal outcomes in Ghanaian women with SCD can be achieved through effective intervention by health care providers with thorough knowledge about predisposing factors toward adverse outcomes.

Background The mortality of severe malaria [cerebral malaria (CM), severe malaria anemia (SMA), acute lung injury (ALI) and acute respiratory distress syndrome (ARDS)] remains high despite the availability associated with adequate treatments. Recent studies in our laboratory and others have revealed a hitherto unknown correlation between chemokine CXCL10/CXCR3, Heme/HO-1 and STAT3 and cerebral malaria severity and mortality. Although Heme/HO-1 and CXCL10/CXCR3 interactions are directly involved in the pathogenesis of CM and fatal disease, the mechanism dictating how Heme/HO-1 and CXCL10/CXCR3 are expressed and regulated under these conditions is still unknown. We therefore tested the hypothesis that these factors share common signaling pathways and may be mutually regulated. Methods We first clarified the roles of Heme/HO-1, CXCL10/CXCR3 and STAT3 in CM pathogenesis utilizing a well established experimental cerebral malaria mouse (ECM, P. berghei ANKA) model. Then, we further determined the mechanisms how STAT3 regulates HO-1 and CXCL10 as well as mutual regulation among them in CRL-2581, a murine endothelial cell line. Results The results demonstrate that (1) STAT3 is activated by P. berghei ANKA (PBA) infection in vivo and Heme in vitro. (2) Heme up-regulates HO-1 and CXCL10 production through STAT3 pathway, and regulates CXCL10 at the transcriptional level in vitro. (3) HO-1 transcription is positively regulated by CXCL10. (4) HO-1 regulates STAT3 signaling. Conclusion Our data indicate that Heme/HO-1, CXCL10/CXCR3 and STAT3 molecules as well as related signaling pathways play very important roles in the pathogenesis of severe malaria. We conclude that these factors are mutually regulated and provide new opportunities to develop potential novel therapeutic targets that could be used to supplement traditional prophylactics and treatments for malaria and improve clinical outcomes while reducing malaria mortality. Our ultimate goal is to develop novel therapies targeting Heme or CXCL10-related biological signaling molecules associated with development of fatal malaria.

The influence of host genetics on susceptibility to Plasmodium falciparum malaria has been extensively studied over the past twenty years. It is now clear that malaria parasites have imposed strong selective forces on the human genome in endemic regions. Different genes have been identified that are associated with different malaria related phenotypes. Factors that promote severity of malaria include parasitaemia, parasite induced inflammation, anaemia and sequestration of parasitized erythrocytes in brain microvasculature. Recent advances in human genome research technologies such as genome-wide association studies (GWAS) and fine genotyping tools have enabled the discovery of several genetic polymorphisms and biomarkers that warrant further study in host-parasite interactions. This review describes and discusses human gene polymorphisms identified thus far that have been shown to be associated with susceptibility or resistance to P. falciparum malaria. Although some polymorphisms play significant roles in susceptibility to malaria, several findings are inconclusive and contradictory and must be considered with caution. The discovery of genetic markers associated with different malaria phenotypes will help elucidate the pathophysiology of malaria and enable development of interventions or cures. Diversity in human populations as well as environmental effects can influence the clinical heterogeneity of malaria, thus warranting further investigations with a goal of developing new interventions, therapies and better management against malaria.