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Stroke, or cerebral infarction, is one of the most serious complications of sickle cell anemia (SCA) in childhood, potentially leading to impaired development and life-long physical and cognitive disabilities. About one in ten children with SCA are at risk for developing overt stroke and an additional 25% may develop silent cerebral infarcts. This is largely due to underlying cerebral injury caused by chronic cerebral ischemia and vascular insult associated with SCA. We previously identified two elevated markers of cerebral injury, plasma brain-derived neurotropic factor (BDNF) and platelet-derived growth factor (PDGF)-AA, in children with SCA and high stroke risk. The objective of this study was to investigate whether neuregulin-1β (NRG-1), an endogenous neuroprotective polypeptide may also be elevated in children with SCA. Neuregulin-1β is involved in the preservation of blood brain barrier integrity and brain microvascular cell viability and is cytoprotective in conditions of heme-induced injury and ischemia. Since elevated plasma heme and ischemia are signature characteristics of SCA, we hypothesized that NRG-1 would be elevated in children with SCA, and that NRG-1 levels would also correlate with our biomarkers of cerebral injury. Plasma NRG-1, BDNF and PDGF-AA levels were measured in children with SCA and healthy Controls. Plasma NRG-1 was found to be nearly five-fold higher in those children with SCA compared to Controls. Neuregulin-1β was also positively correlated with both BDNF and PDGF-AA concentrations, but was not associated with degree of anemia, suggesting that NRG-1 production may be an endogenous response to subclinical cerebral ischemia in SCA warranting further exploration.

In malaria endemic countries, anemia in pregnant women occurs as a result of erythrocyte destruction by Plasmodium infections and other causes including malnutrition. Iron supplementation is recommended as treatment of iron-deficiency anemia. Erythrocyte destruction results in increased release of cytotoxic free heme that is scavenged by haptoglobin (Hp), hemopexin (Hx) and heme oxygenase-1 (HO-1). Paradoxically, iron supplementation in pregnant women has been reported to enhance parasitemia and increase levels of free heme. The relationship between free heme, heme scavengers, and birth outcomes has not been investigated, especially in women who are on iron supplementation. We hypothesized that parasite-infected pregnant women on routine iron supplementation have elevated heme and altered expression of heme scavengers. A cross-sectional study was conducted to determine the association between plasma levels of free heme, HO-1, Hp, Hx, and malaria status in pregnant women who received routine iron supplementation and their birth outcomes. Heme was quantified by colorimetric assay and scavenger protein concentration by ELISA. We demonstrated that iron-supplemented women with asymptomatic parasitemia had increased free heme (mean 75.6 µM; interquartile range [IQR] 38.8–96.5) compared with nonmalaria iron-supplemented women (mean 34.9 µM; IQR 17.4–43.8, P < 0.0001). Women with preterm delivery had lower levels of Hx (mean 656.0 µg/mL; IQR 410.9–861.3) compared with women with full-term delivery (mean: 860.9 µg/mL; IQR 715.2–1055.8, P = 0.0388). Our results indicate that iron supplementation without assessment of circulating levels of free heme and heme scavengers may increase the risk for adverse pregnancy outcomes.

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

Human cerebral malaria (HCM), a severe encephalopathy associated with Plasmodium falciparum infection, has a 20–30% mortality rate and predominantly affects African children. The mechanisms mediating HCM-associated brain injury are difficult to study in human subjects, highlighting the urgent need for non-invasive ex vivo human models. HCM elevates the systemic levels of free heme, which damages the blood-brain barrier and neurons in distinct regions of the brain. We determined the effects of heme on induced pluripotent stem cells (iPSCs) and a three-dimensional cortical organoid system and assessed apoptosis and differentiation. We evaluated biomarkers associated with heme-induced brain injury, including a pro-inflammatory chemokine, CXCL-10, and its receptor, CXCR3, brain-derived neurotrophic factor (BDNF) and a receptor tyrosine-protein kinase, ERBB4, in the organoids. We then tested the neuroprotective effect of neuregulin-1 (NRG-1) against heme treatment in organoids. Neural stem and mature cells differentially expressed CXCL-10, CXCR3, BDNF and ERBB4 in the developing organoids and in response to heme-induced neuronal injury. The organoids underwent apoptosis and structural changes that were attenuated by NRG-1. Thus, cortical organoids can be used to model heme-induced cortical brain injury associated with HCM pathogenesis as well as for testing agents that reduce brain injury and neurological sequelae.