Effects of anti-tau immunotherapy on reactive microgliosis, cerebral endotheliopathy, and cognitive function in an experimental model of cerebral malaria.

Cerebral malaria (CM), a potentially fatal encephalopathy caused primarily by infection with Plasmodium falciparum, results in long-term adverse neuro-psychiatric sequelae. Neural cell injury contributes to the neurological deficits observed in CM. Abnormal regulation of tau, an axonal protein pathologically associated with the formation of neurofibrillary lesions in neurodegenerative diseases, has been linked to inflammation and cerebral microvascular compromise and has been reported in human and experimental CM (ECM). Immunotherapy with a monoclonal antibody to pathological tau (PHF-1 mAB) in experimental models of neurodegenerative diseases has been reported to mitigate cognitive decline. We investigated whether immunotherapy with PHF-1 mAB prevented cerebral endotheliopathy, neural cell injury, and neuroinflammation during ECM. Using C57BL/6 mice infected with either Plasmodium berghei ANKA (PbA), which causes ECM, Plasmodium berghei NK65 (PbN), which causes severe malaria, but not ECM, or uninfected mice (Un), we demonstrated that when compared to PbN infection or uninfected mice, PbA infection resulted in significant memory impairment at 6 days post-infection, in association with abnormal tau phosphorylation at Ser202 /Thr205 (pSer202 /Thr205 ) and Ser396-404 (pSer396-404 ) in mouse brains. ECM also resulted in significantly higher expression of inflammatory markers, in microvascular congestion, and glial cell activation. Treatment with PHF-1 mAB prevented PbA-induced cognitive impairment and was associated with significantly less vascular congestion, neuroinflammation, and neural cell activation in mice with ECM. These findings suggest that abnormal regulation of tau protein contributes to cerebral vasculopathy and is critical in the pathogenesis of neural cell injury during CM. Tau-targeted therapies may ameliorate the neural cell damage and subsequent neurocognitive impairment that occur during disease.

Rapid PCR-free meat species mitochondrial DNA identification using Electric Field Induced Release and Measurement (EFIRM®).

This work details the usage of EFIRM® (Electric Field Induced Release and Measurement) for PCR-free rapid electrochemical detection of mitochondrial DNA. EFIRM® was able to perform highly sensitive detection of animal species for meat contamination testing without multistep sample lysis, DNA extraction, or PCR amplification steps, demonstrating the capability to detect the presence of foreign meat species that only constituted 0.1% of the total mass of a food sample (achieving sensitivity equivalent to that of PCR). The EFIRM® strategy utilizes surface immobilized nucleic acid probes that complement to mitochondrial sequence of Ovis Aries, Sus Scrofa, and Bos Taurus and are immobilized in a polypyrrole matrix on a 96-electrode array. Quantification was performed through amperometric measurement of oxidation-reduction reactions on a streptavidin-peroxidase enzyme chain that completes the nucleic acid complex. All electrochemical procedures were performed using a high-throughput potentiostat system that allows parallelized electrochemical measurement and interfacing to the 96-electrode array.

The novel ETA receptor antagonist HJP-272 prevents cerebral microvascular hemorrhage in cerebral malaria and synergistically improves survival in combination with an artemisinin derivative.

AIM: To investigate the association between vasculopathy and survival during experimental cerebral malaria (ECM), and to determine whether targeting the endothelin-1 (ET-1) pathway alone or in combination with the anti-malaria drug artemether (a semi-synthetic derivative of artemisinin) will improve microvascular hemorrhage and survival. MAIN METHODS: C57BL/6 mice infected with Plasmodium berghei ANKA (PbA) were randomly assigned to four groups: no treatment, artemether treated, ET(A) receptor antagonist (HJP-272) treated, or HJP-272 and artemether treated. The uninfected control mice were treated with HJP-272 and artemether. We analyzed survival, cerebral hemorrhage, weight change, blood glucose levels and parasitemia. KEY FINDINGS: Our studies demonstrated decreased brain hemorrhage in PbA-infected (ECM) mice treated when HJP-272, a 1,3,6-trisubstituted-2-carboxy-quinol-4-one novel ET(A) receptor antagonist synthesized by our group, is used in conjunction with artemether, an anti-malarial agent. In addition, despite adversely affecting parasitemia and weight in non-artemether treated infected mice, HJP-272, seemed to confer some survival benefit when used as adjunctive therapy, though this did not reach significance. SIGNIFICANCE: Previous studies demonstrate that the endothelin pathway is associated with vasculopathy, neuronal injury and inflammation in ECM. As demonstrated here, components of the ET-1 pathway may be important targets for adjunctive therapy in ECM, and may help in preventing hemorrhage and in improving survival when used as adjunctive therapy during malaria infection. The data presented suggest that our novel agent, HJP-272, may ameliorate alterations in the vasculature which can potentially lead to inflammation, neurological dysfunction, and subsequent death in mice with ECM.

Altered regulation of Akt signaling with murine cerebral malaria, effects on long-term neuro-cognitive function, restoration with lithium treatment.

Neurological and cognitive impairment persist in more than 20% of cerebral malaria (CM) patients long after successful anti-parasitic treatment. We recently reported that long term memory and motor coordination deficits are also present in our experimental cerebral malaria model (ECM). We also documented, in a murine model, a lack of obvious pathology or inflammation after parasite elimination, suggesting that the long-term negative neurological outcomes result from potentially reversible biochemical and physiological changes in brains of ECM mice, subsequent to acute ischemic and inflammatory processes. Here, we demonstrate for the first time that acute ECM results in significantly reduced activation of protein kinase B (PKB or Akt) leading to decreased Akt phosphorylation and inhibition of the glycogen kinase synthase (GSK3ß) in the brains of mice infected with Plasmodium berghei ANKA (PbA) compared to uninfected controls and to mice infected with the non-neurotrophic P. berghei NK65 (PbN). Though Akt activation improved to control levels after chloroquine treatment in PbA-infected mice, the addition of lithium chloride, a compound which inhibits GSK3ß activity and stimulates Akt activation, induced a modest, but significant activation of Akt in the brains of infected mice when compared to uninfected controls treated with chloroquine with and without lithium. In addition, lithium significantly reversed the long-term spatial and visual memory impairment as well as the motor coordination deficits which persisted after successful anti-parasitic treatment. GSK3ß inhibition was significantly increased after chloroquine treatment, both in lithium and non-lithium treated PbA-infected mice. These data indicate that acute ECM is associated with abnormalities in cell survival pathways that result in neuronal damage. Regulation of Akt/GSK3ß with lithium reduces neuronal degeneration and may have neuroprotective effects in ECM. Aberrant regulation of Akt/GSK3ß signaling likely underlies long-term neurological sequelae observed in ECM and may yield adjunctive therapeutic targets for the management of CM.

Persistent cognitive and motor deficits after successful antimalarial treatment in murine cerebral malaria.

Human cerebral malaria causes neurological and behavioral deficits which persist long after resolution of infection and clearance of parasites with antimalarial drugs. Previously, we demonstrated that during active infection, mice with cerebral malaria demonstrated negative behavioral outcomes. Here we used a chloroquine treatment model of cerebral malaria to determine whether these abnormal outcomes would be persistent in the mouse model. C57BL/6 mice were infected with Plasmodium berghei ANKA, and treated for ten days. After cessation of chloroquine, a comprehensive assessment of cognitive and motor function demonstrated persistence of abnormal behavioral outcomes, 10 days after successful eradication of parasites. Furthermore, these deficits were still evident forty days after cessation of chloroquine, indicating persistence long after successful treatment, a hallmark feature of human cerebral malaria. Thus, cognitive tests similar to those used in these mouse studies could facilitate the development of adjunctive therapies that can ameliorate adverse neurological outcomes in human cerebral malaria.