Cerebral Malaria; Mechanisms Of Brain Injury And Strategies For Improved Neuro-Cognitive Outcome

Epidemiology

Falciparum malaria is a leading cause of ill health, neuro-disability and death in tropical countries. Although 40% of the world’s population is at risk, most transmission occurs in sub-Saharan Africa where children under the age of 5 years are most affected and the incidence of disease declines in older children with increasing immunity. In South-East Asia, malaria occurs more commonly in adults but the clinical features are different(1).

Every year, there are over 500 million clinical cases. One percent of symptomatic infections may become complicated and develop into severe malaria. Severe malaria may manifest as anemia, hypoglycemia, metabolic acidosis, repeated seizures, coma or multiple organ failure and is estimated to cause over one million deaths annually(1). Cerebral malaria is the most severe neurological manifestation of severe malaria. With an incidence of 1,120/100,000/year in the endemic areas of Africa, children in this region bear the brunt. Peak incidence is in pre-school children and at a minimum, 575,000 children in Africa develop cerebral malaria annually(2). Recent reports however suggest that the incidence of severe malaria is on the decline(3, 4).

Clinical features

The clinical features of cerebral malaria have been reviewed extensively(5, 6). In this paper, we highlight some of the main features.

The World Health Organization defines cerebral malaria as a clinical syndrome characterized by coma at least 1 hour after termination of a seizure or correction of hypoglycemia, asexual forms of Plasmodium falciparum parasites on peripheral blood smears and no other cause to explain the coma(7). In practice, this definition is non-specific. Patients in whom coma is caused by other encephalopathies (e.g. viral encephalitis, poisoning and metabolic disease) or previously unrecognized neurological abnormalities but have incidental parasitemia (a common phenomenon in malaria endemic areas) may be included. Thus, in a postmortem study of Malawian children dying with a clinical diagnosis of cerebral malaria, 24% had other causes of death(8). This lack of specificity is problematic for clinical and pathogenesis studies. The increased specificity accorded by recent descriptions of retinal changes in cerebral malaria is should address this issue(9).

The clinical hallmark of cerebral malaria is impaired consciousness, with coma the most severe manifestation. This is thought to be causes by parasitized red blood cells (pRBCs) sequestered in cerebral micro-circulation, but other authors attribute the impaired consciousness to metabolic factors and inflammatory mediators(10). In African children, coma develops suddenly with seizure onset often, following 1-3 days of fever. A few children develop coma following progressive weakness and prostration. Brain swelling, intracranial hypertension, retinal changes (hemorrhages, peripheral and macular whitening, vessel discoloration and or papilledema) and brainstem signs (abnormalities in posture, pupil size and reaction, ocular movements or abnormal respiratory patterns) are commonly observed. Other systemic complications such as anemia, metabolic acidosis, electrolyte imbalance and hyperpyrexia or hypoglycemia and shock are commonly present. The prognosis is grave in deeply comatose patients with severe metabolic acidosis, shock, hypoglycemia and repeated seizures.

In adults, cerebral malaria is part of a multi-organ disease. Patients develop fever, headache, body ache and progressively, delirium and coma. Compared to African children, seizures papilledema and retinal changes are less common and coma resolution is slower. Anemia, hemoglobinuria, jaundice, shock, renal failure, lactic acidosis, abnormal bleeding, pulmonary edema and adult respiratory distress syndrome may develop. Some develop cortical infarcts and cerebral venous or dural sinus thrombosis as part of a disordered coagulation. Bacterial co-infection may be observed, particularly in those with shock, and accounts for the majority of late deaths(5, 6).

Outcome

Without treatment, cerebral malaria is invariably fatal. In children, parenteral antimalarials (cinchonoids or artemisinin derivatives) are indicated, but even with this treatment, 15-20% die. In adults however, mortality was lower if patients were treated with intravenous artesunate(11). This treatment is currently being evaluated in African children.

Earlier studies suggested that surviving patients fully recover(12) but over the past 20 years, it became clear that many children sustain significant brain injury; 11% are discharged with gross neurological deficits(13, 14). Although some gross deficits, particularly blindness, ataxia and central hypotonia improve with time(15), 25% have long-term impairments especially cognition, motor function or behavior impairments and, epilepsy develops in 10%(15-18), table 1. The prevalence (<5%) and pattern of neurological deficits are different in adults. Thus, other than causing thousands of deaths, cerebral malaria is now considered a leading cause of neuro-disability in African children. The main risk factors include repeated seizures, deep and prolonged coma, intracranial hypertension, and hypoglycemia.

Parasite sequestration in the brain

Parasite sequestration in cerebral microvasculature is thought to be a central factor in pathogenesis and the resulting pathophysiological changes in tissue around the sequestered parasites, which may explain why an intravascular parasite may cause neural dysfunction and why some patients may have a poor outcome. Clearly, there are other factors since sequestration is also observed in patients dying of other complications of falciparum malaria(20).

Sequestration results from adherence of pRBCs to the endothelial lining (cytoadherence) using parasite derived proteins exposed on erythrocyte surface(21). A group of parasite antigens including Plasmodium falciparum erythrocyte membrane protein-1 (PfEMP-1) mediate binding to host receptors of which, intercellular adhesion molecule-1 (ICAM-1) is the most important and whose expression is upregulated in areas adjacent to sequestered parasites. The sequestered parasite mass is further increased when adherent erythrocytes agglutinate with other pRBCs, form rosettes with non-parasitized erythrocytes or use platelet-mediated clumping to bind to each other. Sequestration impairs perfusion and may aggravate coma through hypoxia. Furthermore, the ability of pRBCs to deform and pass through the microvasculature is decreased(22). Therefore, hypoxia and inadequate tissue perfusion may be major pathophysiological events. Although a critical reduction in metabolite supply (oxygen and glucose) may occur, in the majority of children, significant neural tissue necrosis is unlikely since with specific antimalaria treatment, coma is rapidly reversible. However, in the presence of increased metabolic demand such as during seizures and fever, the risk of neural injury is higher and may be worse if the patient is hypoglycemic(23) or if blood flow is further compromised by intracranial hypertension(24).

Cytokines, chemokines and excitotoxicity

Cytokines and chemokines play a complex role in pathogenesis and have both protective and harmful effects. Parasite antigens released at schizogony trigger the release of both pro- and anti-inflammatory cytokines. Although the balance between these mediators is critical for parasite control, their role in pathogenesis of the neuronal damage is unclear. Tumor necrosis factor (TNF), the most extensively studied cytokine in cerebral malaria, upregulates ICAM-1 expression on the cerebral vascular endothelium increasing the cytoadhesion of pRBCs. Near areas of sequestration, there is increased local synthesis. The timing of this is important since early in disease, TNF may be protective but prolonged high levels contribute to complications(25). TNF is also involved in regulating synaptic transmission (strength, scaling and long-term potentiation)(10). Thus, cytokine mediated synaptic changes may contribute to the syndrome of cerebral malaria. Despite the prominence of TNF in pathogenesis, pentoxifylline which decreases macrophage TNF production(26, 27) and monoclonal antibodies to TNF(28), failed decrease mortality.

Several other cytokines and chemokines are important and in particular, interleukin (IL)-1b, IL-6 and IL-10(29) but low levels of the chemokine RANTES is independently associated with mortality(30). The role of nitric oxide (NO) is controversial. The association between NO activity and inducible nitric oxide synthase with pathogenesis have been inconsistent(31, 32). NO is involved in host defense, maintaining vascular status and in neurotransmission and is thought to be an effector for TNF. It is suggested that inflammatory cytokines upregulate inducible NO synthase in brain endothelial cells leading to increased NO production. Nitric oxide can cross the blood brain barrier (BBB), diffuse into brain tissue and interfere with neurotransmission and may therefore partly be responsible for the reversible coma(33).

Other inflammatory products such as the metabolites of the kynurenine pathway - quinolinic and kynurenic acid - may also be important in pathogenesis. Quinolinic acid is a NMDA receptor agonist and an excitotoxin. It causes seizures in animal models of brain disease while kynurenic acid is an antagonist and is generally thought of as neuro-protective. Excitation by quinolinic acid may contribute to convulsions in cerebral malaria. In children, there are graded increases in cerebrospinal fluid concentration across outcome groups of increasing severity(34) although in adults, increased levels were associated with impaired renal function(35). Because of the role of NMDA receptors in modulating neurotransmission and as agonists, high levels of quinolinic acid may have long-term deleterious effects on cognitive function.

Endothelial injury, apoptosis, blood-brain barrier (BBB) dysfunction and intracranial hypertension

Cytoadherence of pRBCs to the endothelium initiates a cascade of events beginning with the transcription of genes involved in inflammation, cell-to-cell signalling and signal transduction, which result in endothelial activation, release of endothelial micro-particles (EMPs) and apoptosis of host cells(36). There is widespread endothelial activation in vessels containing pRBCs and compared to other complications of falciparum malaria, significant increases in circulating EMPs are seen in patients in coma(37). In addition, interactions between pRBCs and platelets (which produce platelet microparticles) cause further injury to endothelial cells through a direct cytotoxic effect(38). Repair of the injured endothelium is also impaired as there is failure to mobilize sufficient circulating endothelial progenitor cells(39) and plasma levels of endothelial regulators, angiopoietin-1 and angiopoietin-2 are altered(40).

In murine models, apoptosis is first observed in endothelial cells and then in neurons and glia(41); the stimulus being contact of pRBCs with the endothelium(42) (figure 1). There is accumulation of activated/effector CD8 lymphocytes. Apoptosis may be induced through a perforin-dependent process, since cerebral symptoms are not seen in perforin-deficient models but only in wild forms that show high increases in perforin mRNA (43). Four patterns of axonal injury have been described; single axons, diffuse or more focal parenchymal patches and neuronal cell bodies(44) and axonal injury correlated with plasma lactate and depth of coma. Axons in children appear to be more susceptible to injury since the median CSF microtubule-associated protein tau level is 3-fold greater than in adults(45). The increased susceptibility to injury may explain the higher prevalence of sequelae in children.

Perivascular macrophages around vessels with parasites express receptors such as sialoadhesin normally present only if there has been contact with plasma proteins(46). Although disruptions at sites of sequestration can expose neurons to plasma proteins, significant leakage of plasma proteins into perivascular spaces has not been seen(47). Despite this, intracranial hypertension is common in African children; up to 40% of children with deep coma have brain swelling on computerized tomography scans(48). BBB dysfunction may contribute to the hypertension although increased cerebral volume could be caused by sequestration and increased cerebral blood flow from seizures, hyperthermia or anemia.

Intracranial hypertension reduces cerebral perfusion pressure, nutrient and oxygen delivery and can lead to global ischemic injury, herniation, brainstem compression and death(24, 49). Ischemic injury is seen on acute computerized tomography and pattern of injury is consistent with a critical reduction in perfusion pressure(48). The convalescent scans in these patients show cerebral atrophy. Many children with severe hypertension are discharged with spastic quadriplegia and subsequently, severe learning disability(24).

Cerebral blood flow and perfusion

Patients with cerebral malaria have an increased cerebral blood flow. This increase is probably an adaptive response to high a metabolic demand to match oxygen and nutrient delivery to requirements since jugular bulb venous oxygen saturation remains within normal(50). Recent studies of the retina have however provided evidence for decreased local perfusion(51, 52). In the eye, multiple discrete areas (100-1000μm) of retinal whitening are observed in most children with cerebral malaria. These areas have impaired capillary perfusion on fluorescein angiography(51), figure 2. Physiologically, reduced local perfusion is associated with an abnormal electroretinograph(52). If the retina mirrors events in the brain, similar obstruction may be present in the brain and coma in cerebral malaria may partly be a result of under-perfusion in multiple but small areas of the brain. Because the patches of brain affected are small, with early treatment and rapid relief of obstruction, there is minimal tissue necrosis and early restoration of perfusion may explain the near complete recovery of gross neurological function in the majority of patients. However, exposure to hypoxia still leaves many children with subtle (e.g. cognitive) deficits. In those who die or develop severe brain injury, the sequestered mass may be higher, blood flow obstruction not readily reversed and hypoxic and ischemic injury more widespread(53).

Regional blood flow may also be altered. In a transcranial Doppler study, sonographic abnormalities were associated with lateralizing deficits in six out of eleven children who developed severe functional deficits on recovery(54), while in patients with severe intracranial hypertension, a linear relationship between cerebral perfusion pressure and blood flow velocity was observed suggesting that auto-regulation was impaired. In addition, sonographic features suggesting progressive intracranial hypertension were seen in some who later died.

Seizures

Plasmodium falciparum is epileptogenic and the risk of seizures increases with parasitemia(55). Seizures are a common feature of childhood cerebral malaria; over 80% are admitted with seizures and seizures recur in 60% during admission(56). In other models, irreversible neuron damage is described following prolonged seizure activity; within days, edema is recognized on magnetic resonance imaging (MRI)(57) but overtime, this is replaced by local atrophy and gliosis(58). There is however no consensus as to whether seizures cause the brain injury or are a manifestation of an injured brain(59, 60). Thus, although prophylactic anticonvulsants in traumatic brain injury (TBI) prevented immediate seizure recurrence, they did not reduce the risk of subsequent epilepsy(61). Similarly, high dose prophylactic Phenobarbital in children with cerebral malaria significantly reduced seizure recurrences(62) but did not improve cognitive outcome(63). In this study however, prophylactic Phenobarbital was associated with increased mortality (from respiratory depression) and the cognitive outcome study included only half of the original cohort. The likely scenario is that brain injury is caused by the seizure initiating noxious agent. Prolonged seizures may worsen this injury and setup a vicious circle of neural injury and more seizures.

Depth, duration and cause of coma

It has been suggested that cerebral malaria is not a single homogenous syndrome rather, four distinct groups: a prolonged post-ictal state, covert status epilepticus, severe metabolic derangement and a primary neurological syndrome(64). A fifth group maybe patients with false cerebral malaria in whom coma has other causes and parasitemia is incidental(8).

Patients with prolonged post-ictal state have coma secondary to seizures, regain consciousness within 6 hours and have a good neurological recovery. Although the seizures are different from simple febrile seizures, the same risk factors may be at play. Those with convert status epilepticus on the other hand present following prolonged seizures. The physical signs of seizure activity are often so minimal that they may not be recognized. Failure to detect the state can be disastrous since these patients are hypoxic and hypercarbic from hypoventilation and at risk of aspiration. The neuro-cognitive outcome may depend on how long the seizures have lasted.

Patients with severe metabolic derangement may regain consciousness a few hours after resuscitation. Impaired consciousness is a secondary phenomenon to an unfavourable environment. Again prolonged periods of either hypoglycemia or acidosis may cause neuronal dysfunction or death and among survivors, outcome may depend on duration of exposure.

Death and neurological sequelae may arise from different mechanisms. In Gambian children, neurological sequelae were associated with repeated seizures and with deep and prolonged coma while death was associated with hypoglycemia and acidosis suggesting that most early deaths in cerebral malaria may result from an overwhelming metabolic derangement(65). Early correction of these derangements may buy time for definitive treatment. Recent phase II trials of albumin as a resuscitative fluid have supported this assertion(66) and has led to the on going Fluid Expansion as Supportive Therapy trial for very sick children in several African hospitals.

Patients with a primary neurological syndrome present with seizures often without severe metabolic disturbances and have the worst neuro-cognitive outcome. They are not severely anemic and coma persists for 24-48 hours well beyond the resolution of seizures. Intracranial hypertension is common. Coma may be a primary consequence of intracranial sequestration of malaria parasites.

Pathogenesis of some specific impairment

Neuro-protective and adjuvant therapies

Brain injury following exposure to noxious insult evolves over hours or days. This has raised the possibility that early intervention may limit injury. However, most neuro-protective interventions in man, particularly in TBI, have been disappointing. A poor understanding of pathophysiological mechanisms, properties of compounds used (e.g. appropriate therapeutic time-window), patient selection or choice of endpoints have been blamed for the failure(76). Studies in cerebral malaria have also been disappointing. These included interventions targeting elements of pathogenesis and risk factors for brain injury. Thus, there have been trials of steroids, immunoglobulin, acetyl salicylic acid, heparin, anti TNF agents, mannitol, iron chelation, micronutrients and prophylactic anticonvulsants. Because prophylactic Phenobarbital was associated with respiratory depression(62), there is an ongoing trial evaluating a less respiratory depressant prophylactic anticonvulsant, fosphenytoin.

More recent studies have examined the possibility of preventing endothelial injury or reversing rossetting and improving blood flow. In a murine model, administration of a low-molecular weight thiol, the B5 complex pro-vitamin pantethine, prevented the development of cerebral malaria. Pantethine appeared to offer this protection by down-regulating platelet reactivity and release of micro-particles from the activated endothelium. Attenuation of these processes left an intact BBB integrity(77). In another study, Glatiramer acetate, an immuno-modulatory agent, was associated with a lower risk of cerebral malaria in treated animals(78). In both cases, human trials are awaited.

Two different anti-apoptotic approaches - treatment with z-VAD (a pan-caspase inhibitor) and over-expression of Bcl-2 (a gene which prevents the release of apoptotic mediators from the mitochondria) - did not reduce cerebral malaria mortality in mice(79). However, the cytokine erythropoietin, which has anti-apoptotic, anti-inflammatory, vaso-dilating and neurotrophic properties and widespread receptors in the brain has shown greater promise. It crosses the BBB and local synthesis is increased during hypoxia. Erythropoietin provided neuro-protection in animal models of TBI, infectious and hypoxic ischemic encephalopathy(80) and offered protection against endothelial injury in the heart, causing NO mediated coronary vasodilatation, increasing blood flow, enhancing repair and reducing apoptosis(81). In adults with stroke, high dose erythropoietin reduced brain-infarct size and improved functional outcome(82). In mice with cerebral malaria, high dose erythropoietin reduced mortality by 40-90%, (reviewed in(83)) and in children, plasma levels >200u/L were associated with ~80% less risk of sequelae(84). A phase I study of erythropoietin, 1500u/kg, in children with cerebral malaria was recently concluded without any major concerns about the safety(85).

Heparan sulfate on endothelial cells mediates binding of pRBCs via DBL1 alpha domain of PfEMP1(86). Interfering with this binding can potentially reduce parasite sequestration. Earlier studies of heparin and acetyl salicylic acid were not continued because they either failed to demonstrate sufficient evidence of benefit or because of toxicity. More recently however, compounds with less anticoagulant effects have been developed. A Swedish group has recently completed a Phase I trial of DFO2, a modified form of heparin with minimal anticoagulant activity and plan to take this to Phase II studies (Anna Vogt, personal communication).

Clearly, a poor understanding of pathogenesis is a major hindrance to progress in cerebral malaria research. MRI offers a way out including a non-invasive study of the structural, metabolic, biochemical and functional features of the brain in vivo(87). A combination of time of flight angiography and diffusion weighted imaging can describe obstruction to flow and the effects of obstruction in distal areas. Cerebral blood flow can be measured using arterial spin labeling while functional MRI can be used to describe neural activity in coma. In addition, proton MR spectroscopy can be used to measure levels of substrates and metabolites.

Rehabilitation

Because of the high mortality associated with cerebral malaria, few studies have focused on rehabilitation for children with sequelae. There are no guidelines to assess impairments or guide rehabilitation(88). With declining child mortality in the region and the growing number of exposed and therefore potentially impaired children, systems are needed to detect impairments, develop and implement rehabilitation. Particular areas include physical and occupational therapy, behavior and speech therapy, cognitive rehabilitation and hearing aids. A preliminary study of cognitive rehabilitation using a computerized program has demonstrated improved neuropsychological and behavioral outcomes in exposed children suggesting that similar interventions are possible(89).