Anna Oommen

Active epilepsy as an index of burden of neurocysticercosis in Vellore district, India.

Objective: To determine the contribution of neurocysticercosis (NCC) to the causation of active epilepsy (AE) in a south Indian community. Methods: We conducted a door-to-door survey of 50,617 people between the ages of 2 and 60 years in a rural (38,105 people) and urban setting (12,512 people) in the Vellore district of the south Indian state of Tamil Nadu to identify patients with AE. Patients with AE were investigated with a contrast-enhanced CT scan and serologic study using enzyme-linked immunotransfer blot (EITB) for cysticercal antibodies. Results: We identified 194 patients with AE. The prevalence of AE was 3.83 per 1,000 people, with the prevalence in the urban clusters more than twice that in the rural clusters (6.23 vs 3.04 per 1,000) (p < 0.0001). A diagnosis of NCC was made in 46 (28.4%) of the 162 patients undergoing a CT scan, and EITB was positive in 21 (13%) patients. Overall, 55 (34%) patients were diagnosed with NCC (11 definitive NCC and 44 probable NCC). There was no significant difference in the prevalence of NCC causing AE in the urban (1.28 per 1,000) and rural (1.02 per 1,000) communities. Conclusions: NCC is the cause of nearly one-third of all cases of AE in both the urban and rural regions. Extrapolating our results to the country as a whole leads to an estimated disease burden of 1 million patients in India with AE attributable to NCC.

Monocrotophos induced oxidative damage associates with severe acetylcholinesterase inhibition in rat brain.

BACKGROUND Neurotoxicity of organophosphate pesticide poisoning, a lead cause of death in South Asia, has not been clearly elucidated. Organophosphates inhibit acetylcholinesterase and neurotoxicity is primarily a result of acetylcholine induced hyperactivation in different regions of the brain. Neurotoxicity also results from oxidative stress induced by acetylcholinesterase inhibition in the brain. Determining the severity of acetylcholinesterase inhibition that induces oxidative damage may help in developing strategies that protect the brain from organophosphate induced toxicity. AIM To determine the level of acetylcholinesterase inhibition that induces oxidative stress in the brain following organophosphate pesticide poisoning. METHODS Brains of rats subject to acute monocrotophos poisoning (0.8 LD(50) by gavage) were assessed for acetylcholinesterase activity, antioxidant response and oxidative damage 2.5 and 8h after poisoning and on recovery from poisoning 24h later after poisoning. Assessments were made in the cortex, striatum and hippocampus, cholinergic rich regions and cerebellum, targets of organophosphate pesticide poisoning. Analysis was in comparison to non poisoned controls. RESULTS High acetylcholinesterase activities were noted in striatum followed by hippocampus, cerebellum and cortex. Acute severe monocrotophos poisoning inhibited acetylcholinesterase 87% in striatum, 67% in hippocampus, 58% in cerebellum, 53% in cortex and increased glutathione levels significantly in all brain regions 2.5h after poisoning. Significant lipid peroxidation and antioxidant enzymes were induced 8h after poisoning, directly correlated to high acetylcholinesterase inhibition (>67%). Recovery from monocrotophos poisoning was associated with absence of lipid peroxidation in the brain although acetylcholinesterase inhibition persisted. CONCLUSIONS Neurotoxicity of monocrotophos poisoning is characterized by oxidative damage in regions of the brain that exhibit high acetylcholinesterase activity and severe acetylcholinesterase inhibition. Recovery from poisoning is associated with prolonged induction of antioxidants that protect against oxidative damage.

Oxidative Stress, NF-κB and the Ubiquitin Proteasomal Pathway in the Pathology of Calpainopathy

The neuromuscular disorder, calpainopathy (LGMD 2A), is a major muscular dystrophy classified under limb girdle muscular dystrophies. Genetic mutations of the enzyme calpain 3 cause LGMD 2A. Calpainopathy is phenotypically observed as progressive muscle wasting and weakness. Pathomechanisms of muscle wasting of calpainopathy remain poorly understood. Oxidative stress, NF-κB and the ubiquitin proteasomal pathway underlie the pathology of several muscle wasting conditions but their role in calpainopathic dystrophy is not known. Oxidative and nitrosative stress, the source of reactive oxygen species, NF-κB signaling and protein ubiquitinylation were studied in 15 calpainopathic and 8 healthy control human muscle biopsies. Oxidative stress and NF-κB/IKK β signaling were increased in calpainopathic muscle and may contribute to increased protein ubiquitinylation and muscle protein loss. Preventing oxidative stress or inhibition of NF-κB signaling could be considered for treatment of LGMD 2A.

A scalable low-cost cGMP process for clinical grade production of the HIV inhibitor 5P12-RANTES in Pichia pastoris

In the continued absence of an effective anti-HIV vaccine, approximately 2 million new HIV infections occur every year, with over 95% of these in developing countries. Calls have been made for the development of anti-HIV drugs that can be formulated for topical use to prevent HIV transmission during sexual intercourse. Because these drugs are principally destined for use in low-resource regions, achieving production costs that are as low as possible is an absolute requirement. 5P12-RANTES, an analog of the human chemokine protein RANTES/CCL5, is a highly potent HIV entry inhibitor which acts by achieving potent blockade of the principal HIV coreceptor, CCR5. Here we describe the development and optimization of a scalable low-cost production process for 5P12-RANTES based on expression in Pichia pastoris. At pilot (150 L) scale, this cGMP compliant process yielded 30 g of clinical grade 5P12-RANTES. As well as providing sufficient material for the first stage of clinical development, this process represents an important step towards achieving production of 5P12-RANTES at a cost and scale appropriate to meet needs for topical HIV prevention worldwide.

Involvement of oxidative stress, nuclear factor kappa B and the ubiquitin proteasomal pathway in dysferlinopathy.

AIMS Dysferlinopathies are autosomal recessive neuromuscular disorders arising from mutations of the protein dysferlin that preferentially affect the limbs which waste and weaken. The pathomechanisms of the diseases are not known and effective treatment is not available. Although free radicals and upstream signaling by the redox sensitive transcription factor, NF-κB, in activation of the ubiquitin pathway are shown to occur in several muscle wasting disorders, their involvement in dysferlinopathy is not known. This study analyzed the role of oxidative stress, NF-κB and the ubiquitin pathway in dysferlinopathic muscle and in dysferlin knockdown human myoblasts and myotubes. MAIN METHODS Fourteen dysferlinopathic muscle biopsies and 8 healthy control muscle biopsies were analyzed for oxidative stress, NF-κB activation and protein ubiquitinylation and human primary myoblasts and myotubes knocked down for dysferlin were studied for their state of oxidative stress. KEY FINDINGS Dysferlinopathic muscle biopsies showed NF-κB p65 signaling induced protein ubiquitinylation in response to oxidative stress. Dysferlin knock down primary muscle cell cultures confirmed that oxidative stress is induced in the absence of dysferlin in muscle. SIGNIFICANCE Anti-oxidants that also inhibit nitrosative stress and NF-κB activation, might prove to be of therapeutic benefit in slowing the progression of muscle wasting that occurs with dysferlinopathy.

Low seroprevalence of systemic cysticercosis among patients with epilepsy in Kerala – South India

PURPOSE Neurocysticercosis (NCC) is considered to be rare in Kerala state, India, although it is an important cause of epilepsy in many other parts of India. Our objective was to test this notion by determining the seroprevalence of cysticercosis (CC) in an unselected sample of persons with epilepsy and comparing it to that of persons without epilepsy living in Kerala. METHODS Individuals with active epilepsy (AE) who had never resided outside Kerala state for more than one month and were attending our center for epilepsy care constituted the cases. Sex-matched persons without epilepsy who had never resided outside Kerala state for more than one month constituted the controls. The demographic details, occupation, and food habits (for the cases and controls), as well as clinical characteristics and imaging (for cases only) were recorded. Sera separated from blood drawn by venipuncture from the cases and controls were assayed for cysticercal antibodies by enzyme-linked immunoelectrotransfer blot (EITB). RESULTS Of the 80 persons with AE, 12 were seropositive for cysticercus antibodies (15%; 95% CI: 8.8-24.4); among the 68 controls, 7 were seropositive (10.3%; 95% CI: 5.1-19.8). The odds ratio (OR) for seropositivity in the epilepsy group (1.54) was not statistically significant (95% CI: 0.6-4.2). Among the 69 patients who had a brain computed tomography (CT) scan or magnetic resonance imaging (MRI), none had features diagnostic of NCC. Gender, diet (vegetarian vs non-vegetarian, consumption of raw vegetables), drinking water status (clean vs unclean), residence (rural vs urban), exposure to manure, and animal rearing including pigs did not have any association with seropositivity. CONCLUSION Among the residents of Kerala, most epilepsy is not related to cysticercosis.

Host Th1/Th2 immune response to Taenia solium cyst antigens in relation to cyst burden of neurocysticercosis

Neurocysticercosis (NCC), Taenia solium larval infection of the brain, is an important cause of acquired seizures in endemic countries, which relate to number, location and degenerating cysts in the brain. Multicyst infections are common in endemic countries although single‐cyst infection prevails in India. Single‐cyst infections in an endemic country suggest a role for host immunity limiting the infection. This study examined ex vivo CD4+ T cells and in vitro Th1 and Th2 cytokine responses to T. solium cyst antigens of peripheral blood mononuclear cells of healthy subjects from endemic and nonendemic regions and of single‐ and multicyst‐infected patients for association with cyst burden of NCC. T. solium cyst antigens elicited a Th1 cytokine response in healthy subjects of T. solium‐endemic and T. solium‐non‐endemic regions and those with single‐cyst infections and a Th2 cytokine response from subjects with multicyst neurocysticercosis. Multicyst neurocysticercosis subjects also exhibited low levels of effector memory CD4+ T cells. Th1 cytokine response of T. solium exposure and low infectious loads may aid in limiting cyst number. Th2 cytokines and low effector T cells may enable multiple‐cyst infections to establish and persist.

The effect of acute severe monocrotophos poisoning on inhibition, expression and activity of acetylcholinesterase in different rat brain regions.

AIM This study examined the acute effects of severe monocrotophos (MCP) poisoning on AChE inhibition, mRNA expression and recovery of acetylcholinesterase activity in different regions of the rat brain. STUDY Wistar rats were administered monocrotophos (0.8LD(50)) by oral gavage to elicit severe effects of acute poisoning and were sacrificed 2.5 h, 24 h, 7 days, 14 days and 1 month after poisoning. Acetylcholinesterse activity, mRNA and protein were assessed in cortex, striatum, hippocampus and cerebellum. RESULTS Acute monocrotophos administration resulted in significant AChE inhibition (50-82%) in the rat brain regions 2.5 h after poisoning. AChE inhibition was associated with down regulation of synaptic AChE mRNA 24 h after poisoning in cortex and striatum. Partial recovery of AChE activity was observed 24 h after poisoning associated with increased catalytic efficiency (K(m)) of the enzyme. The recovery of AChE mRNA and protein levels to normal occurred in 7 days in cortex and cerebellum and over one month in striatum and hippocampus. CONCLUSION Cholinergic neurotoxicity of acute severe monocrotophos poisoning is characterized by high acetylcholinesterase inhibition, downregulation of acetylcholinesterase mRNA and slow recovery of acetylcholinesterase activity in brain regions. De novo synthesized acetylcholinesterase is associated with increased catalytic efficiency that may contribute in restoring cholinergic function.

Economic implications of three strategies for the control of taeniasis

Objective  To evaluate the cost‐effectiveness of three strategies for the control of taeniasis in a community, in terms of cost per case treated.

Monocrotophos toxicity and bioenergetics of muscle weakness in the rat.

Acute organophosphate pesticide poisoning is a common medical emergency with high fatality in agricultural communities of Asia. Organophosphate compounds inhibit acetylcholinesterase and prolonged neuromuscular weakness is a major cause of morbidity and mortality of poisoning. Organophosphate pesticide induced muscle weakness may not only arise from inhibition of acetylcholinesterase but also from non-cholinergic pathomechanisms, particularly mitochondrial dysfunction, affecting the production of sufficient ATP for muscle function. This study examined whether muscle weakness in rats subject to monocrotophos toxicity (0.8LD₅₀) was caused by inhibition of ATP synthesis, by oxidative phosphorylation and glycolysis, in addition to inhibition of muscle acetylcholinesterase. Severe muscle weakness in rats following monocrotophos administration was associated with inhibition of muscle acetylcholinesterase (30-60%) but not with reduced ATP production. The rats rapidly recovered muscle strength with no treatment. The ability of rats to spontaneously reactivate dimethoxy phosphorylated acetylcholinesterase and efficiently detoxify organophosphates may prevent severe inhibition of muscle acetylcholinesterase following acute severe monocrotophos poisoning. This may protect rodents against the development of prolonged muscle weakness induced by organophosphates.