Functional consequences of familial ALS-associated SOD1L84F in neuronal and muscle cells.

Amyotrophic lateral sclerosis is a fatal neurodegenerative disorder characterized by progressive skeletal muscle denervation and loss of motor neurons that results in muscle atrophy and eventual death due to respiratory failure. Previously, we identified a novel SOD1L84F variation in a familial ALS case. In this study, we examined the functional consequences of SOD1L84F overexpression in the mouse motor neuron cell line (NSC-34). The cells expressing SOD1L84F showed increased oxidative stress and increased cell death. Interestingly, SOD1L84F destabilized the native dimer and formed high molecular weight SDS-resistant protein aggregates. Furthermore, SOD1L84F also decreased the percentage of differentiated cells and significantly reduced neurite length. A plethora of evidence suggested active involvement of skeletal muscle in disease initiation and progression. We observed differential processing of the mutant SOD1 and perturbations of cellular machinery in NSC-34 and muscle cell line C2C12. Unlike neuronal cells, mutant protein failed to accumulate in muscle cells probably due to the activated autophagy, as evidenced by increased LC3-II and reduced p62. Further, SOD1L84F altered mitochondrial dynamics only in NSC-34. In addition, microarray analysis also revealed huge variations in differentially expressed genes between NSC-34 and C2C12. Interestingly, SOD1L84F hampered the endogenous FUS autoregulatory mechanism in NSC-34 by downregulating retention of introns 6 and 7 resulting in a two-fold upregulation of FUS. No such changes were observed in C2C12. Our findings strongly suggest the differential processing and response towards the mutant SOD1 in neuronal and muscle cell lines.

Clinical and Genetic Analysis of A Father-Son Duo with Monomelic Amyotrophy: Case Report.

Monomelic Amyotrophy (MMA) is a rare neurological disorder restricted to one upper limb, predominantly affecting young males with an unknown aetiopathogenesis. We report a familial case of father-son duo affected by MMA. Whole exome sequencing identified genetic variations in SLIT1, RYR3 and ARPP21 involved in axon guidance, calcium homeostasis and regulation of calmodulin signaling respectively. This is the first attempt to define genetic modifiers associated with MMA from India and advocates to extend genetic screening to a larger cohort. Deciphering the functional consequences of variations in these genes will be crucial for unravelling the pathogenesis of MMA.

Neuromuscular Junction Dysfunction in Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal neurological disorder characterized by progressive degeneration of motor neurons leading to skeletal muscle denervation. Earlier studies have shown that motor neuron degeneration begins in motor cortex and descends to the neuromuscular junction (NMJ) in a dying forward fashion. However, accumulating evidences support that ALS is a distal axonopathy where early pathological changes occur at the NMJ, prior to onset of clinical symptoms and propagates towards the motor neuron cell body supporting "dying back" hypothesis. Despite several evidences, series of events triggering NMJ disassembly in ALS are still obscure. Neuromuscular junction is a specialized tripartite chemical synapse which involves a well-coordinated communication among the presynaptic motor neuron, postsynaptic skeletal muscle, and terminal Schwann cells. This review provides comprehensive insight into the role of NMJ in ALS pathogenesis. We have emphasized the molecular alterations in cellular components of NMJ leading to loss of effective neuromuscular transmission in ALS. Further, we provide a preview into research involved in exploring NMJ as potential target for designing effective therapies for ALS.

Burden of Stroke in India During 1960 to 2018: A Systematic Review and Meta-Analysis of Community Based Surveys.

BACKGROUND: Stroke is a heterogeneous disorder comprising of clinical subtypes and many risk factors, also alluded to as cerebrovascular disorders (CVDs). Increase in the global burden of stroke in developed and developing countries has been alarming. To galvanize the efforts towards the prevention and treatment, there is a need for robust data on the burden of stroke. OBJECTIVE: The aim of this study was to estimate the burden of stroke, a systematic review of community-based studies was conducted. MATERIAL AND METHODS: Systematic search of PubMed and Google Scholar for studies from January 1960 to December 2018 was done. The articles were screened and the data was retrieved and sorted into incidence, prevalence and mortality rates. Meta-analysis was done on Medcalc statistical software version 19.2.6. RESULTS: Prevalence rate of stroke for total population inclusive of urban and rural population, varied from 44.54 to 150/100000.For the urban population prevalence rate was 45 to 487/100000 and 55 to 388.4/100000 for rural population. The incidence rate varied from 33 to 123/100000 in the urban population and in the rural population it was estimated to be 123.57/100000. The 30 days case fatality rate of stroke varied from 41.08% to 42.06% in urban population and 18% to 46.3%.in the rural population. CONCLUSIONS: Systematic review and meta-analysis reveal that the stroke burden in India is quite high.

Familial Prion Disease: First Indian Kindred with Gerstmann-Sträussler-Scheinker Syndrome.

Gerstmann-Sträussler-Scheinker (GSS) syndrome is a devastating hereditary prion disease, presenting in 4th-5th decade with progressive ataxia and dementia. Pathogenic variants in the PRNP gene lead to aggregation of misfolded prion protein which results in neurodegeneration and death within a few years of onset. A key feature of prion disorders is conversion of normal prion protein (PrPc) into its misfolded form (PrPSc). Genetic modifiers include methionine at position 129 in prion protein and octapeptide repeats. We present an Indian kindred with c. 305C > T, p.Pro102Leu mutation in PRNP gene causing GSS in multiple members and discuss the impact of the polymorphism at position 129 on the severity of illness.

Sleep disorders in amyotrophic lateral sclerosis: A questionnaire-based study from India.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a relatively rare neurological disorder affecting upper and lower motor neurons in the brain and spinal cord with survival for 3-5 years and rarely beyond 10 years. Sleep disturbances in ALS are underreported and undertreated and there is no related data from India. This study aimed to assess the frequency of sleep disorders in patients of ALS and their determinants. METHODS: Patients with definite and probable ALS as per the El Escorial criteria were recruited from May 2014 to April 2016. Functional impairment, presence of sleep specific abnormalities and anxiety and depression were assessed using standardized questionnaires. RESULTS: Forty patients with ALS (23 male; 17 female) with their median age at presentation being 58.5 years (range 44-75 years) and the median duration of illness being 18 months (range: 4-120 months) were includedin the study. Half of the patients had poor sleep quality, which was significantly worse across all components of Pittsburgh Sleep Quality Index (PSQI) compared to controls. Sleep disorders were observed in 70%, insomnia in 65%, sleep disordered breathing/hypoventilation in 52.5% and restless legs syndrome in 5% patients. Night time awakenings attributable to symptoms associated with ALS were noted in 85%, and anxiety and depression in 57.5% patients. Excessive daytime somnolence emerged as an independent predictor for the presence of sleep disorders in ALS patients on multivariate logistic regression [P = 0.043, odd's ratio (OR) 1.435; 95% confidence interval[CI] (1.011-2.036)]. CONCLUSION: This is the first study from India providing insight into the presence of sleep disorders in ALS. About half of the patients of ALS had a poor sleep quality and two-thirds suffered from sleep disturbances.

Expression analysis of protein homeostasis pathways in the peripheral blood mononuclear cells of sporadic amyotrophic lateral sclerosis patients.

Misfolded protein aggregates are the hallmark of Amyotrophic Lateral Sclerosis (ALS) which suggests involvement of protein homeostasis pathways in etiology of ALS. However, status of protein homeostasis in peripheral blood of ALS is not well established. We analyzed expression levels of key genes of proteostasis pathways in peripheral blood mononuclear cells (PBMCs) of sporadic ALS (sALS) patients and healthy controls. Increased protein carbonylation was observed in patients reflecting oxidative damage in PBMCs. We observed increased transcript and protein levels of GRP78 suggesting Endoplasmic reticulum (ER) insult to cells. Further, significant upregulation of spliced XBP1 and two stress sensors: IRE1α/ERN1 and ATF6 indicated induction of unfolded protein response (UPR). Genes involved in autophagosome initiation (ULK1, ULK2, ATG13); nucleation and elongation (BECLIN1, ATG7, ATG16L1, ATG5, ATG10) and vesicular trafficking genes were significantly increased in patients. Increased lipidation of LC3 validated induction of autophagy. Accumulation of low molecular weight ubiquitinated proteins in patients suggested deregulation of proteasome (UPS) pathway. In addition, cytosolic chaperones (HSP70 and HSP27) and HSF1 were elevated in patients. Increased TDP43 indicated role of TDP43 in disease pathology. Our findings suggest that there is oxidative insult and upregulation of UPR, vesicular trafficking and autophagy in PBMCs of sALS patients.

An insight into death wish among patients with amyotrophic lateral sclerosis in India using "Wish-to-Die Questionnaire".

AIMS: In amyotrophic lateral sclerosis (ALS), death wish is expressed in a varying proportion of patients in different countries. In this first study from India, influence of belief system of religion/spirituality and attitude towards death, widely prevalent in the country, in decision making, was evaluated. MATERIAL AND METHODS: Twenty ALS patients were assessed using 'Wish-to-Die Questionnaire' (WDQ) developed to reflect seven domains, namely religion/spirituality, belief in karma, meaning of life, hope, family support, financial support and death wish. Functional impairment, depression, hopelessness and suicidal ideation were assessed by ALS Functional Rating Scale, Beck's Depression Inventory, Beck Hopelessness Scale and The Scale of Suicidal Ideation, respectively. RESULTS: On WDQ, all the 20 patients had belief in religion/spirituality, had hope and family support. Nineteen patients (95%) believed in karma, 16 (80%) still found life meaningful and 15 (75%) had financial support. Six patients (30%) had mild to moderate depression; hopelessness was present in 6 (30%) and suicidal ideation was present in one (5%). The 5 (25%) patients who expressed death wish did not significantly differ from others in 6 domains (religion/spirituality, belief in karma, meaning of life, hope, family support, financial support) of WDQ. The main reason in 3 patients who expressed death wish was lack of financial support. The fourth patient could not find meaning of life after the onset of illness, and the fifth wished to end his life since he had satisfactorily fulfilled all his responsibilities. CONCLUSION: Smaller proportion of patients of ALS expressed death wish in India compared to the Western countries. This may be attributed to belief in religion/spirituality and karma, having meaning of life and family support. As this is the first report from India, useful information may be obtained if similar studies are done on a larger sample.

Analysis of C9orf72 repeat expansion in amyotrophic lateral sclerosis patients from North India.

Pathogenic expansion of a hexanucleotide repeat in C9orf72 is associated with ~30% of familial ALS and ~7% of sporadic ALS patients amongst different populations. This repeat expansion was screened in 75 ALS patients and 115 healthy individuals from North India. On analysis by repeat-primed PCR, pathogenic expansion was not observed either in ALS patients or healthy controls. These observations are similar to the findings in most of the Asian populations.

Identification of L84F mutation with a novel nucleotide change c.255G > T in the superoxide dismutase gene in a North Indian family with amyotrophic lateral sclerosis.

Mutations in the superoxide dismutase (SOD1) gene account for ∼15% and in the transactive response DNA binding protein (TARDBP) gene for ∼5% of familial amyotrophic lateral sclerosis (FALS) cases. These two genes were analysed in two siblings from North India with ALS and a positive family history. The coding region of SOD1 and TARDBP genes was sequenced in both siblings. Genetic variation identified in SOD1 was typed in unaffected family members (n = 11), sporadic ALS patients (n = 48) and healthy controls (n = 35). Molecular dynamic (MD) simulations were performed on wild-type (WT) and mutant monomers of SOD1 to determine structural changes due to the identified mutation. A novel heterozygous nucleotide variation (c.255G > T) was identified in exon 4 of SOD1 in the two siblings and two asymptomatic family members but not in SALS patients and healthy controls. This variation results in a known non-synonymous substitution from leucine to phenylalanine at position 84 (L84F), making it a triallelic variation. Large conformational changes were observed in the zinc loop and electrostatic loop in an L84F mutant compared to WT SOD1 in MD simulations. In conclusion, this is the first report of mutation in SOD1 associated with FALS in India. Structural perturbations in L84F SOD1 may cause dimer destabilization, with decreased metal affinity leading to oligomerization.

Chin fasciculations in Madras motor neuron disease: a new clinical feature.

The characteristic features of Madras motor neuron disease (MMND) are onset in the young in the first two decades, sporadic occurrence, facial and bulbar paralysis, sensorineural hearing impairment, asymmetrical weakness of limbs and pyramidal signs with a slow progression. The majority of the cases reported are from South India. MMND variant has the additional features of optic atrophy and cerebellar signs. We are reporting a 48 year old female of MMND who had persistent fasciculations of chin, with electromyographic features of fasciculations and fibrillations in mentalis muscle. Chin fasciculations, a rare clinical feature, is now described for the first time in Madras motor neuron disease adding a new feature to the clinical constellation of symptoms.

Genetic association analysis of transporters identifies ABCC2 loci for seizure control in women with epilepsy on first-line antiepileptic drugs.

OBJECTIVE: The ATP-binding cassette (ABC) superfamily of transporters is known to efflux antiepileptic drugs (AEDs) primarily in the brain, gastrointestinal tract, liver, and kidneys. In addition, they are also known to be involved in estrogen disposition and may modulate seizure susceptibility and drug response. The objective of the present study was to investigate the role of genetic variants from ABC transporters in seizure control in epilepsy patients treated with monotherapy of first-line AEDs for 12 months. METHODS: On the basis of gene coverage and functional significance, a total of 98 single nucleotide polymorphisms from ABCB1, ABCC1, and ABCC2 were genotyped in 400 patients from North India. Of these, 216 patients were eligible for therapeutic assessment. Genetic variants were compared between the 'no-seizures' and the 'recurrent-seizures' groups. Bonferroni corrections for multiple comparisons and adjustment for covariates were performed before assessment of associations. RESULTS: Functionally relevant promoter polymorphisms from ABCC2: c.-1549G>A and c.-1019A>G either considered alone or in haplotype and diplotype combinations were observed for a significant association with seizure control in women (odds ratio>3.5, P<10, power>95%). Further, low protein-expressing CGT and TGT (c.-24C>T, c.1249G>A, c.3972C>T) haplotypes were always observed to be present in combination with the AG (c.-1549G>A, c.-1019A>G) haplotype that was over-represented in women with 'no seizures'. CONCLUSION: The distribution of the associated variants supports the involvement of ABCC2 in controlling seizures in women possibly by lowering of its expression. The biological basis of this finding could be an altered interaction of ABCC2 with AEDs and estrogens. These results necessitate replication in a larger pool of patients.

Genetic polymorphisms in sex hormone metabolizing genes and drug response in women with epilepsy.

AIMS: It is hypothesized that functionally relevant polymorphisms in genes encoding metabolizing enzymes of sex steroids may influence drug response by directly predisposing women with epilepsy to seizure exacerbation. An alteration in estradiol:progesterone ratio is believed to play a role in seizure occurrence in women. CYP1A1 is a key enzyme involved in the metabolism of estradiol, with variants of the CYP1A1 gene having been reported to play a role in the alteration of sex hormone metabolism in women. The objective of the present study was to test for the association of genetic variants in CYP1A1 with seizure recurrence in patients diagnosed with epilepsy. MATERIALS & METHODS: In the study, the association of five variants in CYP1A1 with seizure control in 228 patients with epilepsy on first-line antiepileptic drug therapy for a minimum period of 12 months was investigated. RESULTS: A significant association of an intronic SNP, IVS1 +606C>A (rs2606345), with respect to seizure recurrence (genotypic: p = 3.3 × 10(-4); allelic: p = 7.2 × 10(-4); OR: 2.86; 95% CI: 1.5-5.3) in women with epilepsy from North India was observed. CONCLUSION: Since CYP1A1 is not involved in the metabolism of any of the first-line antiepileptic drugs, these results imply that variants from genes encoding sex hormone metabolizing enzymes might act as markers for predicting response to antiepileptic drug therapy in women with epilepsy.

Genetic variability in estrogen disposition: Potential clinical implications for neuropsychiatric disorders.

Variability in the physiological levels of neuroactive estrogens is widely believed to play a role in predisposition to several disorders of the central nervous system. Local biosynthesis of estrogens in the brain as well as their circulating serum levels are known to contribute to this pool of neuroactive steroids. It has been well accepted that estrogens modulate neuronal functions by affecting genesis, differentiation, excitability, and degeneration of nerve cells. These actions of estrogens appear to be more prominent in females with higher concentrations and marked variability of circulating serum levels occurring over a woman's lifetime. However, our knowledge regarding the variability of neuroactive steroid levels is very limited. Furthermore, several studies have recently reported differences in the synchronization of circulating and neuronal levels of estradiol. In the absence of reliable circulating steroid levels, knowledge of genetic variability in estrogen disposition may play a determining factor in predicting altered susceptibility or severity of neuropsychiatric disorders in women. Over the past decade, several genetic variants have been linked to both differential serum estrogen levels and predisposition to diverse types of neuropsychiatric disorders in women. Polymorphisms in genes encoding estrogen-metabolizing enzymes as well as estrogen receptors may account for this phenotypic variability. In this review, we attempt to show the contribution of genetics in determining estrogenicity in females with a particular emphasis on the central nervous system. This knowledge will further provide a driving force for unearthing the novel field of "Estrogen Pharmacogenomics." © 2010 Wiley-Liss, Inc.

Genetic profile of patients with epilepsy on first-line antiepileptic drugs and potential directions for personalized treatment.

BACKGROUND: The first-line antiepileptic drugs, although affordable and effective in the control of seizures, are associated with adverse drug effects, and there is large interindividual variability in the appropriate dose at which patients respond favorably. This variability may partly be explained by functional consequences of genetic polymorphisms in the drug-metabolizing enzymes, such as the CYP450 family, microsomal epoxide hydrolase and UDP-glucuronosyltransferases, drug transporters, mainly ATP-binding cassette transporters, and drug targets, including sodium channels. The purpose of this study was to determine the allele and genotype frequencies of such genetic variants in patients with epilepsy from North India administered first-line antiepileptic drugs, such as phenobarbitone, phenytoin, carbamazepine and valproic acid, and compare them with worldwide epilepsy populations. MATERIALS & METHODS: SNP screening of 19 functional variants from 12 genes in 392 patients with epilepsy was carried out, and the patients were classified with respect to the metabolizing rate of their drug-metabolizing enzymes, efflux rate of drug transporters and sensitivity of drug targets. RESULTS: A total of 16 SNPs were found to be polymorphic, and the allelic frequencies for these SNPs were in conformance with Hardy-Weinberg equilibrium. Among all the polymorphisms studied, functional variants from genes encoding CYP2C19, EPHX1, ABCB1 and SCN1A were highly polymorphic in North Indian epilepsy patients, and might account for differential drug response to first-line antiepileptic drugs. CONCLUSION: Interethnic differences were elucidated for several polymorphisms that might be responsible for differential serum drug levels and optimal dose requirement for efficacious treatment.

Organization of neurology services in India: unmet needs and the way forward.

Recognition of the magnitude of the burden and disability and mortality consequent to neurological disorders has led to global initiatives of declaring them as a "global epidemic", emphasizing public health approach and integration of neurology care with general health care. Epidemiological transition with increase in neurological disorders in India, the gross mismatch between the need and the available trained manpower and infrastructural facilities are posing challenges to health planners and policy makers for providing neurological care to the community. Alternative approaches of optimal utilization of rural, community health, satellite clinic and district models of the health care with close interaction with tertiary centres, nongovernmental agencies and private sector may facilitate achieving the goal of taking neurology care to the 'unreached'.