Compound heterozygous variants in Wiskott-Aldrich syndrome like (WASL) gene segregating in a family with early onset Parkinson's disease.

BACKGROUND: Knowledge of genetic determinants in Parkinson's disease is still limited. Familial forms of the disease continue to provide a rich resource to capture the genetic spectrum in disease pathogenesis, and this approach is exploited in this study. METHODS: Informative members from a three-generation family of Indian ethnicity manifesting a likely autosomal recessive mode of inheritance of Parkinson's disease were used for whole exome sequencing. Variant data analysis and in vitro functional characterisation of variant(s) segregating with the phenotype were carried out in HEK-293 and SH-SY5Y cells using gene constructs of interest. RESULTS: Two compound heterozygous variants, a rare missense (c.1139C > T:p.P380L) and a novel splice variant (c.1456 + 2 delTAGA, intron10) in Wiskott-Aldrich syndrome like gene (WASL, 7q31), both predicted to be deleterious were shared among the proband and two affected siblings. WASL, a gene not previously linked to a human Mendelian disorder is known to regulate actin polymerisation via Arp2/3 complex. Based on exon trapping assay using pSPL3 vector in HEK-293 cells, the splice variant showed skipping of exon10. Characterisation of the missense variant in SH-SY5Y cells demonstrated: i) significant alterations in neurite length and number; ii) decreased reactive oxygen species tolerance in mutation carrying cells on Tetrabutylphosphonium hydroxide induction and iii) increase in alpha-synuclein protein. Screening for WASL variants in two independent PD cohorts identified four individuals with heterozygous but none with biallelic variants. CONCLUSION: WASL, with demonstrated functional relevance in neurons may be yet another strong candidate gene for autosomal recessive PD encouraging assessment of its contribution across populations.

Novel and reported variants in Parkinson's disease genes confer high disease burden among Indians.

BACKGROUND: Genetic heterogeneity in Parkinson's disease (PD) has been unambiguously reported across different populations. Assuming a higher genetic load, we tested variant burden in PD genes to an early onset PD cohort from India. METHODS: Whole exome sequencing was performed in 250 PD patients recruited following MDS-UPDRS criteria. The number of rare variants in the 20 known PD genes per exome were used to calculate average rare variant burden with the 616 non-PD exomes available in-house as a comparison group. SKAT-O test was used for gene level analysis. RESULTS: 80 patients harboured rare variants in 20 PD genes, of which six had known pathogenic variants accounting for 2.4% of the cohort. Of 80 patients, 12 had homozygous and nine had likely compound heterozygous variants in recessive PD genes and 59 had heterozygous variants in only dominant PD genes. Of the 16 novel variants of as yet unknown significance identified, four homozygous across ATP13A2, PRKN, SYNJ1 and PARK7; and 12 heterozygous among LRRK2, VPS35, EIF4G1 and CHCHD2 were observed. SKAT-O test suggested a higher burden in GBA (punadjusted = 0.002). Aggregate rare variant analysis including 75 more individuals with only heterozygous variants in recessive PD genes (excluding GBA), with an average of 0.85 protein-altering rare variants per PD patient exome versus 0.51 in the non-PD group, revealed a significant enrichment (p < 0.0001). CONCLUSION: This first study in an early onset PD cohort among Indians identified 16 novel variants in known genes and also provides evidence for a high genetic burden in this ethnically distinct population.

Role of Physical Activity in Parkinson's Disease.

Parkinson's disease (PD) is common, age-dependent neurodegenerative disorder caused by a severe loss of the nigrostriatal dopaminergic neurons. Given the projected increase in the number of people with PD over the coming decades, interventions aimed at minimizing morbidity and improve quality of life are crucial. There is currently no fully proven pharmacological therapy that can modify or slow the disease progression. Physical activity (PA) can complement pharmacological therapy to manage the inherent decline associated with the disease. The evidence indicates that upregulation of neurotrophins and nerve growth factors are potentially critical mediators of the beneficial effects associated with PA. Accumulating evidence suggests that patients with PD might benefit from PA in a number of ways, from general improvements in health to disease-specific effects and potentially, disease-modifying effects. Various forms of PA that have shown beneficial effects in PD include - aerobic exercises, treadmill training, dancing, traditional Chinese exercise, yoga, and resistance training. In this review, we explored available research that addresses the impact of exercise and PA on PD. The original articles with randomized control trials, prospective cohort studies, longitudinal studies, meta-analysis, and relevant review articles from 2005 to 2017 were selected for the present review. Many gaps remain in our understanding of the most effective exercise intervention for PD symptoms, the mechanisms underlying exercise-induced changes and the best way to monitor response to therapy. However, available research suggests that exercise is a promising, cost-effective, and low-risk intervention to improve both motor and nonmotor symptoms in patients with PD. Thus, PA should be prescribed and encouraged in all PD patients.

Epidemiology of Parkinson's Disease-East Versus West.

BACKGROUND: The cause of PD at present remains unknown. A number of epidemiological studies have been conducted across the globe to ascertain the disease burden and the possible risk factors. In this review, we analyze the various studies from East and West with an aim to observe the important similarities and differences in the disease occurrence and risk factor profile. METHODS: A comprehensive search of descriptive and analytical epidemiological studies was undertaken. The descriptive studies and meta-analysis providing the standardised population rates were selected. The demographics, ethnicity and geographical differences between East and West were analysed. In analytical epidemiology, more established and well-studied non-genetic risk factors for PD were reviewed utilising the prospective cohort studies, case control studies and meta-analysis where available. RESULTS AND CONCLUSION: PD is more common with increasing age and shows male predominance, which is more obvious in Western studies. The PD prevalence and incidence rates are slightly lower in the East compared to the West. Incidence studies on different ethnic populations in the same country have also found a lower occurrence of PD amongst Easterners compared to Westerners. Setting methodological differences aside, studies from East and West suggest a role for both environmental and genetic risk factors in PD causation. Smoking, caffeine intake and pesticide exposure are well-established risk factors across regions. There is a robust data for dairy product consumption, urate levels and physical activity in the West while studies on certain risk factors like head injury and alcohol show conflicting and mixed results.

Suicide in Parkinson's Disease.

BACKGROUND: Suicide is a potentially preventable event. Suicidal ideation is common in Parkinson's disease (PD), but literature on completed suicides is scarce. In this case-control study, we compared the clinical characteristics of PD subjects who completed suicide (case) with those who died from natural causes (control). METHODS: PD patients from the National Neurosciences Institute's movement disorders database from 2002 till 2012 were identified. The database was linked to the Singapore National Registry of Disease Office for mortality information, and suicide deaths were confirmed with the coroner's office. The demographic and clinical variables were compared between the cases and controls and the significant factors were further analyzed using logistic regression analysis. RESULTS: During the study period, 366 deaths were recorded and suicide accounted for 11 deaths. Ten subjects with suicide deaths with complete clinical information were compared with randomly selected 30 PD subjects who had died from natural causes. PD suicide patients were younger (65.9 vs. 74.48 years), had less comorbidities (CWI: 2.6 vs. 4.63), better cognition (MMSE: 25.75 vs. 21.36), lower 'ON' UPDRS motor scores (20.83 vs. 41.63), lower H &Y stage (2.16 vs. 3.86), and higher use of Entacapone than the PD non-suicide group. CONCLUSION: Suicide is potentially preventable tragedy. PD patients with the identified clinical characteristics should be closely monitored for suicide ideations. Motor fluctuation is a treatable factor in such patients and should be aggressively managed.

Early Onset Parkinson's disease due to DJ1 mutations: An Indian study.

INTRODUCTION: Early Onset Parkinson's Disease (EOPD) is genetically heterogeneous. PARK2 mutations are the commonest cause of autosomal recessive EOPD followed by PINK1.DJ1 mutations is rare and there is scarce literature on its phenotype and long term outcome. OBJECTIVES: We undertook a retrospective study to determine the prevalence of DJ1 mutation(s) in an Indian population and describe the clinical features and long term outcome of EOPD patients with these mutations. METHODS: One hundred EOPD patients and 114 controls were evaluated. All the seven coding exons of DJ1 gene were screened for novel and reported mutations by PCR- Sanger sequencing. RESULTS: A novel homozygous missense mutation (c.313 A > T, p. Ile105Phe) in exon 5 was seen in one patient and four unrelated patients had a homozygous missense single nucleotide variant rs71653619 (c.293 G > A, p.Arg98Gln). The clinical phenotype comprised of asymmetrical onset, slowly progressive Parkinsonism with levodopa induced motor restlessness in a patient with the novel mutation (c.313 A > T, p. Ile105Phe) while subjects with c.293 G > A, p.Arg98Gln had early onset levodopa responsive symmetrical Parkinsonism. CONCLUSION: DJ1 mutations account for ∼5% of EOPD patients from the Indian population. This study further adds to the clinical spectrum of EOPD with DJ1 mutations.

Hydro-dissection - A simple Solution in Difficult Laparoscopic Cholecystectomy.

This Quasi-experimental study was done to assess the effectiveness of hydro-dissection in difficult laparoscopic cholecystectomies in Hamdard University Hospital, Karachi, Pakistan, from April 2012 to March 2014. All consecutive patients who presented with cholelithiasis and planned for laparoscopic cholecystectomy were enrolled in this study. Per-operatively the degree of difficulty of the operation was assessed by Cuschieri's scale after grading; Grade II, III and IV cholecystectomies were included in this study. Hydro dissection with saline jet through 5mm simple irrigation and suction probe was used, Operative findings and the total number of patients, in whom anatomy of calot's triangle was clearly displayed with hydro-dissection, was recorded. A total 55 patients were included in the study after assessing the degree of difficulty per operatively by Cuschieri Scale. Thirty one (31) patients were in Group II, 22 in Group III and 02 were included in group IV of Cuschieri scale in which hydro-dissection was used. This method cleared the obscure anatomy in all patients in Group II but in 3 patients of Group III, dense adhesions required sharp dissection to clear the operative field. Two patients, in whom conversion was required, were grouped in Cuschieri's scale IV. Methods of dissection in difficult cholecystectomies are of paramount importance to avoid iatrogenic injuries. Hydro-dissection using suction irrigation probe is a safe and effective technique to clear the difficult anatomy.

A semisynthetic fetal calf serum-free liquid medium for in vitro cultivation of Leishmania promastigotes.

A semisynthetic, autoclavable, liquid medium that does not require inclusion of fetal calf serum (FCS) for the successful cultivation of Leishmania promastigotes in vitro is described. The parasites were grown in commercially available medium 199 supplemented with 10% FCS and 2% mammalian urine and the growth was compared with the same medium without FCS. The growth of the parasites in the semidefined medium without FCS was found to be as good as or slightly better that with medium 199 supplemented with FCS.

Determination of phosphatidylcholine in a flow injection system using immobilized enzyme reactors.

Two alternative procedures are described for the quantitative determination of phosphatidylcholine in a flow-injection system utilizing immobilized enzymes. Phospholipase C from Bacillus cereus and phospholipase D from cabbage were covalently bound to the surface of controlled-pore glass beads and the enzyme-derivatized beads were packed in small columns. In the first procedure, the phospholipase C column was connected with a second column containing coimmobilized alkaline phosphatase and choline oxidase. In the alternative procedure, the column packed with immobilized phospholipase D was connected with a column packed with immobilized choline oxidase. The hydrogen peroxide produced through the action of choline oxidase in both flow-injection systems was detected amperometrically. Both procedures are suitable for an accurate and rapid quantitation of phosphatidylcholine. The sensitivity of the method based on phospholipase C and alkaline phosphatase is higher than that using phospholipase D. Quantitation of phosphatidylcholine at the nanomole level can be easily obtained using the first method.

An improved procedure for the purification of ethanolaminephosphotransferase. Reconstitution of the purified enzyme with lipids.

Ethanolaminephosphotransferase (CDPethanolamine:1,2-diacylglycerol ethanolaminephosphotransferase, EC 2.7.8.1) has been purified in active form from rat brain microsomes by a two-step chromatographic procedure. Enzyme preparations characterized by high specific activity and stability were obtained supplementing the solubilization and elution buffers, containing 1% Triton X-100, with 0.01% 2,6-di-tert-butyl-4-methylphenol. The specific activity of the purified enzyme was about 1200-times higher than that of the crude solubilized enzyme. The lipid dependence of ethanolaminephosphotransferase was studied both in the presence of Triton X-100 and in detergent-free enzyme preparations. The activity of the detergent-solubilized ethanolaminephosphotransferase was strongly modified by phospholipids. The kinetic behaviour of the enzyme was also dependent on the lipids contained in the aggregates obtained by removal of the detergent from detergent/lipid/protein suspensions. A regulatory role of phospholipids on the activity of the membrane-bound ethanolaminephosphotransferase is discussed.