Off the Clock: the Non-canonical Roles of Cyclin-Dependent Kinases in Neural and Glioma Stem Cell Self-Renewal.

Glioma stem cells (GSCs) are thought to drive growth and therapy resistance in glioblastoma (GBM) by "hijacking" at least a subset of signaling pathways active in normal neural stem cells (NSCs). Though the origins of GSCs still remain elusive, uncovering the mechanisms of self-renewing division and cell differentiation in normal NSCs has shed light on their dysfunction in GSCs. However, the distinction between self-renewing division pathways utilized by NSC and GSC becomes critical when considering options for therapeutically targeting signaling pathways that are specifically active or altered in GSCs. It is well-established that cyclin-dependent kinases (CDKs) regulate the cell cycle, yet more recent studies have shown that CDKs also play important roles in the regulation of neuronal survival, metabolism, differentiation, and self-renewal. The intimate relationship between cell cycle regulation and the cellular programs that determine self-renewing division versus cell differentiation is only beginning to be understood, yet seems to suggest potential differential vulnerabilities in GSCs. In this timely review, we focus on the role of CDKs in regulating the self-renewal properties of normal NSCs and GSCs, highlighting novel opportunities to therapeutically target self-renewing signaling pathways specifically in GBM.

TOP2B Enzymatic Activity on Promoters and Introns Modulates Multiple Oncogenes in Human Gliomas.

PURPOSE: The epigenetic mechanisms involved in transcriptional regulation leading to malignant phenotype in gliomas remains poorly understood. Topoisomerase IIB (TOP2B), an enzyme that decoils and releases torsional forces in DNA, is overexpressed in a subset of gliomas. Therefore, we investigated its role in epigenetic regulation in these tumors. EXPERIMENTAL DESIGN: To investigate the role of TOP2B in epigenetic regulation in gliomas, we performed paired chromatin immunoprecipitation sequencing for TOP2B and RNA-sequencing analysis of glioma cell lines with and without TOP2B inhibition and in human glioma specimens. These experiments were complemented with assay for transposase-accessible chromatin using sequencing, gene silencing, and mouse xenograft experiments to investigate the function of TOP2B and its role in glioma phenotypes. RESULTS: We discovered that TOP2B modulates transcription of multiple oncogenes in human gliomas. TOP2B regulated transcription only at sites where it was enzymatically active, but not at all native binding sites. In particular, TOP2B activity localized in enhancers, promoters, and introns of PDGFRA and MYC, facilitating their expression. TOP2B levels and genomic localization was associated with PDGFRA and MYC expression across glioma specimens, which was not seen in nontumoral human brain tissue. In vivo, TOP2B knockdown of human glioma intracranial implants prolonged survival and downregulated PDGFRA. CONCLUSIONS: Our results indicate that TOP2B activity exerts a pleiotropic role in transcriptional regulation of oncogenes in a subset of gliomas promoting a proliferative phenotype.

Methods for in vitro modeling of glioma invasion: Choosing tools to meet the need.

Widespread tumor cell invasion is a fundamental property of diffuse gliomas and is ultimately responsible for their poor prognosis. A greater understanding of basic mechanisms underlying glioma invasion is needed to provide insights into therapies that could potentially counteract them. While none of the currently available in vitro models can fully recapitulate the complex interactions of glioma cells within the brain tumor microenvironment, if chosen and developed appropriately, these models can provide controlled experimental settings to study molecular and cellular phenomena that are challenging or impossible to model in vivo. Therefore, selecting the most appropriate in vitro model, together with its inherent advantages and limitations, for specific hypotheses and experimental questions achieves primary significance. In this review, we describe and discuss commonly used methods for modeling and studying glioma invasion in vitro, including platforms, matrices, cell culture, and visualization techniques, so that choices for experimental approach are informed and optimal.

Surfen-mediated blockade of extratumoral chondroitin sulfate glycosaminoglycans inhibits glioblastoma invasion.

Invasive spread of glioblastoma (GBM) is linked to changes in chondroitin sulfate (CS) proteoglycan (CSPG)-associated sulfated glycosaminoglycans (GAGs) that are selectively up-regulated in the tumor microenvironment (TME). We hypothesized that inhibiting CS-GAG signaling in the TME would stem GBM invasion. Rat F98 GBM cells demonstrated enhanced preferential cell invasion into oversulfated 3-dimensional composite of CS-A and CS-E [4- and 4,6-sulfated CS-GAG (COMP)] matrices compared with monosulfated (4-sulfated) and unsulfated hyaluronic acid matrices in microfluidics-based choice assays, which is likely influenced by differential GAG receptor binding specificities. Both F98 and human patient-derived glioma stem cells (GSCs) demonstrated a high degree of colocalization of the GSC marker CD133 and CSPGs. The small molecule sulfated GAG antagonist bis-2-methyl-4-amino-quinolyl-6-carbamide (surfen) reduced invasion and focal adhesions in F98 cells encapsulated in COMP matrices and blocked CD133 and antichondroitin sulfate antibody (CS-56) detection of respective antigens in F98 cells and human GSCs. Surfen-treated F98 cells down-regulated CSPG-binding receptor transcripts and protein, as well as total and activated ERK and protein kinase B. Lastly, rats induced with frontal lobe tumors and treated with a single intratumoral dose of surfen demonstrated reduced tumor burden and spread compared with untreated controls. These results present a first demonstration of surfen as an inhibitor of sulfated GAG signaling to stem GBM invasion.-Logun, M. T., Wynens, K. E., Simchick, G., Zhao, W., Mao, L., Zhao, Q., Mukherjee, S., Brat, D. J., Karumbaiah, L. Surfen-mediated blockade of extratumoral chondroitin sulfate glycosaminoglycans inhibits glioblastoma invasion.

CDK5 Inhibition Resolves PKA/cAMP-Independent Activation of CREB1 Signaling in Glioma Stem Cells.

Cancer stem cells promote neoplastic growth, in part by deregulating asymmetric cell division and enhancing self-renewal. To uncover mechanisms and potential therapeutic targets in glioma stem cell (GSC) self-renewal, we performed a genetic suppressor screen for kinases to reverse the tumor phenotype of our Drosophila brain tumor model and identified dCdk5 as a critical regulator. CDK5, the human ortholog of dCdk5 (79% identity), is aberrantly activated in GBMs and tightly aligned with both chromosome 7 gains and stem cell markers affecting tumor-propagation. Our investigation revealed that pharmaceutical inhibition of CDK5 prevents GSC self-renewal in vitro and in xenografted tumors, at least partially by suppressing CREB1 activation independently of PKA/cAMP. Finally, our TCGA GBM data analysis revealed that CDK5, stem cell, and asymmetric cell division markers segregate within non-mesenchymal patient clusters, which may indicate preferential dependence on CDK5 signaling and sensitivity to its inhibition in this group.

Groundwater Arsenic Contamination in the Ganga River Basin: A Future Health Danger.

This study highlights the severity of arsenic contamination in the Ganga River basin (GRB), which encompasses significant geographic portions of India, Bangladesh, Nepal, and Tibet. The entire GRB experiences elevated levels of arsenic in the groundwater (up to 4730 µg/L), irrigation water (~1000 µg/L), and in food materials (up to 3947 µg/kg), all exceeding the World Health Organization's standards for drinking water, the United Nations Food and Agricultural Organization's standard for irrigation water (100 µg/L), and the Chinese Ministry of Health's standard for food in South Asia (0.15 mg/kg), respectively. Several individuals demonstrated dermal, neurological, reproductive, cognitive, and cancerous effects; many children have been diagnosed with a range of arsenicosis symptoms, and numerous arsenic-induced deaths of youthful victims are reported in the GRB. Victims of arsenic exposure face critical social challenges in the form of social isolation and hatred by their respective communities. Reluctance to establish arsenic standards and unsustainable arsenic mitigation programs have aggravated the arsenic calamity in the GRB and put millions of lives in danger. This alarming situation resembles a ticking time bomb. We feel that after 29 years of arsenic research in the GRB, we have seen the tip of the iceberg with respect to the actual magnitude of the catastrophe; thus, a reduced arsenic standard for drinking water, testing all available drinking water sources, and sustainable and cost-effective arsenic mitigation programs that include the participation of the people are urgently needed.

Molecular Programs Underlying Asymmetric Stem Cell Division and Their Disruption in Malignancy.

Asymmetric division of stem cells is a highly conserved and tightly regulated process by which a single stem cell produces two unequal daughter cells. One retains its stem cell identity while the other becomes specialized through a differentiation program and loses stem cell properties. Coordinating these events requires control over numerous intra- and extracellular biological processes and signaling networks. In the initial stages, critical events include the compartmentalization of fate determining proteins within the mother cell and their subsequent passage to the appropriate daughter cell in order to direct their destiny. Disturbance of these events results in an altered dynamic of self-renewing and differentiation within the cell population, which is highly relevant to the growth and progression of cancer. Other critical events include proper asymmetric spindle assembly, extrinsic regulation through micro-environmental cues, and non-canonical signaling networks that impact cell division and fate determination. In this review, we discuss mechanisms that maintain the delicate balance of asymmetric cell division in normal tissues and describe the current understanding how some of these mechanisms are deregulated in cancer.

TRIM8 regulates stemness in glioblastoma through PIAS3-STAT3.

Glioblastoma (GBM) is the most malignant form of primary brain tumor, and GBM stem-like cells (GSCs) contribute to the rapid growth, therapeutic resistance, and clinical recurrence of these fatal tumors. STAT3 signaling supports the maintenance and proliferation of GSCs, yet regulatory mechanisms are not completely understood. Here, we report that tri-partite motif-containing protein 8 (TRIM8) activates STAT3 signaling to maintain stemness and self-renewing capabilities of GSCs. TRIM8 (also known as 'glioblastoma-expressed ring finger protein') is expressed equally in GBM and normal brain tissues, despite its hemizygous deletion in the large majority of GBMs, and its expression is highly correlated with stem cell markers. Experimental knockdown of TRIM8 reduced GSC self-renewal and expression of SOX2, NESTIN, and p-STAT3, and promoted glial differentiation. Overexpression of TRIM8 led to higher expression of p-STAT3, c-MYC, SOX2, NESTIN, and CD133, and enhanced GSC self-renewal. We found that TRIM8 activates STAT3 by suppressing the expression of PIAS3, an inhibitor of STAT3, most likely through E3-mediated ubiquitination and proteasomal degradation. Interestingly, we also found that STAT3 activation upregulates TRIM8, providing a mechanism for normalized TRIM8 expression in the setting of hemizygous gene deletion. These data demonstrate that bidirectional TRIM8-STAT3 signaling regulates stemness in GSC.

Fate of over 480 million inhabitants living in arsenic and fluoride endemic Indian districts: Magnitude, health, socio-economic effects and mitigation approaches.

During our last 27 years of field survey in India, we have studied the magnitude of groundwater arsenic and fluoride contamination and its resulting health effects from numerous states. India is the worst groundwater fluoride and arsenic affected country in the world. Fluoride results the most prevalent groundwater related diseases in India. Out of a total 29 states in India, groundwater of 20 states is fluoride affected. Total population of fluoride endemic 201 districts of India is 411 million (40% of Indian population) and more than 66 million people are estimated to be suffering from fluorosis including 6 million children below 14 years of age. Fluoride may cause a crippling disease. In 6 states of the Ganga-Brahmaputra Plain (GB-Plain), 70.4 million people are potentially at risk from groundwater arsenic toxicity. Three additional states in the non GB-Plain are mildly arsenic affected. For arsenic with substantial cumulative exposure can aggravate the risk of cancers along with various other diseases. Clinical effects of fluoride includes abnormal tooth enamel in children; adults had joint pain and deformity of the limbs, spine etc. The affected population chronically exposed to arsenic and fluoride from groundwater is in danger and there is no available medicine for those suffering from the toxicity. Arsenic and fluoride safe water and nutritious food are suggested to prevent further aggravation of toxicity. The World Health Organization (WHO) points out that social problems arising from arsenic and fluoride toxicity eventually create pressure on the economy of the affected areas. In arsenic and fluoride affected areas in India, crisis is not always having too little safe water to satisfy our need, it is the crisis of managing the water.

Drosophila Brat and Human Ortholog TRIM3 Maintain Stem Cell Equilibrium and Suppress Brain Tumorigenesis by Attenuating Notch Nuclear Transport.

Cancer stem cells exert enormous influence on neoplastic behavior, in part by governing asymmetric cell division and the balance between self-renewal and multipotent differentiation. Growth is favored by deregulated stem cell division, which enhances the self-renewing population and diminishes the differentiation program. Mutation of a single gene in Drosophila, Brain Tumor (Brat), leads to disrupted asymmetric cell division resulting in dramatic neoplastic proliferation of neuroblasts and massive larval brain overgrowth. To uncover the mechanisms relevant to deregulated cell division in human glioma stem cells, we first developed a novel adult Drosophila brain tumor model using brat-RNAi driven by the neuroblast-specific promoter inscuteable Suppressing Brat in this population led to the accumulation of actively proliferating neuroblasts and a lethal brain tumor phenotype. brat-RNAi caused upregulation of Notch signaling, a node critical for self-renewal, by increasing protein expression and enhancing nuclear transport of Notch intracellular domain (NICD). In human glioblastoma, we demonstrated that the human ortholog of Drosophila Brat, tripartite motif-containing protein 3 (TRIM3), similarly suppressed NOTCH1 signaling and markedly attenuated the stem cell component. We also found that TRIM3 suppressed nuclear transport of active NOTCH1 (NICD) in glioblastoma and demonstrated that these effects are mediated by direct binding of TRIM3 to the Importin complex. Together, our results support a novel role for Brat/TRIM3 in maintaining stem cell equilibrium and suppressing tumor growth by regulating NICD nuclear transport. Cancer Res; 76(8); 2443-52. ©2016 AACR.

Cancer stem cell division: when the rules of asymmetry are broken.

Asymmetric division of stem cells is a highly conserved and tightly regulated process by which a single stem cell produces two daughter cells and simultaneously directs the differential fate of both: one retains its stem cell identity while the other becomes specialized and loses stem cell properties. Coordinating these events requires control over numerous intra- and extracellular biological processes and signaling networks. In the initial stages, critical events include the compartmentalization of fate determining proteins within the mother cell and their subsequent passage to the appropriate daughter cell. Disturbance of these events results in an altered dynamic of self-renewing and differentiation within the cell population, which is highly relevant to the growth and progression of cancer. Other critical events include proper asymmetric spindle assembly, extrinsic regulation through micro-environmental cues, and noncanonical signaling networks that impact cell division and fate determination. In this review, we discuss mechanisms that maintain the delicate balance of asymmetric cell division in normal tissues and describe the current understanding how some of these mechanisms are deregulated in cancer. The universe is asymmetric and I am persuaded that life, as it is known to us, is a direct result of the asymmetry of the universe or of its indirect consequences. The universe is asymmetric. -Louis Pasteur.

Emerging functional similarities and divergences between Drosophila Spargel/dPGC-1 and mammalian PGC-1 protein.

Peroxisome Proliferator Activated Receptor Gamma Co-activator-1 (PGC-1) is a well-conserved protein among all chordates. Entire Drosophila species subgroup carries a PGC-1 homolog in their genome called spargel/dPGC-1 showing very little divergence. Recent studies have reported that significant functional similarities are shared between vertebrate and invertebrate PGC-1's based on their role in mitochondrial functions and biogenesis, gluconeogenesis, and most likely in transcription and RNA processing. With the help of genetic epistasis analysis, we established that Drosophila Spargel/dPGC-1 affects cell growth process as a terminal effector in the Insulin-TOR signaling pathway. The association between Spargel/dPGC-1 and Insulin signaling could also explain its role in the aging process. Here we provided a further comparison between Spargel/dPGC-1 and PGC-1 focusing on nuclear localization, oxidative stress resistance, and a possible role of Spargel/dPGC-1 in oogenesis reminiscing the role of Spargel in reproductive aging like many Insulin signaling partners. This led us to hypothesize that the discovery of newer biological functions in Drosophila Spargel/dPGC-1 will pave the way to uncover novel functional equivalents in mammals.

Spargel/dPGC-1 is a new downstream effector in the insulin-TOR signaling pathway in Drosophila.

Insulin and target of rapamycin (TOR) signaling pathways converge to maintain growth so a proportionate body form is attained. Insufficiency in either insulin or TOR results in developmental growth defects due to low ATP level. Spargel is the Drosophila homolog of PGC-1, which is an omnipotent transcriptional coactivator in mammals. Like its mammalian counterpart, Spargel/dPGC-1 is recognized for its role in energy metabolism through mitochondrial biogenesis. An earlier study demonstrated that Spargel/dPGC-1 is involved in the insulin-TOR signaling, but a comprehensive analysis is needed to understand exactly which step of this pathway Spargel/PGC-1 is essential. Using genetic epistasis analysis, we demonstrated that a Spargel gain of function can overcome the TOR and S6K mediated cell size and cell growth defects in a cell autonomous manner. Moreover, the tissue-restricted phenotypes of TOR and S6k mutants are rescued by Spargel overexpression. We have further elucidated that Spargel gain of function sets back the mitochondrial numbers in growth-limited TOR mutant cell clones, which suggests a possible mechanism for Spargel action on cells and tissue to attain normal size. Finally, excess Spargel can ameliorate the negative effect of FoxO overexpression only to a limited extent, which suggests that Spargel does not share all of the FoxO functions and consequently cannot significantly rescue the FoxO phenotypes. Together, our observation established that Spargel/dPGC-1 is indeed a terminal effector in the insulin-TOR pathway operating below TOR, S6K, Tsc, and FoxO. This led us to conclude that Spargel should be incorporated as a new member of this growth-signaling pathway.

A muscle-specific p38 MAPK/Mef2/MnSOD pathway regulates stress, motor function, and life span in Drosophila.

Molecular mechanisms that concordantly regulate stress, life span, and aging remain incompletely understood. Here, we demonstrate that in Drosophila, a p38 MAP kinase (p38K)/Mef2/MnSOD pathway is a coregulator of stress and life span. Hence, overexpression of p38K extends life span in a MnSOD-dependent manner, whereas inhibition of p38K causes early lethality and precipitates age-related motor dysfunction and stress sensitivity, that is rescued through muscle-restricted (but not neuronal) add-back of p38K. Additionally, mutations in p38K are associated with increased protein carbonylation and Nrf2-dependent transcription, while adversely affecting metabolic response to hypoxia. Mechanistically, p38K modulates expression of the mitochondrial MnSOD enzyme through the transcription factor Mef2, and predictably, perturbations in MnSOD modify p38K-dependent phenotypes. Thus, our results uncover a muscle-restricted p38K-Mef2-MnSOD signaling module that influences life span and stress, distinct from the insulin/JNK/FOXO pathway. We propose that potentiating p38K might be instrumental in restoring the mitochondrial detoxification machinery and combating stress-induced aging.

SOD2, the principal scavenger of mitochondrial superoxide, is dispensable for embryogenesis and imaginal tissue development but essential for adult survival.

Definitive evidence on the impact of MnSOD/SOD2-deficiency and the consequent effects of high flux of mitochondrial reactive oxygen species (ROS) on pre-natal/pre-adult development has yet to be reported for either Drosophila or mice. Here we report that oocytes lacking maternal SOD2 protein develop into adults just like normal SOD2-containing oocytes suggesting that maternal SOD2-mediated protection against mitochondrial ROS is not essential for oocyte viability. However, the capacity of SOD2-null larvae to undergo successful metamorphosis into adults is negatively influenced in the absence of SOD2. We therefore determined the impact of a high superoxide environment on cell size, progression through the cell cycle, cell differentiation, and cell death and found no difference between SOD2-null and SOD2+ larva and pupa. Thus loss of SOD2 activity clearly has no effect on pre-adult imaginal tissues. Instead, we found that the high mitochondrial superoxide environment arising from the absence of SOD2 leads to the induction of autophagy. Such autophagic response may underpin the resistance of pre-adult tissues to unscavenged ROS. Finally, while our data establish that SOD2 activity is less essential for normal development, the mortality of Sod2-/- neonates of both Drosophila and mice suggests that SOD2 activity is indeed essential for the viability of adults. We therefore asked if the early mortality of SOD2-null young adults could be rescued by activation of SOD2 expression. The results support the conclusion that the early mortality of SOD2-null adults is largely attributable to the absence of SOD2 activity in the adult per se. This finding somewhat contradicts the widely held notion that failure to scavenge the high volume of superoxide emanating from the oxidative demands of development would be highly detrimental to developing tissues.

Status of groundwater arsenic contamination in Bangladesh: a 14-year study report.

Since 1996, 52,202 water samples from hand tubewells were analyzed for arsenic (As) by flow injection hydride generation atomic absorption spectrometry (FI-HG-AAS) from all 64 districts of Bangladesh; 27.2% and 42.1% of the tubewells had As above 50 and 10 µg/l, respectively; 7.5% contained As above 300 µg/l, the concentration predicting overt arsenical skin lesions. The groundwater of 50 districts contained As above the Bangladesh standard for As in drinking water (50 µg/l), and 59 districts had As above the WHO guideline value (10 µg/l). Water analyses from the four principal geomorphological regions of Bangladesh showed that hand tubewells of the Tableland and Hill tract regions are primarily free from As contamination, while the Flood plain and Deltaic region, including the Coastal region, are highly As-contaminated. Arsenic concentration was usually observed to decrease with increasing tubewell depth; however, 16% of tubewells deeper than 100 m, which is often considered to be a safe depth, contained As above 50 µg/l. In tubewells deeper than 350 m, As >50 µg/l has not been found. The estimated number of tubewells in 50 As-affected districts was 4.3 million. Based on the analysis of 52,202 hand tubewell water samples during the last 14 years, we estimate that around 36 million and 22 million people could be drinking As-contaminated water above 10 and 50 µg/l, respectively. However for roughly the last 5 years due to mitigation efforts by the government, non-governmental organizations and international aid agencies, many individuals living in these contaminated areas have been drinking As-safe water. From 50 contaminated districts with tubewell As concentrations >50 µg/l, 52% of sampled hand tubewells contained As <10 µg/l, and these tubewells could be utilized immediately as a source of safe water in these affected regions provided regular monitoring for temporal variation in As concentration. Even in the As-affected Flood plain, sampled tubewells from 22 thanas in 4 districts were almost entirely As-safe. In Bangladesh and West Bengal, India the crisis is not having too little water to satisfy our needs, it is the challenge of managing available water resources. The development of community-specific safe water sources coupled with local participation and education are required to slow the current effects of widespread As poisoning and to prevent this disaster from continuing to plague individuals in the future.

Arsenic groundwater contamination and its health effects in the state of Uttar Pradesh (UP) in upper and middle Ganga plain, India: a severe danger.

This communication presents results of our 2-year survey on groundwater arsenic contamination in three districts Ballia, Varanasi and Gazipur of Uttar Pradesh (UP) in the upper and middle Ganga plain, India. Analyses of 4,780 tubewell water samples revealed that arsenic concentrations in 46.5% exceeded 10 microg/L, in 26.7%, 50 microg/L and in 10% 300 microg/L limits. Arsenic concentrations up to 3,192 microg//L were observed. The age of tubewells (n=1,881) ranged from less than a year to 32 years, with an average of 6.5 years. Our study shows that older tubewells had a greater chance of contamination. Depth of tubewells (n=3,810) varied from 6 to 60.5 m with a mean of 25.75 m. A detailed study in three administrative units within Ballia district, i.e. block, Gram Panchayet, and village was carried out to assess the magnitude of the contamination. Before our survey the affected villagers were not aware that they were suffering from arsenical toxicity through contaminated drinking water. A preliminary clinical examination in 11 affected villages (10 from Ballia and 1 from Gazipur district) revealed typical arsenical skin lesions ranging from melanosis, keratosis to Bowens (suspected). Out of 989 villagers (691 adults, and 298 children) screened, 137 (19.8%) of the adults and 17 (5.7%) of the children were diagnosed to have typical arsenical skin lesions. Arsenical neuropathy and adverse obstetric outcome were also observed, indicating severity of exposure. The range of arsenic concentrations in hair, nail and urine was 137-10,900, 764-19,700 microg/kg, and 23-4,030 microg/L, respectively. The urine, hair and nail concentrations of arsenic correlated significantly (r=0.76, 0.61, and 0.55, respectively) with drinking water arsenic concentrations. The similarity to previous studies on arsenic contamination in West Bengal, Bihar and Bangladesh indicates that people from a significant part of the surveyed areas in UP are suffering and this will spread unless drives to raise awareness of arsenic toxicity are undertaken and an arsenic safe water supply is immediately introduced.

Molecular pathogenesis of Parkinson's disease: identification of mutations in the Parkin gene in Indian patients.

Parkinson's disease (PD), the second most common neurodegenerative disorder, affects at least 1% of the population over the age of 50. However, very little information is available regarding the molecular basis of PD among Indians. Since the largest number of mutations have been detected in the Parkin gene among all known PD loci, we aim to use Parkin as the candidate gene to assess its role in PD-related pathogenesis in Indian patients. A total of 138 PD patients, with the mean age of onset being 47+/-14 (age range, 5-77 years), and 100 controls were recruited for the study from eastern India. Parkin mutations were detected by amplification of exons of the gene along with the flanking splice junctions by polymerase chain reaction, single-stranded conformation polymorphism and DNA sequencing. A total of 18 nucleotide variants including six novel changes were detected. These include five missense mutations (Gln34Arg, Arg42Cys, Arg42His, Tyr143Cys and Arg334Cys) detected in eight patients in heterozygous condition and a homozygous deletion encompassing exons 3 and 4 in two sibs affected with PD. Clinical features of the Parkin mutants were compared. Among eastern Indian PD patients, mutation in Parkin was identified in 7.24% cases.

An eight-year study report on arsenic contamination in groundwater and health effects in Eruani village, Bangladesh and an approach for its mitigation.

Based on several surveys during 1997-2005 and visits of a medical team to Eruani village, Laksham upazila, Comilla district, Bangladesh, the arsenic contamination situation and consequent clinical manifestations of arsenicosis among the villagers, including dermatology, neuropathy, and obstetric outcome, are reported here. Analysis of biological samples from patients and non-patients showed high body burden of arsenic. Even after eight years of known exposure, village children were still drinking arsenic-contaminated water, and many of them had arsenical skin lesions. There were social problems due to the symptoms of arsenicosis. The last survey established that there is a lack of proper awareness among villagers about different aspects of arsenic toxicity. The viability of different options of safe water, such as dugwells, deep tubewells, rainwater harvesting, and surface water with watershed management in the village, was studied. Finally, based on 19 years of field experience, it was felt that, for any successful mitigation programme, emphasis should be given to creating awareness among villagers about the arsenic problem, role of arsenic-free water, better nutrition from local fruits and vegetables, and, above all, active participation of women along with others in the struggle against the arsenic menace.