Correction: Aktar et al. Gene Expression Analysis of Immune Regulatory Genes in Circulating Tumour Cells and Peripheral Blood Mononuclear Cells in Patients with Colorectal Carcinoma. Int. J. Mol. Sci. 2023, 24, 5051.

Faysal Bin Hamid was not included as an author in the original publication [...].

Gene Expression Analysis of Immune Regulatory Genes in Circulating Tumour Cells and Peripheral Blood Mononuclear Cells in Patients with Colorectal Carcinoma.

Information regarding genetic alterations of driver cancer genes in circulating tumour cells (CTCs) and their surrounding immune microenvironment nowadays can be employed as a real-time monitoring platform for translational applications such as patient response to therapeutic targets, including immunotherapy. This study aimed to investigate the expression profiling of these genes along with immunotherapeutic target molecules in CTCs and peripheral blood mononuclear cells (PBMCs) in patients with colorectal carcinoma (CRC). Expression of p53, APC, KRAS, c-Myc, and immunotherapeutic target molecules PD-L1, CTLA-4, and CD47 in CTCs and PBMCs were analysed by qPCR. Their expression in high versus low CTC-positive patients with CRC was compared and clinicopathological correlations between these patient groups were analysed. CTCs were detected in 61% (38 of 62) of patients with CRC. The presence of higher numbers of CTCs was significantly correlated with advanced cancer stages (p = 0.045) and the subtypes of adenocarcinoma (conventional vs. mucinous, p = 0.019), while being weakly correlated with tumour size (p = 0.051). Patients with lower numbers of CTCs had higher expression of KRAS. Higher KRAS expression in CTCs was negatively correlated with tumour perforation (p = 0.029), lymph node status (p = 0.037), distant metastasis (p = 0.046) and overall staging (p = 0.004). CTLA-4 was highly expressed in both CTCs and PBMCs. In addition, CTLA-4 expression was positively correlated with KRAS (r = 0.6878, p = 0.002) in the enriched CTC fraction. Dysregulation of KRAS in CTCs might evade the immune system by altering the expression of CTLA-4, providing new insights into the selection of therapeutic targets at the onset of the disease. Monitoring CTCs counts, as well as gene expression profiling of PBMCs, can be helpful in predicting tumour progression, patient outcome and treatment.

Identification of ZBTB24 protein domains and motifs for heterochromatin localization and transcriptional activation.

Immunodeficiency, centromeric instability, facial anomalies (ICF) syndrome is a rare autosomal recessive disorder caused by mutations in either DNMT3B, ZBTB24, CDCA7, HELLS or an unknown gene(s). Among the known causative genes, ZBTB24 encodes a member of the BTB-zinc finger (ZF) transcription factor family. The protein possesses a BTB domain, an AT-hook and eight C2H2 ZF motifs. All ZBTB24 mutations reported in ICF patients are predicted to disrupt at least one ZF motif. Here, we show that both AT-hook and distinct ZF motifs, particularly the 6th motif, of human and mouse ZBTB24 proteins are important for their heterochromatin localization. On the other hand, the 6th and 7th ZF motifs, and not the AT-hook or the BTB domain, of the human and mouse proteins are essential for transcriptional activation of CDCA7, another ICF causative gene and a known target of ZBTB24. By deletion analysis of the human CDCA7 promoter, we show that two motifs for ZBTB24 binding are important for transcriptional activation of this gene. These results reveal the evolutionarily conserved domains and motifs important for the biological function of ZBTB24, which provides a basis for understanding the molecular mechanisms underlying the pathogenesis of ICF syndrome.

Elevated concentrations of serum matrix metalloproteinase-2 and -9 and their associations with circulating markers of cardiovascular diseases in chronic arsenic-exposed individuals.

BACKGROUND: Cardiovascular diseases (CVDs) and cancers are the major causes of chronic arsenic exposure-related morbidity and mortality. Matrix metalloproteinase-2 (MMP-2) and -9 (MMP-9) are deeply involved in the pathogenesis of CVDs and cancers. This study has been designed to evaluate the interactions of arsenic exposure with serum MMP-2 and MMP-9 concentrations especially in relation to the circulating biomarkers of CVDs. METHODS: A total of 373 human subjects, 265 from arsenic-endemic and 108 from non-endemic areas in Bangladesh were recruited for this study. Arsenic concentrations in the specimens were measured by inductively coupled plasma mass spectroscopy (ICP-MS) and serum MMPs were quantified by immunoassay kits. RESULTS: Serum MMP-2 and MMP-9 concentrations in arsenic-endemic population were significantly (p < 0.001) higher than those in non-endemic population. Both MMPs showed significant positive interactions with drinking water (r s = 0.208, p < 0.001 for MMP-2; r s = 0.163, p < 0.01 for MMP-9), hair (r s = 0.163, p < 0.01 for MMP-2; r s = 0.173, p < 0.01 for MMP-9) and nail (r s = 0.160, p < 0.01 for MMP-2; r s = 0.182, p < 0.001 for MMP-9) arsenic of the study subjects. MMP-2 concentrations were 1.02, 1.03 and 1.05 times, and MMP-9 concentrations were 1.03, 1.06 and 1.07 times greater for 1 unit increase in log-transformed water, hair and nail arsenic concentrations, respectively, after adjusting for covariates (age, sex, BMI, smoking habit and hypertension). Furthermore, both MMPs were increased dose-dependently when the study subjects were split into three (≤10, 10.1-50 and > 50 µg/L) groups based on the regulatory upper limit of water arsenic concentration set by WHO and Bangladesh Government. MMPs were also found to be significantly (p < 0.05) associated with each other. Finally, the concentrations of both MMPs were correlated with several circulating markers related to CVDs. CONCLUSIONS: This study showed the significant positive associations and dose-response relationships of arsenic exposure with serum MMP-2 and MMP-9 concentrations. This study also showed the interactions of MMP-2 and MMP-9 concentrations with the circulating markers of CVDs suggesting the MMP-2 and MMP-9 -mediated mechanism of arsenic-induced CVDs.

Elevated levels of plasma uric acid and its relation to hypertension in arsenic-endemic human individuals in Bangladesh.

Blood uric acid has been recognized as a putative marker for cardiovascular diseases (CVDs). CVDs are the major causes of arsenic-related morbidity and mortality. However, the association of arsenic exposure with plasma uric acid (PUA) levels in relation to CVDs has not yet been explored. This study for the first time demonstrated the associations of arsenic exposure with PUA levels and its relationship with hypertension. A total of 483 subjects, 322 from arsenic-endemic and 161 from non-endemic areas in Bangladesh were recruited as study subjects. Arsenic concentrations in the drinking water, hair and nails of the study subjects were measured by inductively coupled plasma mass spectroscopy. PUA levels were measured using a colorimetric method. We found that PUA levels were significantly (p<0.001) higher in males and females living in arsenic-endemic areas than those in non-endemic area. Arsenic exposure (water, hair and nail arsenic) levels showed significant positive correlations with PUA levels. In multiple regression analyses, arsenic exposure levels were found to be the most significant contributors on PUA levels among the other variables that included age, body mass index, blood urea nitrogen, and smoking. There were dose-response relationships between arsenic exposure and PUA levels. Furthermore, diastolic and systolic blood pressure showed significant positive correlations with PUA levels. Finally, the average PUA levels were significantly higher in the hypertensive group than those in the normotensive group in both males and females living in arsenic-endemic areas. These results suggest that arsenic exposure-related elevation of PUA levels may be implicated in arsenic-induced CVDs.

Hydrogeochemical quality and suitability studies of groundwater in northern Bangladesh.

Agriculture, rapid urbanization and geochemical processes have direct or indirect effects on the chemical composition of groundwater and aquifer geochemistry. Hydro-chemical investigations, which are significant for assessment of water quality, were carried out to study the sources of dissolved ions in groundwater of Dinajpur district, northern Bangladesh. The groundwater samplish were analyzed for physico-chemical properties like pH, electrical conductance, hardness, alkalinity, total dissolved solids and Ca2+, Mg2+, Na+, K+, CO3(2-), HCO3(-), SO4(2-) and Cl- ions, respectively. Based on the analyses, certain parameters like sodium adsorption ratio, soluble sodium percentage, potential salinity, residual sodium carbonate, Kelly's ratio, permeability index and Gibbs ratio were also calculated. The results showed that the groundwater of study area was fresh, slightly acidic (pH 5.3-6.4) and low in TDS (35-275 mg I(-1)). Ground water of the study area was found suitable for irrigation, drinking and domestic purposes, since most of the parameters analyzed were within the WHO recommended values for drinking water. High concentration of NO3- and Cl- was reported in areas with extensive agriculture and rapid urbanization. Ion-exchange, weathering, oxidation and dissolution of minerals were major geochemical processes governing the groundwater evolution in study area. Gibb's diagram showed that all the samples fell in the rock dominance field. Based on evaluation, it is clear that groundwater quality of the study area was suitable for both domestic and irrigation purposes.

Transcriptomics and biochemical evidence of trigonelline ameliorating learning and memory decline in the senescence-accelerated mouse prone 8 (SAMP8) model by suppressing proinflammatory cytokines and elevating neurotransmitter release.

In recent years, exploring natural compounds with functional properties to ameliorate aging-associated cognitive decline has become a research priority to ensure healthy aging. In the present study, we investigated the effects of Trigonelline (TG), a plant alkaloid, on memory and spatial learning in 16-week-old senescence-accelerated mouse model SAMP8 using an integrated approach for cognitive and molecular biology aspects. After 30 days of oral administration of TG at the dose of 5 mg/kg/day, the mice were trained in Morris Water Maze task. TG-treated SAMP8 mice exhibited significant improvement in the parameters of escape latency, distance moved, and annulus crossing index. Next, we performed a whole-genome transcriptome profiling of the mouse hippocampus using microarrays. Gene ontology analyses showed that a wide range of biological processes, including nervous system development, mitochondrial function, ATP synthesis, and several signaling pathways related to inflammation, autophagy, and neurotransmitter release, were significantly enriched in TG-treated SAMP8 compared to nontreated. Further, a nonlinear dimensionality reduction technique, Uniform Manifold Approximation and Projection (UMAP), was applied to identify clusters of functions that revealed TG primarily regulated pathways related to inflammation, followed by those involved in neurotransmitter release. In addition, a protein-protein interaction network analysis indicated that TG may exert its biological effects through negatively modulating Traf6-mediated NF-κB activation. Finally, ELISA test showed that TG treatment significantly decreased proinflammatory cytokines- TNFα and IL6 and increased neurotransmitters- dopamine, noradrenaline, and serotonin in mouse hippocampus. Altogether, our integrated bio-cognitive approach highlights the potential of TG in alleviating age-related memory and spatial impairment.

Current Status of Circulating Tumor Cells in Head and Neck Squamous Cell Carcinoma: A Review.

OBJECTIVE: Circulating tumor cells (CTCs) are found in the blood of patients with cancer, including head and neck squamous cell carcinomas (HNSCCs). The aim is to review the most up-to-date status of CTCs for applications in patients with HNSCC. DATA SOURCES: English articles in PubMed. REVIEW METHODS: All the studies on CTCs in HNSCCs in the literature were reviewed. CONCLUSIONS: There is emerging information on the diagnostic and prognostic value of CTCs in HNSCCs. Evidence also highlights the advantages of various downstream analysis approaches over circulating tumor DNA (ctDNA), such as single-CTC analysis, ex vivo, and in vivo expansion of CTCs. Multiple phenotypic surface markers (cytokeratins, EpCAM, vimentin, etc.), used for CTCs characterization using different immunoassays, could predict disease progression as well as patients' response to treatment efficacy. Immune checkpoint inhibitors' status in CTCs could also provide better insight into treatment. Clonal expansion of CTCs and single-cell analysis of CTCs are the most emerging fields nowadays which may offer an understanding of the mechanism of tumor evolution as well as therapeutic efficacy. Although several clinical trials are ongoing, limitations still exist in the detection and characterization of CTCs. Due to the lack of a gold standard protocol, the sensitivity and specificity of CTC enumeration methods vary. IMPLICATIONS FOR PRACTICE: Prospective clinical trials are still needed before CTCs can be employed as diagnostic and prognostic markers in the clinical management of patients with HNSCC.

Correlation between KRAS Mutation and CTLA-4 mRNA Expression in Circulating Tumour Cells: Clinical Implications in Colorectal Cancer.

Combination strategies of KRAS inhibition with immunotherapy in treating advanced or recurrent colorectal carcinoma (CRC) may need to be assessed in circulating tumour cells (CTCs) to achieve better clinical outcomes. This study aimed to investigate the genomic variations of KRAS in CTCs and matched CRC tissues and compared mRNA expression of KRAS and CTLA-4 between wild-type and KRAS-mutated CTCs and CRC tissues. Clinicopathological correlations were also compared. Six known mutations of KRAS were identified at both codon 12 and codon 13 (c.35G>T/G12V, c.35G>A7/G12D, c.35G>C/G12A, c.34G>A/G12S, c.38G>C/G13A, and c.38G>A/G13D). Three CTC samples harboured the identified mutations (16.7%; 3/18), while fifteen matched primary tumour tissues (65.2%, 15/23) showed the mutations. CTCs harbouring the KRAS variant were different from matched CRC tissue. All the mutations were heterozygous. Though insignificant, CTLA-4 mRNA expression was higher in patients carrying KRAS mutations. Patients harbouring KRAS mutations in CTCs were more likely to have poorly differentiated tumours (p = 0.039) and with lymph node metastasis (p = 0.027) and perineural invasion (p = 0.014). KRAS mutations in CTCs were also significantly correlated with overall pathological stages (p = 0.027). These findings imply the genetic basis of KRAS with immunotherapeutic target molecules based on a real-time platform. This study also suggests the highly heterogeneous nature of cancer cells, which may facilitate the assessment of clonal dynamics across a single patient's disease.

Biological and clinical significance of the glypican-3 gene in human lung adenocarcinoma: An in silico analysis.

Glypican-3 (GPC3), a membrane-bound heparan sulfate proteoglycan, has long been found to be dysregulated in human lung adenocarcinomas (LUADs). Nevertheless, the function, mutational profile, epigenetic regulation, co-expression profile, and clinicopathological significance of the GPC3 gene in LUAD progression are not well understood. In this study, we analyzed cancer microarray datasets from publicly available databases using bioinformatics tools to elucidate the above parameters. We observed significant downregulation of GPC3 in LUAD tissues compared to their normal counterparts, and this downregulation was associated with shorter overall survival (OS) and relapse-free survival (RFS). Nevertheless, no significant differences in the methylation pattern of GPC3 were observed between LUAD and normal tissues, although lower promoter methylation was observed in male patients. GPC3 expression was also found to correlate significantly with infiltration of B cells, CD8+, CD4+, macrophages, neutrophils, and dendritic cells in LUAD. In addition, a total of 11 missense mutations were identified in LUAD patients, and ~1.4% to 2.2% of LUAD patients had copy number amplifications in GPC3. Seventeen genes, mainly involved in dopamine receptor-mediated signaling pathways, were frequently co-expressed with GPC3. We also found 11 TFs and 7 miRNAs interacting with GPC3 and contributing to disease progression. Finally, we identified 3 potential inhibitors of GPC3 in human LUAD, namely heparitin, gemcitabine and arbutin. In conclusion, GPC3 may play an important role in the development of LUAD and could serve as a promising biomarker in LUAD.

AHR gene expression and the polymorphism rs2066853 are associated with clinicopathological parameters in colorectal carcinoma.

The relationship between red and processed meat and its risk toward colorectal carcinoma (CRC) is not fully explored in literature. Polycyclic aromatic hydrocarbons (PAHs) are procarcinogenic molecules that are ingested with meat cooked at high temperatures. The metabolic conversion of PAHs to carcinogenic diol epoxides is in part mediated by the aryl hydrocarbon receptor (AhR)-dependent induction of CYP1A1. This study aims to examine the expression profiles and polymorphisms of the AHR (aryl hydrocarbon receptor) gene which is involved in the metabolic conversion of PAHs in patients with CRC. Genetic analysis was done in matched cancer and non-neoplastic tissues from 79 patients diagnosed with CRCs. Low AHR mRNA expression was associated with mucinous colorectal adenocarcinoma. Exon 10 of AHR showed that 27% of patients had the rs2066853 single-nucleotide polymorphism resulting in an arginine-to-lysine change at codon 554. This variant was significantly associated with a lower likelihood of perineural invasion, presence of synchronous cancer, and multiple colorectal polyps. Furthermore, rs2066853 individuals were significantly more likely to be of more advanced age and have a more favorable tumor grade and pathological stage. These results imply the pathogenic roles of AHR in PAH-associated colorectal carcinogenesis.

Polymorphisms in PAH metabolising enzyme CYP1A1 in colorectal cancer and their clinicopathological correlations.

CYP1A1 enzyme is integral to the biotransformation of polycyclic aromatic hydrocarbons to carcinogenic compounds. This study aimed to screen mutations in exon 7 (ex7) of CYP1A1 and investigate its clinicopathological correlations in fresh tissue samples from 85 patients (42 women; 43 men) with colorectal carcinoma (CRC). Tumour tissues and matched non-neoplastic mucosa tissues were collected prospectively. Genomic DNA was extracted from all tissues, and subject to high-resolution melt curve analysis for CYP1A1-ex7. Sanger sequencing was employed to detect specific mutations. Three known single nucleotide polymorphisms (SNPs) were identified in both tumour and matched non-neoplastic tissue for the same individual. Of the 85 patients, one third (n = 28) harboured either rs1048943, rs1799814, or rs41279188. Patients who had a SNP at ex7 of CYP1A1 were significantly more likely to be over 65 years of age (p = 0.015). Furthermore, individuals harbouring a SNP at exon7 showed a low incidence of perineural cancer infiltration (p = 0.025) when compared to the wild-type population. Overall, polymorphisms at exon 7 of CYP1A1 are present in patients with CRC and associated with a few clinicopathological characteristics.

Heme oxygenase-1 & 2 and their potential contribution in heme induced colorectal carcinogenesis.

BACKGROUND: Literature suggests Heme oxygenase (HO) system to be a double-edged sword which can promote both cytoprotection as well as carcinogenicity. The aim of this study was to investigate the role of heme in HO-1 and HO-2 induced colorectal carcinogenesis and the clinicopathological significance of their expressions in patients with colorectal carcinoma (CRC). METHODS: HO-1 and HO-2 expression alterations in normal colonic epithelial (FHC) and colon cancer cells (SW480) were explored following treatment with 0 µM, 25 µM, 100 µM and 250 µM concentrations of hemin, using qPCR. Fifty paired CRC and adjacent non-neoplastic samples were subjected to qPCR to determine the HO-1 and HO-2 expression. Clinicopathological associations of HO-1 and HO-2 expression levels were determined. RESULTS: Low concentrations of hemin caused upregulation and high concentration caused downregulation of HO-1 expression, whereas HO-2 expression was significantly downregulated with all hemin concentrations in FHC. HO-1 expression in SW480 was increased with all hemin concentrations and HO-2 expression was downregulated at the highest hemin concentration. HO-1 and HO-2 expressions in adjacent non-neoplastic tissue was significantly higher than that of CRC. Expression of HO-1 was significantly higher than HO-2, in both CRC and adjacent non-neoplastic tissue. Sex, HFE expression and lymph-vascular invasion were significantly correlated with HO-1 expression. HO-2 expression showed significant associations with staging, local spread and recurrence of tumour. CONCLUSION: HO-1 and HO-2 expression is respectively induced and repressed by exogenous hemin in normal colon and colon cancer cells. HO-1 and HO-2 expression profiles in CRC are correlated with the assessed clinicopathological features of CRC, suggesting the possible implications of HO expression status in CRC pathogenesis.

Incorporation of Plasmid DNA Into Bacterial Membrane Vesicles by Peptidoglycan Defects in Escherichia coli.

Membrane vesicles (MVs) are released by various prokaryotes and play a role in the delivery of various cell-cell interaction factors. Recent studies have determined that these vesicles are capable of functioning as mediators of horizontal gene transfer. Outer membrane vesicles (OMVs) are a type of MV that is released by Gram-negative bacteria and primarily composed of outer membrane and periplasm components; however, it remains largely unknown why DNA is contained within OMVs. Our study aimed to understand the mechanism by which DNA that is localized in the cytoplasm is incorporated into OMVs in Gram-negative bacteria. We compared DNA associated with OMVs using Escherichia coli BW25113 cells harboring the non-conjugative, non-mobilized, and high-copy plasmid pUC19 and its hypervesiculating mutants that included ΔnlpI, ΔrseA, and ΔtolA. Plasmid copy per vesicle was increased in OMVs derived from ΔnlpI, in which peptidoglycan (PG) breakdown and synthesis are altered. When supplemented with 1% glycine to inhibit PG synthesis, both OMV formation and plasmid copy per vesicle were increased in the wild type. The bacterial membrane condition test indicated that membrane permeability was increased in the presence of glycine at the late exponential phase, in which cell lysis did not occur. Additionally, quick-freeze deep-etch and replica electron microscopy observations revealed that outer-inner membrane vesicles (O-IMVs) are formed in the presence of glycine. Thus, two proposed routes for DNA incorporation into OMVs under PG-damaged conditions are suggested. These routes include DNA leakage due to increased membrane permeation and O-IMV formation. Additionally, our findings contribute to a greater understanding of the vesicle-mediated horizontal gene transfer that occurs in nature and the utilization of MVs for DNA cargo.

HFE variants in colorectal cancer and their clinicopathological correlations.

The study aimed to screen mutation of human homeostatic iron regulator (HFE) in colorectal carcinoma (CRC) and detect their associations with clinicopathological parameters. Expression of HFE was determined by quantitative polymerase chain reaction in matched CRC and non neoplastic colorectal mucosal tissue of 76 patients. Genomic DNA extracted were subjected to high high-resolution melt curve analysis and Sanger sequencing to detect mutations in HFE. The associations of the identified mutations with a variety of clinical features were determined. Approximately 60% of CRC showed low HFE expression. Of the ten 10 mutations identified in exons 2 and 4, c.187C>G (H63D), c845G>A (C282Y), c.193A>T (S65C), g.3828T>C, g.5795T>C, and g.5728G>A were known mutations. Four novel mutations were discovered; : c.184G>A, c.220T>G, c.322A>C, and c.324T>C. Heterozygous H63D and C282Y mutations were seen in 71% and 49% of cancer tissue, respectively. Tumour site (p = 0.048) and gender (p = 0.039) were significantly associated with H63D and C282Y mutation status, respectively. Local spread of cancer was significantly associated with C282Y mutations in CRC cancer and adjacent non-neoplastic tissue (p = 0.029 & and p = 0.004, respectively). There was a statistically significant association between H63D and C282Y negativity in matched non-neoplastic colorectal mucosa tissue and pathological staging of cancer (p = 0.047 & and p = 0.001, respectively). Patients with H63D and C282Y mutations in cancer tissue tend to have higher survival rates. Hence HFE mutations are common in CRC and are associated with clinicopathological parameters, implying the potential clinical significance of HFE mutations in colorectal carcinogenesis.

Environmental contamination of lead in dairy farms in Narayangonj, Bangladesh.

OBJECTIVE: In recent years, lead (Pb) has arisen as a foremost contaminant due to overpopulation, rapid industrialization, and expansion that could contaminate the human food chain. However, the correlation between the environmental contamination of Pb and its spatial transfer to the dairy products is still unmapped. In this paper, we intend to evaluate the concentration of environmental Pb and its spatial distribution in dairy feed and products in Narayangonj, a highly polluted district of Bangladesh. MATERIALS AND METHODS: A total of 125 samples of soil, water, forage, and milk were collected from five upazilas (Narayangonj Sadar, Bondor, Rupgonj, Araihazar, and Sonargoan) of Narayangonj. The samples were digested by acid digestion, and Pb was detected by flame atomic absorption spectrophotometer and graphite furnace atomic absorption spectrometer. The bioconcentration factor of the samples was also calculated. RESULTS: In this study, 25% of the soil, 20% of the water, 5% of the forage, and 2% of the milk samples contained Pb at a variable level. Among the environmental samples, the highest concentration (26-39 µg/kg) of Pb was detected in the soil, followed by in the water (0.023-0.059 µg/kg) and forage (0.017-0.035 µg/kg). The contamination (0.041-0.068 µg/kg) in the milk, however, was lower than the soil but higher than the water and forage. The concentration of Pb in all the samples was within a safer limit. None of the forage samples was the potential bioaccumulator. CONCLUSION: Although no linear correlation was established between the environmental samples, forage, and milk, the study identifies the potentials of the spatial distribution of Pb from the environment to the dairy feed and products. Therefore, feasible procedures should be adapted to cease the residue to the human food chain.

Butyrylcholinesterase-a potential plasma biomarker in manganese-induced neurobehavioral changes.

Groundwater particularly drinking water contamination with metals has created an environmental disaster in Bangladesh. Manganese (Mn), an essential trace element, plays a key role in the development and function of the brain. Excess Mn exposure is reported to be associated with complex neurological disorders. Here, we have found a notably large extent of Mn above the permissive limit in the tube-well water of Rajshahi and Naogaon districts in Bangladesh. Higher levels of Mn in hair and nail samples, and a decreasing level of butyrylcholinesterase (BChE) activity were detected in plasma samples of the human subjects recruited from Naogaon district. Mn concentrations in water, hair, and nails were negatively correlated with the plasma BChE levels in Mn-exposed populations. To compare and validate these human studies, an animal model was used to determine the in vivo effects of Mn on neurobehavioral changes and blood BChE levels. In elevated plus maze, the time spent was significantly reduced in open arms and increased in closed arms of Mn-exposed mice compared to control group. The mean latency time to find the platform was declined significantly in control mice compared to Mn-treated group during 7 days in Morris water maze test, and Mn-exposed group also spent significantly less time in the desired quadrant as compared to the control group in probe trial. BChE activity was significantly reduced in Mn-exposed mice compared to control mice. Taken together, these results suggest that plasma BChE levels may serve as reliable biomarker of Mn-induced neurotoxicity related to behavioral changes.

A New Electron Microscopic Method to Observe the Distribution of Phosphatidylinositol 3,4-bisphosphate.

Phosphatidylinositol 3,4-bisphosphate [PtdIns(3,4)P2] is a phosphoinositide that plays important roles in signal transduction, endocytosis, and cell migration among others. The intracellular distribution of PtdIns(3,4)P2 has mainly been studied by observing the distribution of GFP-tagged PtdIns(3,4)P2-binding protein domains in live cells and by labeling with anti-PtdIns(3,4)P2 antibody in fixed cell samples, but these methods only offer low spatial resolution results and may have pitfalls. In the present study, we developed an electron microscopic method to observe the PtdIns(3,4)P2 distribution using the SDS-treated freeze-fracture replica labeling method. The recombinant GST-tagged pleckstrin homology (PH) domain of TAPP1 was used as the binding probe, and its binding to PtdIns(3,4)P2 in the freeze-fracture replica was confirmed by using liposomes containing different phosphoinositides and by the lack of labeling by a mutant probe, in which one amino acid in the PH domain was substituted. The method was applied to NIH3T3 cell samples and showed that the increase of PtdIns(3,4)P2 in cells treated with hydrogen peroxide occurs in the cytoplasmic leaflet of the plasma membrane, except in the caveolar membrane. The present method can define the distribution of PtdIns(3,4)P2 at a high spatial resolution and will facilitate our understanding of the physiological function of this less studied phosphoinositide.

Individual and Combined Effects of Arsenic and Lead on Behavioral and Biochemical Changes in Mice.

Arsenic (As) toxicity has caused an environmental tragedy affecting millions of people in the world. Little is known about the toxic effects of As on neurobehavioral and biochemical changes in vivo. Along this line of metal toxicity, co-exposure of lead (Pb) could aggravate the situation in the host. The present study was designed to explore the combined effects of As and Pb on behavioral changes like anxiety, spatial memory and learning impairment, and blood indices related to organ dysfunction. Exposure of mice to As (10 mg/kg body weight), Pb (10 mg/kg body weight), and As + Pb via drinking water significantly decreased the time spent exploring the open arms while it increased the time spent in the closed arms compared to control mice in the elevated plus maze. The mean latency time of the control group to find the platform decreased significantly during the learning for 7 days compared to all three treated groups in the Morris water maze test, and the As-exposed group spent significantly less time in the desired quadrant as compared to the control group in the probe trial. Both metals posed an anxiety-like behavior and deficits in spatial memory and learning, and also altered blood indices related to liver and kidney dysfunction, and a combined exposure of these metals inhibited the individual accumulation of As and Pb. Taken together, these data suggest that As has more toxic effects on neurobehavioral and biochemical changes than Pb, and there may be antagonism in the effects and accumulation between these two toxicants.

Arsenic-induced Histological Alterations in Various Organs of Mice.

Deposition of arsenic in mice through groundwater is well documented but little is known about the histological changes of organs by the metalloid. Present study was designed to evaluate arsenic-induced histological alterations in kidney, liver, thoracic artery and brain of mice which are not well documented yet. Swiss albino male mice were divided into 2 groups and treated as follows: Group 1: control, 2: arsenic (sodium arsenite at 10 mg/kg b.w. orally for 8 wks). Group 2 showed marked degenerative changes in kidney, liver, thoracic artery, and brain whereas Group 1 did not reveal any abnormalities on histopathology. We therefore concluded that arsenic induces histological alterations in the tested organs.