O-GlcNAcylation of GLI transcription factors in hyperglycemic conditions augments Hedgehog activity.

Modification of proteins by O-linked ß-N-acetylglucosamine (O-GlcNAc) promotes tumor cell survival, proliferation, epigenetic changes, angiogenesis, invasion, and metastasis. Here we demonstrate that in conditions of elevated glucose, there is increased expression of key drug resistance proteins (ABCB1, ABCG2, ERCC1, and XRCC1), all of which are regulated by the Hedgehog pathway. In elevated glucose conditions, we determined that the Hedgehog pathway transcription factors, GLI1 and GLI2, are modified by O-GlcNAcylation. This modification functionally enhanced their transcriptional activity. The activity of GLI was enhanced when O-GlcNAcase was inhibited, while inhibiting O-GlcNAc transferase caused a decrease in GLI activity. The metabolic impact of hyperglycemic conditions impinges on maintaining PKM2 in the less active state that facilitates the availability of glycolytic intermediates for biosynthetic pathways. Interestingly, under elevated glucose conditions, PKM2 directly influenced GLI activity. Specifically, abrogating PKM2 expression caused a significant decline in GLI activity and expression of drug resistance proteins. Cumulatively, our results suggest that elevated glucose conditions upregulate chemoresistance through elevated transcriptional activity of the Hedgehog/GLI pathway. Interfering in O-GlcNAcylation of the GLI transcription factors may be a novel target in controlling cancer progression and drug resistance of breast cancer.

Neurofibromin (NF1) genetic variant structure-function analyses using a full-length mouse cDNA.

Neurofibromatosis type 1 (NF1) is caused by pathogenic variants or mutations in the NF1 gene that encodes neurofibromin. We describe here a new approach to determining the functional consequences of NF1 genetic variants. We established a heterologous cell culture expression system using a full-length mouse Nf1 cDNA (mNf1) and human cell lines. We demonstrate that the full-length murine cDNA produces a > 250 kDa neurofibromin protein that is capable of modulating Ras signaling. We created mutant cDNAs representing NF1 patient variants with different clinically relevant phenotypes, and assessed their ability to produce mature neurofibromin and restore Nf1 activity in NF1-/- cells. These cDNAs represent variants in multiple protein domains and various types of clinically relevant predicted variants. This approach will help advance research on neurofibromin structure and function, determine pathogenicity for missense variants, and allow for the development of activity assays and variant-directed therapeutics.

Programmable nanowire circuits for nanoprocessors.

A nanoprocessor constructed from intrinsically nanometre-scale building blocks is an essential component for controlling memory, nanosensors and other functions proposed for nanosystems assembled from the bottom up. Important steps towards this goal over the past fifteen years include the realization of simple logic gates with individually assembled semiconductor nanowires and carbon nanotubes, but with only 16 devices or fewer and a single function for each circuit. Recently, logic circuits also have been demonstrated that use two or three elements of a one-dimensional memristor array, although such passive devices without gain are difficult to cascade. These circuits fall short of the requirements for a scalable, multifunctional nanoprocessor owing to challenges in materials, assembly and architecture on the nanoscale. Here we describe the design, fabrication and use of programmable and scalable logic tiles for nanoprocessors that surmount these hurdles. The tiles were built from programmable, non-volatile nanowire transistor arrays. Ge/Si core/shell nanowires coupled to designed dielectric shells yielded single-nanowire, non-volatile field-effect transistors (FETs) with uniform, programmable threshold voltages and the capability to drive cascaded elements. We developed an architecture to integrate the programmable nanowire FETs and define a logic tile consisting of two interconnected arrays with 496 functional configurable FET nodes in an area of ∼960 µm(2). The logic tile was programmed and operated first as a full adder with a maximal voltage gain of ten and input-output voltage matching. Then we showed that the same logic tile can be reprogrammed and used to demonstrate full-subtractor, multiplexer, demultiplexer and clocked D-latch functions. These results represent a significant advance in the complexity and functionality of nanoelectronic circuits built from the bottom up with a tiled architecture that could be cascaded to realize fully integrated nanoprocessors with computing, memory and addressing capabilities.

Nanowire nanocomputer as a finite-state machine.

Implementation of complex computer circuits assembled from the bottom up and integrated on the nanometer scale has long been a goal of electronics research. It requires a design and fabrication strategy that can address individual nanometer-scale electronic devices, while enabling large-scale assembly of those devices into highly organized, integrated computational circuits. We describe how such a strategy has led to the design, construction, and demonstration of a nanoelectronic finite-state machine. The system was fabricated using a design-oriented approach enabled by a deterministic, bottom-up assembly process that does not require individual nanowire registration. This methodology allowed construction of the nanoelectronic finite-state machine through modular design using a multitile architecture. Each tile/module consists of two interconnected crossbar nanowire arrays, with each cross-point consisting of a programmable nanowire transistor node. The nanoelectronic finite-state machine integrates 180 programmable nanowire transistor nodes in three tiles or six total crossbar arrays, and incorporates both sequential and arithmetic logic, with extensive intertile and intratile communication that exhibits rigorous input/output matching. Our system realizes the complete 2-bit logic flow and clocked control over state registration that are required for a finite-state machine or computer. The programmable multitile circuit was also reprogrammed to a functionally distinct 2-bit full adder with 32-set matched and complete logic output. These steps forward and the ability of our unique design-oriented deterministic methodology to yield more extensive multitile systems suggest that proposed general-purpose nanocomputers can be realized in the near future.

Nonclassical activation of Hedgehog signaling enhances multidrug resistance and makes cancer cells refractory to Smoothened-targeting Hedgehog inhibition.

The Hedgehog (Hh) pathway is critical in normal development. However, it has been reported to be up-regulated in numerous cancers and implicated in tumorigenicity and metastasis. Classical activation of Hh signaling initiated by Hh ligands results in activation of Smoothened (SMOH) and culminates in the activation of the GLI transcription factors. Classical Hh signaling is autocrine or paracrine (involving interaction between tumor cells and their stroma/microenvironment). The tumor milieu is rich in inflammatory cytokines that can modulate tumor cell behavior. Here, we show for the first time that the Hh pathway can be nonclassically up-regulated by the inflammatory cytokine, osteopontin (OPN). OPN-initiated Akt-GSK3ß signaling mediates the subcellular distribution and activation of GLI1 resulting in the modulation of epithelial mesenchymal plasticity and drug resistance. Interestingly, the SMOH inhibitor cyclopamine was unable to uncouple the effects of OPN on Hh signaling, indicating that OPN nonclassically activates GLI-mediated transcription. Given the fact that OPN is itself transcriptionally activated upon Hh signaling, our current findings highlight the possibility of a feedforward vicious cycle such that the Hh pathway might be turned on nonclassically by stimuli from the tumor milieu. Thus, drugs that target the classical Hh ligand-mediated activation of Hh signaling may be compromised in their ability to interfere with the functioning of the pathway.

Hedgehog signaling induced by breast cancer cells promotes osteoclastogenesis and osteolysis.

Bone integrity is maintained by a dynamic equilibrium between the activities of bone-forming osteoblasts and bone-resorbing osteoclasts. Osteolytic lesions are a painful consequence of metastasis of breast cancer cells to bone in an overwhelming majority of breast cancer patients. Factors secreted by breast cancer cells propel a cascade of events that trigger osteoclastogenesis and elevated bone resorption. In the present study, we show that the Hedgehog (Hh) ligands secreted by breast cancer cells promote osteoclast differentiation and potentiate the activity of mature osteoclasts. Paracrine Hh signaling induced by breast cancer cells mediates a detrimental chain of events by the up-regulation of osteopontin (OPN), which in turn enhances osteoclastic activity by up-regulating cathepsin K and MMP9. Hh signaling is essential for osteoclasts because blocking the Hh pathway using the pharmacological Hh inhibitor, cyclopamine, results in an overall decrease in osteoclastogenesis and resorptive activity. Our studies suggest that inhibiting Hh signaling interferes with the ability of pre-osteoclasts to respond to the stimulatory effects of the breast cancer cells, indicating that Hh signaling is vital to osteoclast activity.

Loss of tumor suppressor Merlin in advanced breast cancer is due to post-translational regulation.

Unlike malignancies of the nervous system, there have been no mutations identified in Merlin in breast cancer. As such, the role of the tumor suppressor, Merlin, has not been investigated in breast cancer. We assessed Merlin expression in breast cancer tissues by immunohistochemistry and by real-time PCR. The expression of Merlin protein (assessed immunohistochemically) was significantly decreased in breast cancer tissues (although the transcript levels were comparable) simultaneous with increased expression of the tumor-promoting protein, osteopontin (OPN). We further demonstrate that the loss of Merlin in breast cancer is brought about, in part, due to OPN-initiated Akt-mediated phosphorylation of Merlin leading to its proteasomal degradation. Restoring expression of Merlin resulted in reduced malignant attributes of breast cancer, characterized by reduced invasion, migration, motility, and impeded tumor (xenograft) growth in immunocompromised mice. The possibility of developing a model using the relationship between OPN and Merlin was tested with a logistic regression model applied to immunohistochemistry data. This identified consistent loss of immunohistochemical expression of Merlin in breast tumor tissues. Thus, we demonstrate for the first time a role for Merlin in impeding breast malignancy, identify a novel mechanism for the loss of Merlin protein in breast cancer, and have developed a discriminatory model using Merlin and OPN expression in breast tumor tissues.

Quantitative Longitudinal Imaging Reveals that Inhibiting Hedgehog Activity Alleviates the Hypoxic Tumor Landscape.

Metastases account for the majority of mortalities related to breast cancer. The onset and sustained presence of hypoxia strongly correlates with increased incidence of metastasis and unfavorable prognosis in patients with breast cancer. The Hedgehog (Hh) signaling pathway is dysregulated in breast cancer, and its abnormal activity enables tumor progression and metastasis. In addition to programming tumor cell behavior, Hh activity enables tumor cells to craft a metastasis-conducive microenvironment. Hypoxia is a prominent feature of growing tumors that impacts multiple signaling circuits that converge upon malignant progression. We investigated the role of Hh activity in crafting a hypoxic environment of breast cancer. We used radioactive tracer [18F]-fluoromisonidazole (FMISO) positron emission tomography (PET) to image tumor hypoxia. We show that tumors competent for Hh activity are able to establish a hypoxic milieu; pharmacologic inhibition of Hh signaling in a syngeneic mammary tumor model mitigates tumor hypoxia. Furthermore, in hypoxia, Hh activity is robustly activated in tumor cells and institutes increased HIF signaling in a VHL-dependent manner. The findings establish a novel perspective on Hh activity in crafting a hypoxic tumor landscape and molecularly navigating the tumor cells to adapt to hypoxic conditions. IMPLICATIONS: Importantly, we present a translational strategy of utilizing longitudinal hypoxia imaging to measure the efficacy of vismodegib in a preclinical model of triple-negative breast cancer.

The hedgehog pathway transcription factor GLI1 promotes malignant behavior of cancer cells by up-regulating osteopontin.

The role of Hedgehog (Hh) signaling as a developmental pathway is well established. Several recent studies have implicated a role for this pathway in multiple cancers. In this study we report that expression of GLI1 and osteopontin (OPN) increase progressively with the progression of melanoma from primary cutaneous cancer to metastatic melanoma in clinically derived specimens. We have further determined that OPN is a direct transcriptional target of GLI1. We have observed that OPN expression is stimulated in the presence of Hh ligands and inhibited in the presence of the Smoothened (SMO) inhibitor, cyclopamine. Transcriptional silencing of GLI1 negatively impacts OPN expression and compromises the ability of cancer cells to proliferate, migrate, and invade in vitro and interferes with their ability to grow as xenografts and spontaneously metastasize in nude mice. These altered attributes could be rescued by re-expressing OPN in the GLI1-silenced cells, suggesting that OPN is a critical downstream effector of active GLI1 signaling. Our observations lead us to conclude that the GLI1-mediated up-regulation of OPN promotes malignant behavior of cancer cells.

Multifunctional effect of epigallocatechin-3-gallate (EGCG) in downregulation of gelatinase-A (MMP-2) in human breast cancer cell line MCF-7.

AIMS: The tumor inhibiting property of green tea polyphenol epigallocatechin-3-gallate (EGCG) is well documented. Studies reveal that matrix-metalloproteinases (MMPs) play pivotal roles in tumor invasion through degradation of basement membranes and extracellular matrix (ECM). We studied the effect of EGCG on matrixmetalloproteinases-2 (MMP-2), the factors involved in activation, secretion and signaling molecules that might be involved in the regulation of MMP-2 in human breast cancer cell line, MCF-7. MAIN METHODS: MCF-7 was treated with EGCG (20 muM, 24 h), the effect of EGCG on MMP-2 expression, activity and its regulatory molecules were studied by gelatin zymography, Western blot, quantitative and semi-quantitative real time RT-PCR, immunoflourescence and cell adhesion assay. KEY FINDINGS: EGCG treatment reduced the activity, protein expression and mRNA expression level of MMP-2. EGCG treatment reduced the expression of focal adhesion kinase (FAK), membrane type-1-matrix metalloproteinase (MT1-MMP), nuclear factor-kappa B (NF-kB), vascular endothelial growth factor (VEGF) and reduced the adhesion of MCF-7 cells to ECM, fibronectin and vitronectin. Real time RT-PCR revealed a reduced expression of integrin receptors alpha5, beta1, alphav and beta3 due to EGCG treatment. SIGNIFICANCE: Down regulation of expression of MT1-MMP, NF-kB, VEGF and disruption of functional status of integrin receptors may indicate decreased MMP-2 activation; low levels of FAK expression might indicate disruption in FAK-induced MMP-2 secretion and decrease in activation of phosphatidyl-inositol-3-kinase (PI-3K), extracellular regulated kinase (ERK) indicates probable hindrance in MMP-2 regulation and induction. We propose EGCG as potential inhibitor of expression and activity of pro-MMP-2 by a process involving multiple regulatory molecules in MCF-7.

Performance simulation and analysis of a CMOS/nano hybrid nanoprocessor system.

This paper provides detailed simulation results and analysis of the prospective performance of hybrid CMOS/nanoelectronic processor systems based upon the field-programmable nanowire interconnect (FPNI) architecture. To evaluate this architecture, a complete design was developed for an FPNI implementation using 90 nm CMOS with 15 nm wide nanowire interconnects. Detailed simulations of this design illustrate that critical design choices and tradeoffs exist beyond those specified by the architecture. This includes the selection of the types of junction nanodevices, as well as the implementation of low-level circuits. In particular, the simulation results presented here show that only nanodevices with an 'on/off' current ratio of 200 or more are suitable to produce correct system-level behaviour. Furthermore, the design of the CMOS logic gates in the FPNI system must be customized to accommodate the resistances of both 'on'-state and 'off'-state nanodevices. Using these customized designs together with models of suitable nanodevices, additional simulations demonstrate that, relative to conventional 90 nm CMOS FPGA systems, performance gains can be obtained of up to 70% greater speed or up to a ninefold reduction in energy consumption.

Gold Nanoparticles sensitize pancreatic cancer cells to gemcitabine.

Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest solid cancers with dismal prognosis. Several mechanisms that are mainly responsible for aggressiveness and therapy resistance of PDAC cells include epithelial to mesenchymal transition (EMT), stemness and Mitogen Activated Protein Kinase (MAPK) signaling. Strategies that inhibit these mechanisms are critically important to improve therapeutic outcome in PDAC. In the current study, we wanted to investigate whether gold nanoparticles (AuNPs) could sensitize pancreatic cancer cells to the chemotherapeutic agent gemcitabine. We demonstrated that treatment with AuNPs of 20 nm diameter inhibited migration and colony forming ability of pancreatic cancer cells. Pre-treatment with AuNPs sensitized pancreatic cancer cells to gemcitabine in both viability and colony forming assays. Mechanistically, pre-treatment of pancreatic cancer cells with AuNPs decreased gemcitabine induced EMT, stemness and MAPK activation. Taken together, these findings suggest that AuNPs could be considered as a potential agent to sensitize pancreatic cancer cells to gemcitabine.

Rapid expression and activation of MMP-2 and MMP-9 upon exposure of human breast cancer cells (MCF-7) to fibronectin in serum free medium.

Interactions between tumour cells and the extracellular matrix (ECM) strongly influence tumour development, affecting cell survival, proliferation and migration. Many of these interactions are mediated through a family of cell surface receptors named integrins. Fibronectin and its integrin receptors play important roles in tumour development. The alpha5beta 1 integrin interacts with the central cell adhesive region of fibronectin and requires both the RGD and synergy sites for maximal binding. Matrix metalloproteinases (MMPs) are a family of zinc dependent endopeptidases. They are capable of digesting the different components of the ECM and basement membrane. The ECM gives structural support to cells and plays a central role in cell adhesion, differentiation, proliferation and migration. Binding of ECM to integrins modulates expression and activity of the different MMPs. Our experimental findings demonstrate that cultivation of human breast cancer cells, MCF-7, in serum free medium in the presence of fibronectin upregulates the activity of MMP-2 and MMP-9. Blocking of alpha5beta 1 integrin with anti-alpha5 monoclonal antibody inhibits the fibronectin-induced MMP activation response appreciably. This strongly indicates alpha5beta 1 mediated signalling events in activation of MMP-2 and MMP-9. Phosphorylation of FAK and PI-3 kinase and the nuclear translocation of ERK and NF-kappaB upon fibronectin binding demonstrate possible participation of the FAK/PI-3K/ERK signalling pathways in the regulation of MMP-2 activity.

Culture of human A375 melanoma cells in the presence of fibronectin causes expression of MMP-9 and activation of MMP-2 in culture supernatants.

UNLABELLED: Interactions between tumor cell surface integrin receptors and extracellular matrix (ECM) ligands play an important role in tumor development, affecting cell survival, proliferation, and migration. Integrin-ECM ligand interaction leads to phosphorylation of focal adhesion kinase (FAK) and activation of mitogen-activated protein kinase (MAPK) pathways. It has been reported that integrins also regulate expression and function of matrix metalloproteinases (MMPs). In this present work, we cultured human A375 melanoma cells in the presence of fibronectin to study fibronectin-integrin mediated modulation of MMP activity. METHODS: A375 cells were cultured in serum-free culture medium (SFCM) in the presence of fibronectin (25 microg/0.75 ml), SFCM was collected and gelatin zymography was performed. Western blot and RT-PCR were performed with A375 cells cultured in the presence of fibronectin. RESULTS: Culture of A375 cells in the presence of fibronectin led to expression of MMP-9 and activation of MMP-2 within 2 h. When cells were treated with ERK inhibitor (PD98059) or PI-3K inhibitor (LY294002) and grown in the presence of fibronectin, MMP-9 expression and MMP-2 activation was inhibited. Tyrosine phosphorylation of FAK and ERK were increased in A375 cells grown in the presence of fibronectin. Increased MMP-9 mRNA expression and processing of MT1-MMP were also observed in A375 cells grown in the presence of fibronectin. CONCLUSIONS: Our findings indicate culture of A375 cells in SFCM in the presence of fibronectin perhaps generates a signaling cascade that leads to expression of MMP-9 and activation of MMP-2 in culture supernatants within 2 h. The signaling pathway activated is probably the FAK/ERK pathway.

Loss of Merlin induces metabolomic adaptation that engages dependence on Hedgehog signaling.

The tumor suppressor protein Merlin is proteasomally degraded in breast cancer. We undertook an untargeted metabolomics approach to discern the global metabolomics profile impacted by Merlin in breast cancer cells. We discerned specific changes in glutathione metabolites that uncovered novel facets of Merlin in impacting the cancer cell metabolome. Concordantly, Merlin loss increased oxidative stress causing aberrant activation of Hedgehog signaling. Abrogation of GLI-mediated transcription activity compromised the aggressive phenotype of Merlin-deficient cells indicating a clear dependence of cells on Hedgehog signaling. In breast tumor tissues, GLI1 expression enhanced tissue identification and discriminatory power of Merlin, cumulatively presenting a powerful substantiation of the relationship between these two proteins. We have uncovered, for the first time, details of the tumor cell metabolomic portrait modulated by Merlin, leading to activation of Hedgehog signaling. Importantly, inhibition of Hedgehog signaling offers an avenue to target the vulnerability of tumor cells with loss of Merlin.

Culture of human cervical cancer cells, SiHa, in the presence of fibronectin activates MMP-2.

PURPOSE: Several studies indicate that integrin receptors are involved in the regulation of matrix metalloproteinase (MMP) expression. Integrin-ECM ligand interaction leads to phosphorylation of focal adhesion kinase (FAK) and activation of mitogen activated protein kinase pathways. In this present communication, we cultured human cervical cancer cells, SiHa, in the presence of fibronectin to study fibronectin-integrin mediated modulation of MMP activity. METHODS: SiHa cells were cultured in serum-free medium (SFCM) in the presence of fibronectin, SFCM was collected and gelatin zymography was performed. Western blot, RT-PCR and immunocytochemistry were performed with SiHa cells cultured in the presence of fibronectin. RESULTS: The culture of SiHa cells in the presence of 50 microg/1.5 ml fibronectin led to expression of pro-MMP-9 and activation of MMP-2 within 2 h. When cells were treated with ERK inhibitor (PD98059) and grown in the presence of fibronectin MMP-2 activation was partially inhibited, but when cells were treated with PI-3K inhibitor (LY294002) and grown in the presence of fibronectin MMP-2 activation was appreciably reduced. Tyrosine phosphorylation of FAK, PI-3K and ERK and nuclear trafficking of ERK were increased in SiHa cells grown in the presence of fibronectin. Increased MT1-MMP mRNA expression and processing of MT1-MMP were also observed in SiHa cells grown in the presence of fibronectin. CONCLUSIONS: Our findings indicate that the culture of SiHa cells in SFCM in the presence of fibronectin perhaps generates a signalling cascade which leads to the expression of pro-MMP-9 and the activation of MMP-2 within 2 h. The signalling pathways activated seem to be the FAK/ERK/PI-3K pathway.

Loss of tumor suppressor Merlin results in aberrant activation of Wnt/ß-catenin signaling in cancer.

The expression of the tumor suppressor Merlin is compromised in nervous system malignancies due to genomic aberrations. We demonstrated for the first time, that in breast cancer, Merlin protein expression is lost due to proteasome-mediated elimination. Immunohistochemical analysis of tumor tissues from patients with metastatic breast cancer revealed characteristically reduced Merlin expression. Importantly, we identified a functional role for Merlin in impeding breast tumor xenograft growth and reducing invasive characteristics. We sought to determine a possible mechanism by which Merlin accomplishes this reduction in malignant activity. We observed that breast and pancreatic cancer cells with loss of Merlin show an aberrant increase in the activity of ß-catenin concomitant with nuclear localization of ß-catenin. We discovered that Merlin physically interacts with ß-catenin, alters the sub-cellular localization of ß-catenin, and significantly reduces the protein levels of ß-catenin by targeting it for degradation through the upregulation of Axin1. Consequently, restoration of Merlin inhibited ß-catenin-mediated transcriptional activity in breast and pancreatic cancer cells. We also present evidence that loss of Merlin sensitizes tumor cells to inhibition by compounds that target ß-catenin-mediated activity. Thus, this study provides compelling evidence that Merlin reduces the malignant activity of pancreatic and breast cancer, in part by suppressing the Wnt/ß-catenin pathway. Given the potent role of Wnt/ß-catenin signaling in breast and pancreatic cancer and the flurry of activity to test ß-catenin inhibitors in the clinic, our findings are opportune and provide evidence for Merlin in restraining aberrant activation of Wnt/ß-catenin signaling.

Distribution pattern of ambient cadmium in wetland ponds distributed along an industrial complex.

Water and sediment samples collected from 18 wetland ponds within and outside industrial areas were examined for cadmium concentration and water quality parameters during the period of January to July 1996. The Cd contents in gill, liver, mantle and shell of freshwater mussel (Lamellidens marginalis) as well as leaves and roots of water hyacinth Eichhornia those occurred in these ponds were also estimated. Cd concentration ranged from 0.006 to 0.7025 mg/l in water and from 7 to 77 microg/gdw in sediments of all the ponds investigated. The amount of Cd occurring in water and sediment was much higher in concentrations in the ponds located in Captain Bheri and Mudiali farm close to industrial areas, compared to remaining ponds located outside the industrial belt. Lamellidens marginalis procured from Mudiali and Captain Bheri ponds showed regardless of size, tissue and season of collection significantly higher Cd concentration than did those from other ponds. Likewise, tissue Cd in Eichhornia collected from Mudiali pond was as high as 125-152 microg/gdw in root and 21-63 microg/gdw in leaves compared to 40-108 microg/gdw in root and 9-43 microg/gdw in leaves in the remaining ponds. Seasonal variability of Cd was clear-cut; the concentration was relatively higher in water and sediment in all ponds during summer than during monsoon season or winter. Size-wise, smaller groups showed the highest concentrations of Cd in all tissues of Lamellidens compared with medium and large size groups. Concentration factor for all tissues of Lamellidens regardless of size and season, was inversely proportional with the ambient Cd concentrations. Concentration factor estimated for all tissues in all ponds and all seasons was in the order: liver>gill>shell>mantle. As all ponds located outside the industrial belt showed Cd concentrations ranging from 0.006 to 0.049 mg/l, it is suggested that these wetlands do not pose serious risk to the environment.

In situ cadmium reclamation by freshwater bivalve Lamellidens marginalis from an industrial pollutant-fed river canal.

The biofilter potential of the freshwater bivalve, Lamellidens marginalis was examined in cage experiments conducted in a river canal (Ichhapore, 24-Parganas, West Bengal, India) receiving industrial effluents from steel and metal factories as well as from an ordinance factory. Cadmium is one of the major contaminants in this river canal. Lamellidens collected from pollution free natural ponds, were sorted into three size groups (large: 59+/-3.2 g, 10+/-2.3 cm; medium: 30+/-2 g, 6+/-1.7 cm and small: 13+/-1.5 g, 4+/-1.2 cm) were held in cages at three different sites along a cadmium concentration gradient. Concentrations of cadmium were measured from water, sediment and different tissues of Lamellidens at weekly intervals using atomic absorption spectrophotometric methods. Cadmium uptake by Lamellidens in all media were highly concentration dependent in both summer and winter months. For all three size groups, cadmium uptake was maximum in the gills at the beginning of experiment, and liver at the later phase. Cadmium uptake was maximum in the small bivalves and minimum in the large bivalves groups. Cadmium uptake was 11-67% higher during summer than during the monsoon season for all tissues and size groups. Estimation of concentration factor revealed that tissues were saturated with cadmium during the 13-14th week after Lamellidens introduction during summer, but remained unsaturated during the monsoon season. It is concluded that Lamellidens might be considered as an efficient biofilter for reclamation of aquatic environment having sub-lethal concentrations of cadmium.

Oxygen uptake and filtration rate as animal health biomarker in Lamellidens marginalis (Lamarck).

The freshwater bivalve, L. marginalis was experimentally exposed to 10 and 30 ppm concentrations of CdCl2 to examine filtration rate, oxygen uptake and glycogen level of liver and gills for health assessment for their reuse in the reclamation of cadmium intoxicated environments. In situ experiment was also performed for better appraisal of the filtration rate in the lake. Oxygen uptake in the treated group exceeded that of control by 15-22% during the early 24 hr after cadmium exposure, but followed an essential decline (23-30%) thereafter. The reduction of filtration rate ranged from 12-62% in laboratory to 83-85% in field trials. At the tissue level, glycogen content was reduced by 61-72% in liver and 52-63% in gill. In both tissues, glycogen content was inversely proportional to the cadmium contents of the animal. Critical appraisal of data suggests that the threshold values of cadmium in gill and liver were 50-80 microg/g dw for oxygen uptake and 50-60 microg/g for filtration rate because of marked reduction of these parameters beyond the values of cadmium. It is concluded that filtration rate, oxygen uptake of the freshwater bivalve, L. marginalis can be used as biomarker for animal health assessment and for possible reuse of the stock animals.