Effect of Cichorium intybus seeds supplementation on the markers of glycemic control, oxidative stress, inflammation, and lipid profile in type 2 diabetes mellitus: A randomized, double-blind placebo study.

Diabetes mellitus is associated with increased levels of inflammation and oxidative stress in patients. The aim of the present study was to test the hypothesis that aqueous extract of Cichorium intybus seeds (AECIS) would have add-on beneficial effect in type 2 diabetes mellitus (T2DM). In this double-blind randomized clinical study, 150 subjects were enrolled to assess the add-on efficacy and safety of AECIS in T2DM patients. The subjects were randomized (1:1) to the AECIS (n = 51) and placebo (n = 49) groups. The subjects in both groups continued to take prescribed doses of metformin. The standardization of AECIS was carried out by liquid chromatography-mass spectrometry and phytochemical analysis. The mean hemoglobin A1c (HbA1c) level in the AECIS and placebo groups at baseline was 8.6% and 8.5%, respectively. Mean values of HbA1c at the end of 12 weeks of intervention were 7.42% in the AECIS group (a reduction of 1.18% from baseline) and 8.4% in the placebo group (mean reduction of 0.1% from baseline). Besides, significant reduction in inflammation, oxidative stress, and hypertriglyceridemia was seen in the AECIS group (p < .05). The study shows for the first time that AECIS supplementation ameliorates the disease progression and it is beneficial as a potential adjunct dietary supplement for the management of T2DM.

Assessment of cellular and serum proteome from tongue squamous cell carcinoma patient lacking addictive proclivities for tobacco, betel nut, and alcohol: Case study.

The intriguing molecular pathways involved in oral carcinogenesis are still ambiguous. The oral squamous cell carcinoma (OSCC) ranks as the most common type constituting more than 90% of the globally diagnosed oral cancers cases. The elevation in the OSCC incidence rate during past 10 years has an alarming impression on human healthcare. The major challenges associated with OSCC include delayed diagnosis, high metastatic rates, and low 5-year survival rates. The present work foundations on reverse genetic strategy and involves the identification of genes showing expressional variability in an OSCC case lacking addictive proclivities for tobacco, betel nut, and/or alcohol, major etiologies. The expression modulations in the identified genes were analyzed in 16 patients comprising oral pre-cancer and cancer histo-pathologies. The genes SCCA1 and KRT1 were found to down regulate while DNAJC13, GIPC2, MRPL17, IG-Vreg, SSFA2, and UPF0415 upregulated in the oral pre-cancer and cancer pathologies, implicating the genes as crucial players in oral carcinogenesis.

Novel mutations in transthyretin gene associated with hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the major health problems with increasing incidence worldwide. We report the elevation in transthyretin (TTR) expression following HCC induction using diethylnitrosamine (DEN) and 2-aminoacetylfluorine (2-AAF) in male Wistar rats. The increase in its expression took place at very early stage and remained elevated throughout HCC progression. The analysis of TTR gene in HCC bearing rats revealed four novel mutations that alter three amino acids at positions 61, 100, and 115. While these mutations do not directly affect the binding of TTR to thyroxine (T4 ), the mutation in amino acid 115 interferes with TTR tetramer formation that leads to its accumulation. Further, the mutated TTR is unable to cleave C-terminal of apolipoprotein A1 (APOA1) that results in abnormal lipid metabolism. This has correlation with development of liver cirrhosis and HCC. Furthermore, the mutated TTR seems to have potential as biomarker for early detection of HCC.

Virus-like particles derived from Pichia pastoris-expressed dengue virus type 1 glycoprotein elicit homotypic virus-neutralizing envelope domain III-directed antibodies.

BACKGROUND: Four antigenically distinct serotypes (1-4) of dengue viruses (DENVs) cause dengue disease. Antibodies to any one DENV serotype have the potential to predispose an individual to more severe disease upon infection with a different DENV serotype. A dengue vaccine must elicit homotypic neutralizing antibodies to all four DENV serotypes to avoid the risk of such antibody-dependent enhancement in the vaccine recipient. This is a formidable challenge as evident from the lack of protective efficacy against DENV-2 by a tetravalent live attenuated dengue vaccine that has completed phase III trials recently. These trial data underscore the need to explore non-replicating subunit vaccine alternatives. Recently, using the methylotrophic yeast Pichia pastoris, we showed that DENV-2 and DENV-3 envelope (E) glycoproteins, expressed in absence of prM, implicated in causing severe dengue disease, self-assemble into virus-like particles (VLPs), which elicit predominantly virus-neutralizing antibodies and confer significant protection against lethal DENV challenge in an animal model. The current study extends this work to a third DENV serotype. RESULTS: We cloned and expressed DENV-1 E antigen in P. pastoris, and purified it to near homogeneity. Recombinant DENV-1 E underwent post-translational processing, namely, signal peptide cleavage and glycosylation. Purified DENV-1 E self-assembled into stable VLPs, based on electron microscopy and dynamic light scattering analysis. Epitope mapping with monoclonal antibodies revealed that the VLPs retained the overall antigenic integrity of the virion particles despite the absence of prM. Subtle changes accompanied the efficient display of E domain III (EDIII), which contains type-specific neutralizing epitopes. These VLPs were immunogenic, eliciting predominantly homotypic EDIII-directed DENV-1-specific neutralizing antibodies. CONCLUSIONS: This work demonstrates the inherent potential of P. pastoris-expressed DENV-1 E glycoprotein to self-assemble into VLPs eliciting predominantly homotypic neutralizing antibodies. This work justifies an investigation of the last remaining serotype, namely, DENV-4, to assess if it also shares the desirable vaccine potential manifested by the remaining three DENV serotypes. Such efforts could make it possible to envisage the development of a tetravalent dengue vaccine based on VLPs of P. pastoris-expressed E glycoproteins of the four DENV serotypes.

A small molecule inhibitor of dengue virus type 2 protease inhibits the replication of all four dengue virus serotypes in cell culture.

BACKGROUND: Dengue has emerged as the most significant of arboviral diseases in the 21st century. It is endemic to >100 tropical and sub-tropical countries around the world placing an estimated 3.6 billion people at risk. It is caused by four genetically similar but antigenically distinct, serotypes of dengue viruses. There is neither a vaccine to prevent nor a drug to treat dengue infections, at the present time. The major objective of this work was to explore the possibility of identifying a small molecule inhibitor of the dengue virus protease and assessing its ability to suppress viral replication in cultured cells. METHODS: We cloned, expressed and purified recombinant dengue virus type 2 protease. Using an optimized and validated fluorogenic peptide substrate cleavage assay to monitor the activity of this cloned dengue protease we randomly screened ~1000 small molecules from an 'in-house' library to identify potential dengue protease inhibitors. RESULTS: A benzimidazole derivative, named MB21, was found to be the most potent in inhibiting the cloned protease (IC50 = 5.95 µM). In silico docking analysis indicated that MB21 binds to the protease in the vicinity of the active site. Analysis of kinetic parameters of the enzyme reaction suggested that MB21 presumably functions as a mixed type inhibitor. Significantly, this molecule identified as an inhibitor of dengue type 2 protease was also effective in inhibiting each one of the four serotypes of dengue viruses in infected cells in culture, based on analysis of viral antigen synthesis and infectious virus production. Interestingly, MB21 did not manifest any discernible cytotoxicity. CONCLUSIONS: This work strengthens the notion that a single drug molecule can be effective against all four dengue virus serotypes. The molecule MB21 could be a potential candidate for 'hit-to-lead' optimization, and may pave the way towards developing a pan-dengue virus antiviral drug.

1,8-Naphthyridine Derivatives: A Review of Multiple Biological Activities.

The 1,8-naphthyridine group of compounds have gained special attention of researchers on account of their demonstrating a variety of interesting biological activities. A wide range of biological properties establishes them as potent scaffolds in therapeutic and medicinal research. The broad spectrum of activities primarily includes antimicrobial, antiviral, anticancer, anti-inflammatory, and analgesic activities. 1,8-Naphthyridine derivatives have also exhibited potential applications in neurological disorders such as Alzheimer's disease, multiple sclerosis, and depression. In addition, these synthetic derivatives have been found to possess activities such as anti-osteoporotic (α(v)ß(3) antagonists), anti-allergic, antimalarial, gastric antisecretory, bronchodilator, anticonvulsant, anti-hypertensive, platelet aggregation inhibition, anti-oxidant, EGFR inhibition, protein kinase inhibition, ionotropic agent, ß-3 antagonist, MDR modulator, adenosine receptor agonist, adrenoceptor antagonist, and pesticide activities. In spite of the widespread application of the 1,8-naphythyridine scaffolds, only a limited number of review articles are available till date. In this review, we attempt to compile and discuss the key data available in the literature for the multiple biological activities of 1,8-naphthyridine derivatives, in a chronological manner. This review compilation (with 199 references) may be helpful in understanding the diverse biological properties of 1,8-naphthyridines and provide insights into their mechanism of action. This may direct future research in the synthesis of new derivatives and exploring this scaffold for other possible biological activities.

Ochratoxin A-induced cell proliferation and tumor promotion in mouse skin by activating the expression of cyclin-D1 and cyclooxygenase-2 through nuclear factor-kappa B and activator protein-1.

Our prior studies have indicated that ochratoxin A (OTA), a mycotoxin, has skin tumor initiating activity. In the present investigation, skin tumor promoting activity of OTA and the mechanism/(s) involved therein was undertaken. A single topical application of OTA (100 nmol/mouse) caused significant enhancement in short-term markers of skin tumor promotion such as ornithine decarboxylase activity, DNA synthesis, hyperplasia as well as expression of cyclin-D1 and COX-2 in mouse skin. In a two-stage mouse skin tumorigenesis protocol, twice-weekly exposure of OTA (50 nmol/mouse) to 7,12-dimethylbenz[α]anthracene (120 nmol/mouse) initiated mice skin for 24 weeks leads to tumor formation. Further, exposure of primary murine keratinocytes (PMKs) with non-cytotoxic dose of OTA (5.0 µM) caused (i) significant enhancement of DNA synthesis, (ii) enhanced phosphorylation and subsequent activation of epidermal growth factor receptor (EGFR) and its downstream signaling pathways viz Akt, ERK1/2, p38 and JNK mitogen-activated protein kinases (MAPKs), (iii) overexpression of c-jun, c-fos, cyclin-D1 and COX-2 and (iv) increased binding of nuclear factor-kappaB (NF-κB) and AP-1 transcription factors to the promoter region of cyclin-D1 and COX-2 genes. It was also observed that knocking down the messenger RNA expression of NF-κB, c-jun, c-fos, cyclin-D1 and COX-2 results in significant inhibition in OTA-induced PMKs proliferation. These results suggest that OTA has cell proliferative and tumor-promoting potential in mouse skin, which involves EGFR-mediated MAPKs and Akt pathways along with NF-κB and AP-1 transcription factors and that cyclin-D1 and COX-2 are the target genes responsible for tumor-promoting activity of OTA.

Dengue-specific subviral nanoparticles: design, creation and characterization.

BACKGROUND: Dengue is today the most significant of arboviral diseases. Novel tools are necessary to effectively address the problem of dengue. Virus-like particles (VLP) offer a versatile nanoscale platform for developing tools with potential biomedical applications. From the perspective of a potentially useful dengue-specific tool, the dengue virus envelope protein domain III (EDIII), endowed with serotype-specificity, host receptor recognition and the capacity to elicit virus-neutralizing antibodies, is an attractive candidate. METHODS: We have developed a strategy to co-express and co-purify Hepatitis B virus surface (S) antigen in two forms: independently and as a fusion with EDIII. We characterized these physically and functionally. RESULTS: The two forms of the S antigen associate into VLPs. The ability of these to display EDIII in a functionally accessible manner is dependent upon the relative levels of the two forms of the S antigen. Mosaic VLPs containing the fused and un-fused components in 1:4 ratio displayed maximal functional competence. CONCLUSIONS: VLPs armed with EDIII may be potentially useful in diagnostic, therapeutic and prophylactic applications.

Cytochrome P450 2A isoenzymes in freshly prepared blood lymphocytes isolated from rats and validation as a biomarker for clinical studies in humans.

1. The present study aimed to identify the expression of carcinogen metabolizing cytochrome P4502A (CYP2A) isoenzymes in freshly prepared rat peripheral blood lymphocytes (PBL) isolated from adult rats and investigate similarities in the regulation of lymphocyte CYP2A-isoenzymes with the tissue enzyme. 2. qRT-PCR studies demonstrated significant constitutive mRNA expression of CYP2A-isoenzymes in PBL isolated from male and female rats which further increases significantly after pretreatment with nicotine or 3-methylcholanthrene (MC) indicating responsiveness of CYP2A-isoenzymes in PBL. This increase in the CYP2A expression was associated with an increase in the protein expression and CYP2A3-dependent coumarin hydroxylase (COH) activity in PBL. 3. Clinical studies further demonstrated significant increase in the expression of CYP2A6 and associated enzyme activity in PBL isolated from lung cancer patients. Our data thus provided evidence for similarities in the regulation of carcinogen metabolizing CYP2A-isoenzymes in PBL with the tissue enzymes. Further, responsiveness of blood CYP2A6 in human blood lymphocytes isolated from lung cancer patients has led us to suggest that associating expression profiles of CYP2A6 and other polycyclic aromatic hydrocarbons (PAH)-responsive CYPs in PBL with the genotyping data could lead to the development of a possible screen to monitor and predict environment-induced diseases and toxicity in humans.

Ethanol induced induction of cytochrome P450 2E1 and activation of mitogen activated protein kinases in peripheral blood lymphocytes.

Cytochrome P450 2E1 (CYP2E1), which induces oxidative stress that leads to alcohol-mediated toxicity in liver, is expressed in peripheral blood lymphocytes. To validate blood lymphocyte CYP2E1 as a biomarker of alcohol-induced diseases, studies were initiated to investigate similarities in CYP2E1 induction and associated cell signalling pathways in freshly prepared blood lymphocytes with the liver in rats exposed to alcohol. Acute or chronic treatment of ethanol produced significant increase in enzyme activity and lipid peroxidation in blood lymphocytes. As observed in liver, this increase was associated with the enrichment of CYP2E1 protein and mRNA. Similar pattern of increase in the mRNA and protein expression of c-jun and c-fos was also observed in blood lymphocytes and liver. Acute exposure to ethanol activated ERK and JNK MAP kinases and c-jun in the blood lymphocytes and liver. The present data demonstrating similarities in the induction of CYP2E1 and lipid peroxidation and activation of MAP Kinases in blood lymphocytes with liver after acute or chronic exposure of ethanol have suggested that blood lymphocytes could be used to monitor ethanol induced CYP2E1 induction and associated oxidative stress in liver.

Inhibitory effect of antibodies against human chorionic gonadotropin on the growth of colorectal tumour cells.

Human chorionic gonadotropin (hCG) was initially believed to be secreted exclusively by the embryo with its primary function being "rescue" of the corpus luteum. However, recently it has been found that the hormone (or its individual subunits) is also secreted by many cancers and that in many cases secretion is associated with poor patient prognosis. In this study, we assessed the presence of hCG in colorectal cancer cells (CCL-253) and evaluated the anti-tumour effects of anti-hCG antibodies in vitro and in vivo. Anti-hCG antibodies were reactive with CCL-253, as revealed by confocal immunoflourescence microscopy; both cell surface and intracellular expression were observed. Western blot analysis showed that antibodies appeared to interact with several moieties, indicating a level of cross-reactivity. Anti-hCG antiserum specifically reduced the viability of tumor cells and the addition of complement increased in vitro anti-tumor effects. In nude mice implanted with CCL-253 cells, administration of anti-hCG antiserum caused a significant reduction in tumor volume; all treated animals survived, while mortality was observed in control animals. Results suggest that anti-hCG antibodies can mediate significant anti-tumor activity both in vitro and in vivo and lend support to the rationale of anti-hCG immunization in the therapy of gonadotropin- sensitive cancers.

Novel derivative of benzofuran induces cell death mostly by G2/M cell cycle arrest through p53-dependent pathway but partially by inhibition of NF-kappaB.

The Dracaena resin is widely used in traditional medicine as an anticancer agent, and benzofuran lignan is the active component. In this report, we provide evidence that the synthetic derivative of benzofuran lignan (Benfur) showed antitumor activities. It induced apoptosis in p53-positive cells. Though it inhibited endotoxin-induced nuclear factor kappaB (NF-kappaB) activation in both p53-positive and -negative cells, the activation of caspase 3 was observed in p53-positive cells. It showed partial cell death effect in both p53-positive and -negative cells through inhibition of NF-kappaB. Cell cycle analysis using flow cytometry showed that treatment with this novel benozofuran lignan derivative to Jurkat T-cells, but not U-937 cells, resulted in a G2/M arrest in a dose- and time-dependent manner. It increased amounts of p21, p27, and cyclin B, but not phospho-Rb through p53 nuclear translocation in Jurkat T-cells, but not in U-937 cells. It inhibited amounts of MDM2 (murine double minute 2) by repressing the transcription factor Sp1, which was also proved in silico. It induced cell death in tumor cells, but not in primary T-cells. Overall, our data suggest that Benfur-mediated cell death is partially dependent upon NF-kappaB, but predominantly dependent on p53. Thus, this novel benzofuran lignan derivative can be effective chemopreventive or chemotherapeutic agent against malignant T-cells.

Expression profiling of toxicity pathway genes by real-time PCR array in cypermethrin-exposed mouse brain.

Cypermethrin, a type II pyrethroid, has been shown to exert genotoxic effects in the central nervous system of non-target species such as mouse and Drosophila. To unravel the gene expression of toxicity-related pathways in cypermethrin-exposed Swiss albino mouse brain, transcriptional profiling was carried out through pathway-focused real-time PCR arrays (DNA damage signaling, oxidative stress/antioxidants, and stress/toxicity pathways). The real-time PCR array data revealed a significant (p < 0.05) modulation in transcript levels of 61 genes involved in DNA replication and repair, apoptosis, cell cycle, oxidative stress, and toxicity pathways. Cypermethrin also produced oxidative stress in brain, as was evident by a significant (p < 0.05) elevation (66%) in lipid peroxidation and reduction of glutathione (GSH) content (10.6%) as well as catalase activity (56.7%). The results demonstrate that cypermethrin alters the expression of stress- and toxicity-related genes as well as induces oxidative stress which may lead to DNA damage. These observations also point to complex metabolic networks involved in genotoxic manifestations by cypermethrin.

Novel caffeic acid ester derivative induces apoptosis by expressing FasL and downregulating NF-KappaB: Potentiation of cell death mediated by chemotherapeutic agents.

In the present study, we demonstrate the biological activity of esterified caffeic acid with methyl vanillate also termed as caffeic acid methyl vanillate ester (CAMVE). CAMVE potentiates TNF-induced cell death as analyzed by cell viability assay and blocks inflammatory stimuli-induced nuclear transcription factor kappaB (NF-kappaB) activation and NF-kappaB-dependent genes expression. CAMVE-mediated inhibition of NF-kappaB or induction of cell death is not cell type specific. CAMVE inhibits cell proliferation by inhibiting G1 to S phase progression. It suppresses TNF-induced Bcl-2 expression and potentiates chemotherapeutic agents-mediated cell death. CAMVE enhances intracellular free Ca(2+) and thereby activates calcineurin. Calcineurin, in turns, activates nuclear transcription factor NF-AT and its dependent genes such as FasL, which induces cell death. The data demonstrate that CAMVE is one of the combinatorial products, which is able to inhibit NF-kappaB regulated genes and cell proliferation. The combinatorial synthesis of novel caffeic ester derivatives can be a useful approach to generate potent chemotherapeutic agents and designing CAMVE as potent therapeutic agent for combination therapy may be useful to treat tumors.

Expression of alphaVbeta3 integrin in rat endometrial epithelial cells and its functional role during implantation.

The alpha(V)beta(3) integrin as a marker of endometrial receptivity has been well established in human and other mammalian species; however, its expression is still not known in rats. Our objective was to establish the expression of alpha(V)beta(3) integrin as a marker of endometrial receptivity in rat and to further prove its role in implantation by function-blocking studies in this species. Immunocytochemical, immunohistochemical and flow-cytometric studies were performed in rat endometrial epithelial cells (EEC) to demonstrate the expression of alpha(V)beta(3) integrin during non-receptive, pre-receptive and receptive phases of the uterus. Results revealed positive immunocytochemical staining for alpha(v) and beta(3) subunits on the surface of EEC of days 4 and 5p.c. (post-coitum), but the intensity was higher in cells of day 5p.c. Flow-cytometric study revealed higher level of alpha(V)beta(3) on day 5p.c. as compared to day 4p.c. and non-pregnant animals. Immunohistochemical analysis of uterine tissue also revealed that the alpha(V)beta(3) expression in LE was higher on day 5p.c. morning as compared to that observed on day 4p.c. In addition, the expression of beta(3) subunit was not evident in rats receiving ormeloxifene, an agent known to inhibit the uterine receptivity. Immunoblotting experiments also revealed higher expression of uterine beta(3) on day 5p.c. On day 6, expression of beta(3) was high in implantation sites than on inter-implantation sites. In immature ovariectomized rats, alpha(V)beta(3) was up-regulated by progesterone and by a combination of estrogen and progesterone. The expression of alpha(V)beta(3) was also up-regulated in EEC co-cultured with blastocysts. All the agents used for function-blocking studies showed significant reduction in the number of implantation sites in treated horn as compared to sham control horn. The present study has successfully demonstrated the expression of alpha(V)beta(3) in rat EEC as a marker of endometrial receptivity and showed that this molecule is indispensable for the process of implantation in this species.

Elevated Expression of A-Raf and FA2H in Hepatocellular Carcinoma is Associated with Lipid Metabolism Dysregulation and Cancer Progression.

BACKGROUND: Identification of events leading to hepatocellular carcinoma (HCC) progression is essential for understanding its pathophysiology. The aims of this study are to identify and characterize differentially expressed proteins in serum of HCC-bearing rats and the corresponding controls during cancer initiation, progression and tumorigenesis. METHODS: Chemical carcinogens, N-Nitrosodiethylamine and 2-aminoacetylfluorine are administered to induce HCC to male Wistar rats. The 2D-Electrophoresis and PD-Quest analyses are performed to identify several differentially expressed proteins in serum of HCC-bearing animals. These proteins are further characterized by MALDI-TOF-MS/MS analyses. Using pathwaylinker a HCC-specific network is analyzed among the MALDITOF- MS/MS characterized proteins and their interactors. RESULTS: Carcinogen administration caused inflammation leading to liver injury and HCC development. Liver inflammation was confirmed by increase in the levels of TNF-α and IL-6 in carcinogen treated rats. We report significant increase in expression of two differentially expressed proteins, namely, A-Raf and Fatty Acid 2- Hydroxylase (FA2H), at early stage of HCC initiation, during its progression and at tumor stage. Real-time PCR analysis of mRNA for these proteins confirmed up-regulation of their transcripts. Further, we validated our experimental data with sera of clinically confirmed liver cancer patients. CONCLUSION: The study suggests that FA2H and A-Raf play a major role in the progression of HCC.

Interactome Analysis of the Differentially Expressed Proteins in Uterine Leiomyoma.

BACKGROUND: Recent advances in proteomics present enormous opportunities to discover proteome related disparities and thus understanding the molecular mechanisms related to a disease. Uterine leiomyoma is a benign monoclonal tumor, located in the pelvic region, and affecting 40% of reproductive aged female. OBJECTIVE: Identification and characterization of the differentially expressed proteins associated with leiomyogenesis by comparing uterine leiomyoma and normal myometrium. METHODS: Paired samples of uterine leiomyoma and adjacent myometrium retrieved from twenty-five females suffering from uterine leiomyoma (n=50) were submitted to two-dimensional electrophoresis (2-DE), matrixassisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and to reverse transcription polymerase chain reaction (RT-PCR). RESULTS: Comparison of protein patterns revealed seven proteins with concordantly increased spot intensities in leiomyoma samples. E3 ubiquitin-protein ligase MIB2 (MIB2), Mediator of RNA polymerase II transcription subunit 10 (MED10), HIRA-interacting protein (HIRP3) and Fatty acid binding protein brain (FABP7) were found to be upregulated. While, Biogenesis of lysosome-related organelles complex 1 subunit 2 (BL1S2), Shadow of prion protein (SPRN) and RNA binding motif protein X linked like 2 (RMXL2) were found to be exclusively present in leiomyoma sample. The expression modulations of the corresponding genes were further validated which corroborated with the 2-DE result showing significant upregulation in leiomyoma. We have generated a master network showing the interactions of the experimentally identified proteins with their close neighbors and further scrutinized the network to prioritize the routes leading to cell proliferation and tumorigenesis. CONCLUSION: This study highlights the importance of identified proteins as potential targets for therapeutic purpose. This work provides an insight into the mechanism underlying the overexpression of the proteins but warrants further investigations.

HCC Specific Protein Network Involving Interactions of EGFR with A-Raf and Transthyretin: Experimental Analysis and Computational Biology Correlates.

BACKGROUND: The network interactions link human disease proteins to regulatory cellular pathways leading to better understanding of protein functions and cellular processes. Revealing the network of signaling pathways in cancer through protein-protein interactions at molecular level enhances our understanding of Hepatocellular Carcinoma (HCC). OBJECTIVE: A rodent model for study of HCC was developed to identify differentially expressed proteins at very early stage of cancer initiation and throughout its progression. METHODOLOGY: HCC was induced by administrating N-Nitrosodiethylamine (DEN) and 2-aminoacetylfluorine (2-AAF) to male Wistar rats. Proteomic approaches such as 2D-Electrophoresis, PD-Quest, MALDI-TOF-MS and Western blot analyses have been used to identify, characterize and validate the differentially expressed proteins in HCC-bearing animals vis-a-vis controls. RESULTS: The step-wise analysis of morphological and histological parameters revealed HCC induction and tumorigenesis at 1 and 4 months after carcinogen treatment, respectively. We report a novel protein network of 735 different proteins out of which eight proteins are characterized by MALDI-TOF-MS analysis soon after HCC was chemically induced in rats. We have analyzed four different novel routes representing the association of experimentally identified proteins with HCC progression. CONCLUSION: The study suggests that A-Raf, transthyretin and epidermal growth factor receptor play major role in HCC progression by regulating MAPK signaling pathway and lipid metabolism leading to continuous proliferation, neoplastic transformation and tumorigenesis.

Structural and Biochemical Characterization of Apicomplexan Inorganic Pyrophosphatases.

Inorganic pyrophosphatases (PPase) participate in energy cycling and they are essential for growth and survival of organisms. Here we report extensive structural and functional characterization of soluble PPases from the human parasites Plasmodium falciparum (PfPPase) and Toxoplasma gondii (TgPPase). Our results show that PfPPase is a cytosolic enzyme whose gene expression is upregulated during parasite asexual stages. Cambialistic PfPPase actively hydrolyzes linear short chain polyphosphates like PPi, polyP3 and ATP in the presence of Zn2+. A remarkable new feature of PfPPase is the low complexity asparagine-rich N-terminal region that mediates its dimerization. Deletion of N-region has an unexpected and substantial effect on the stability of PfPPase domain, resulting in aggregation and significant loss of enzyme activity. Significantly, the crystal structures of PfPPase and TgPPase reveal unusual and unprecedented dimeric organizations and provide new fundamental insights into the variety of oligomeric assemblies possible in eukaryotic inorganic PPases.

Priming with DNA Enhances Considerably the Immunogenicity of hCG ß-LTB Vaccine.

PROBLEM: Necessity to elicit antibody response above the protective threshold titres by sexually active women immunized to prevent pregnancy. METHOD OF STUDY: Recombinant hCGß-LTB vaccine expressed as both DNA and protein. Balb C mice employed for testing immunogenicity. RESULTS: Necessity to give three primary injections of the vaccine to elicit proper antibody response. Immunization twice with DNA form of the vaccine at fortnightly interval followed by the protein elicits a distinctly higher antibody response than proteinic vaccine alone. Antibodies generated are bio-effective against hCG. CONCLUSION: Immunization with the DNA form of the recombinant hCGß-LTB vaccine twice at fortnightly interval followed by the proteinic form of the vaccine induces distinctly higher antibody response.