Synthesis of aminomethyl linked (+)-usnic acid derivatives via the Mannich reaction and evaluation of their biological activities.

(+)-Usnic acid (UA), a natural dibenzofuran derivative, abundantly produced by lichens and possess wide number of biomedical applications including antibacterial, anti-inflammatory, anti-oxidant and anticancer activities. In the present study, as series of usnic acid derivatives (3a-3i) were synthesised using Mannich reaction assessed for their antioxidant, α-glucosidase, and anticancer activities. The in vitro antioxidant activity showed that compound 3d displayed potent antioxidant activity by scavenging the activities of DPPH and ABTS+. The compounds 3d and 3e showed potent cytotoxic activity against HepG2 cancer cells by arresting the cell cycle at S phase and regulating the Bax/BcL2 expression and subsequently induce the apoptosis. Overall, the results clearly indicated that (+)-usnic acid derivatives bearing secondary amines are useful scaffolds for the development of drug candidates for treatment of oxidative stress mediated cancer and metabolic disorders.

Synthesis and biological evaluation of 1,2,3-triazole hybrids of 4-methoxy ethyl cinnamate isolated from Hedychium spicatum (Sm) rhizomes: identification of antiproliferative lead actives against prostate cancer.

A series of 1, 2, 3- triazole hybrids (9a-9n) were synthesised from major phenolic constituent, 4-methoxy ethyl cinnamate (5) isolated from rhizomes of Hedychium spicatum (Sm), a traditional medicinal plant used in variety of disease conditions. All the synthesised analogues were tested for their in vitro antiproliferative potential against HCT 116 (colon cancer), A549 (lung cancer), DU-145 (prostate cancer), Hep G2 (hepatoma) and HEK-293 (normal) cell lines. Among the compounds tested, compounds 9i and 9k potently arrested proliferation of DU-145 (prostate cancer) cell line. Compound 9i displayed 20 times better antiproliferative potential than parent compound and almost identical inhibitory activity to that of the standard drug, doxorubicin. The flow cytometric analysis revealed that 9i arrested cells in G2/M phase of cell cycle and induced apoptosis. Overall, the hybrid derivative 9i was found to be a potential antiproliferative lead against prostate cancer.

A 1.6 Mb region on SSC2 is associated with antibody response to classical swine fever vaccination in a mixed pig population.

Classical Swine Fever (CSF) is a contagious viral disease of pigs which is endemic in several parts of the world, including India. Prophylactic vaccination using live attenuated vaccine is the preferred method of control. However, there is significant inter-individual variation in the antibody response to vaccination. In this study, we measured the E2 antibody blocking percentage after 21 days of CSF vaccination in a mixed pig population consisting of Landrace, indigenous Ghurrah pigs, and their crossbreds. A Genome Wide Association Study (GWAS) carried out using single-SNP and haplotype based methods detected a 1.6 Mb region on SSC2 (28.92-30.52 Mb) as significantly associated with antibody response to CSF vaccination. The significant region and 1 Mb flanking sequences encompass 3 genes - EIF3M, DNAJC24 and ARL14EP, which code for proteins involved in Pestivirus replication and host immune response system. Our results combined with previous studies on immune response of pigs present this region as a suitable candidate for future functional investigations.

Novel peptide (RATH) mediated delivery of peptide nucleic acids for antiviral interventions.

The peptide nucleic acid (PNA) is a chimeric molecule with the nucleobases connected by peptide bonds. This chimeric nature gives the PNA certain therapeutic advantages over natural antisense nucleic acid molecules. The PNA probes are known for its better and stronger complementation with target nucleic acids. However, cellular delivery of PNA is a major hurdle due to the charge-neutral nature of the PNA. For cellular delivery of PNA, peptide-PNA conjugates are used. This approach may face some practical limitation in terms of PNA antisense activity. In this study, we propose a novel RATH-2 peptide-based non-covalent PNA delivery mechanism. We observed RATH-2 shows a favorable molecular interaction with PNA at 16:1 (peptide:PNA) molar ratio resulting in co-centric nanoparticle formation. With this combination, we could achieve as high as 93% cellular delivery of the PNA. The proposed non-covalent RATH:PNA delivery model showed endocytic entrapment free delivery of PNA. The study further demonstrated the therapeutic application of PNA with in vitro antiviral intervention model. Using RATH-2 non-covalent PNA delivery system, we could inhibit 69.5% viral load. The present study demonstrates a cell-penetrating peptide:PNA interaction can lead to nanoparticle formations that facilitated cellular delivery of PNA.Key points• A novel cell-penetrating peptide (RATH-2) was identified for non-covalent delivery of PNA.• RATH-2 and PNA formed co-centric nanoparticles at appropriate molar combination.• PNA delivered through the RATH-2 inhibited the viral gene expression and reduced the viral load.

Anti-hyperglycemic and genotoxic studies of 1-O-methyl chrysophanol, a new anthraquinone isolated from Amycolatopsis thermoflava strain SFMA-103.

The compound 1-O-methyl chrysophanol (OMC) which belongs to a class of hydroxyanthraquinones was isolated from Amycolatopsis thermoflava strain SFMA-103 and studied for their anti-diabetic properties. OMC was evaluated as an anti-diabetic agent based on in silico studies which initially predicted the binding energy with α-amylase (-188.81 KJ mol-1) and with α-glucosidase (70.53 KJ mol-1). Further, these results were validated based on enzyme inhibition assays where OMC demonstrated enzyme inhibitory activity towards α-amylase (IC50 3.4 mg mL-1) and α-glucosidase (IC50 38.49 µg mL-1). To confirm the anti-diabetic activity, in vivo studies (oral dose in Wistar rats) revealed that OMC inhibited significantly the increase in glucose concentration at 100 mg/kg as compared to starch control (p < 0.05). Further, to understand the safety of OMC as a therapeutic agent, the genotoxic analysis was performed in both in vitro Chinese Hamster Ovary cells (250, 500, and 1000 µM/mL) and in vivo Swiss albino mice (250, 500, and 1000 mg/kg). In vitro results showed that OMC concentration of up to 250 µM/mL did not elicit significant changes in CAs, MI, and MN counts in CHO cells. Similarly, in mice experiments (i.p. injection), no significant changes in CAs, MI, and MN induction were observed till 500 mg/kg of OMC when compared with chrysophanic acid (Cy) (200 mg/kg). In addition, mice that received the lowest dose of OMC (250 mg/kg) did not show any histological changes in liver, kidney, and heart. The study concluded that five times higher therapeutic dose (100 mg/kg) of OMC can be utilized against hyperglycemia with no genotoxic effects.

Isolation, semi-synthesis, free-radicals scavenging, and advanced glycation end products formation inhibitory constituents from Parmotrema tinctorum.

Bioassay-guided separation of acetone extract from lichen Parmotrema tinctorum (Delise ex Nyl.) Hale led to the isolation of six major phenolic constituents (1-6). Compounds structures were established using NMR and mass spectral techniques. Further, to develop libraries on these scaffolds, a series of semi-synthetic derivatives were prepared (1a-1f, 2a-2b, 3a, 5a) and investigated for their free-radicals (2,2-diphenyl-1-picrylhydrazyl and 2,2'-azinobis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS)) scavenging and advanced glycation end products (AGEs) formation inhibitory activities. Amongst tested derivatives, 1a, 1d, 1e, 2a, and 5a showed strong ABTS scavenging potentials comparable to Trolox. In addition, these derivatives also manifested moderate AGEs formation inhibitory activities. [Formula: see text].

Synthesis and biological evaluation of bergenin-1,2,3-triazole hybrids as novel class of anti-mitotic agents.

In continuation of our investigation of pharmacologically-motivated natural products, we have isolated bergenin (1) as a major compound from Mallotus philippensis, which is deployed in different Indian traditional systems of medicine. Here, a series of bergenin-1,2,3-triazole hybrids were synthesized and evaluated for their potentials against a panel of cancer cell lines. Several of the hybrid derivatives were found more potent in comparison to parent compound bergenin (1). Among them, 4j demonstrated potent activity against A-549 and HeLa cell lines with IC50 values of 1.86 µM and 1.33 µM, respectively, and was equipotent to doxorubicin. Cell cycle analysis showed that 4j arrested HeLa cells at G2/M phase and lead to accumulation of Cyclin B1 protein. Cell based tubulin polymerization assays and docking studies demonstrated that 4j disrupts tubulin assembly by occupying colchicine binding pocket of tubulin.

Comprehensive Analysis of Secondary Metabolites in Usnea longissima (Lichenized Ascomycetes, Parmeliaceae) Using UPLC-ESI-QTOF-MS/MS and Pro-Apoptotic Activity of Barbatic Acid.

Considering the importance of ultra-performance liquid chromatography-electrospray ionization-quadrupole time of flight-tandem mass spectrometry (UPLC-ESI-QTOF-MS/MS) hyphenated techniques for analysis of secondary metabolites from crude extracts, the present study was aimed at identification of secondary metabolites in acetone extract of the lichen Usnea longissima. From our study, 19 compounds were tentatively identified through comparison of exact molecular masses from their MS/MS spectra, mass fragmentation studies and comparison with literature data. In addition, potent cytotoxic activity of U. longissima extract prompted us to isolate four compounds, 18R-hydroxy-dihydroalloprotolichesterinic acid (19), neuropogolic acid (20), barbatic acid (21), and usnic acid (22) from this extract which were adequately identified through mass spectrometry and NMR spectroscopy. All four compounds displayed cytotoxic activity. Barbatic acid (21) manifested doxorubicin equivalent activity against A549 lung cancer cell line with IC50 of 1.78 µM and strong G0/G1 accumulation of cells. Poly ADP-ribose polymerase (PARP) cleavage confirmed that it induced cytotoxic activity via apoptosis. Finally, our work has discerned the depside, barbatic acid (21) from crude extract as a candidate anti-cancer molecule, which induces cell death by stepping up apoptosis.

Sequence diversity of major histo-compatibility complex class II DQA1 in Indian Tharparkar cattle: novel alleles and in-silico analysis.

Exon 2 of MHC class II gene codes for the first domain of the molecule that forms the peptide-binding groove and its polymorphism partly explains functional MHC diversity. A 850 bp DQA1 gene fragment spanning from intron I to exon III was typed by sequencing of 40 Tharparkar cattle of various agro-climatic zones of northern India along with 10 Tharparkar crossbreds. On analysis of nucleotide sequences, a total of 30 polymorphic sites (1 insertion and 29 SNPs) were identified in 14 MHC alleles leading to amino acid changes in 5 places in 249 bp (exon 2). Five new BoLa DQA1 alleles were identified and reported. The within group mean distance was highest in Tharparkar herd of Bikaner (0.045) and lowest (0.020) in that of Surathgarh (breeding tract) whereas, between groups mean distance was highest in Bikaner Tharparkar-Suratgarh Tharparkar pair. There was excess of nonsynonymous over synonymous nucleotide substitutions in the present study. The effects of these substitutions were predicted using I-Mutant and Panther online resources. The mean ratio of dN/dS was found to be >1.0 at 12 codons with two mutation hotspots at 13th codon (P = 0.002) and 64th codon (P = 0.01). The phylo-geographic analysis revealed that alleles 5, 7 and 13 formed a different cluster with alleles 7 and 13 grouped by the most frequent allele (BoLa-DQA*1401).

Development of multiplex PCR assay for concurrent detection of tick borne haemoparasitic infections in bovines.

This study describes development and evaluation of a multiplex PCR assay for simultaneous detection of Theileria annulata, Babesia bigemina and Anaplasma marginale infections in bovines. The assay was developed using parasites specific genomic DNA and three sets of PCR primers targeting the Tams1, 18S rRNA and 16S rRNA genes of T. annulata, B. bigemina and A. marginale, respectively. Blood samples collected from a total of 461 bovines, suspected for haemoparasitic infections, were examined microscopically to record the status of infection and simultaneously, genomic DNA extracted from these blood samples were utilized for the optimization and validation of multiplex PCR assay. Microscopic examination of blood samples revealed presence of single and multiple species of haemoparasites in 25.8% and 2.4% samples, respectively. Results of multiplex PCR revealed the presence of single haemoparasitic species infection in 159 cases (34.5%), whereas mixed infection was recorded in 82 (17.8%) samples. Occurrence of individual species infection detected by mPCR in the study was 26.03% (120/461) for T. annulata, 3.25% (15/461) for B. bigemina and 5.20% (24/461) for A. marginale. The detection limit of multiplex PCR assay was at the template dilutions of 10-6, 10-6 and 10-4, which corresponded to 0.1 pg, 0.1 pg and 10.0 pg of DNA for T. annulata, A. marginale, and B. bigemina, respectively. Based on the high diagnostic sensitivity and throughput, multiplex PCR assay developed in the present study could be exploited as a tool to conduct large-scale epidemiological survey for tick-borne haemoparasitic infection of bovines.

In vitro study on role of σB protein in avian reovirus pathogenesis.

Avian reoviruses, members of Orthoreovirus genus was known to cause diseases like tenosynovitis, runting-stunting syndrome in chickens. Among eight structural proteins, the proteins of S-class are mainly associated with viral arthritis but the significance of σB protein in arthritis is not established till date. In this infection pathological condition together with infection of joints often leads to arthritis because joints consists of cartilage which forms lubricating surface between two bones, and has limited metabolic, replicative and repair capacity. To establish the role of σB protein in arthritis, an in-vitro microarray study was conducted consisting four groups viz. virus infected and control; pDsRed-Express-N1-σB and empty pDs-Red transfected, CEF cells. With cut-off value as FC ≥2, p value <0.05, 6709 and 4026 numbers of DEGs in virus and σB, respectively were identified. The Ingenuity Pathway Analysis gave an idea about the involvement of σB protein in "osteoarthritis pathway", which was activated with z-score with 3.151. The pathway "Role of IL-17A in arthritis pathway" was also enriched with -log (p-value) 1.64. Among total 122 genes involved in osteoarthritis pathway, 28 upregulated and 11 downregulated DEGs were common to both virus and σB treated cells. Moreover, 14 upregulated and 7 downregulated were unique in σB transfected cells. Using qRT-PCR for IL-1B, BMP2, SMAD1, SPP1 genes, the microarray data was validated. We concluded that during ARV infection σB protein, if not fully partially leads to molecular alteration of various genes of host orchestrating the different molecular pattern in joints, leading to tenosynovitis syndrome.

UPLC-MS/MS quantitative analysis and structural fragmentation study of five Parmotrema lichens from the Eastern Ghats.

Comparative phytochemical analysis of five lichen species [Parmotrema tinctorum (Delise ex Nyl.) Hale, P. andinum (Mull. Arg.) Hale, P. praesorediosum (Nyl.) Hale, P. grayanum (Hue) Hale, P. austrosinense (Zahlbr.) Hale] of Parmotrema genus were performed using two complementary UPLC-MS systems. The first system consists of high resolution UPLC-QToF-MS/MS spectrometer and the second system consisted of UPLC-MS/MS in Multiple Reaction Monitoring (MRM) mode for quantitative analysis of major constituents in the selected lichen species. The individual compounds (47 compounds) were identified using Q-ToF-MS/MS, via comparison of the exact molecular masses from their MS/MS spectra, the comparison of literature data and retention times to those of standard compounds which were isolated from crude extract of abundant lichen, P. tinctorum. The analysis also allowed us to identify unknown peaks/compounds, which were further characterized by their mass fragmentation studies. The quantitative MRM analysis was useful to have a better discrimination of species according to their chemical profile. Moreover, the determination of antioxidant activities (ABTS+ inhibition) and Advance Glycation Endproducts (AGEs) inhibition carried out for the crude extracts revealed a potential antiglycaemic activity to be confirmed for P. austrosinense.

Development of a recombinant σB protein based dot-ELISA for the diagnosis of avian reovirus (ARV).

Avian reovirus (ARV) causes significant economic losses to the poultry industry worldwide. The ARV proteins fall into three different classes based on their sizes:λ (large); µ (medium) and σ (small). σB, an outer capsid protein of the ARV contains group specific neutralizing epitopes and induces strong immune response in naturally infected chickens. This study describes the development of a rapid dot-enzyme linked immunosorbent assay (dot-ELISA) using recombinant σB protein antigen of 54 kDa (approx). The assay is rapid (4-5 h) and results can be read by the naked eye. Sixteen ARV positive serum samples (group A) produced strong reaction in the dot-ELISA while twenty of the ARV negative serum samples (group B) collected from SPF chickens showed no reaction. Seventy six randomly collected serum samples were tested with a commercial indirect ELISA kit and the in-house developed dot-ELISA. A total of sixty eight serum samples were found to be positive by indirect ELISA and sixty five serum samples were found to be positive by dot-ELISA. Therefore, using the commercial ELISA as the reference test, the dot-ELISA had a diagnostic sensitivity of 83.8% and specificity of 88.6%. This dot-ELISA can be used as a simple, reliable and inexpensive alternative to commercial ELISA kits for serodiagnosis of ARV where the facilities for standard ELISA are not available.

Synthesis and biological evaluation of Schizandrin derivatives as potential anti-cancer agents.

A new series of Schizandrin (1) derivatives were synthesized utilizing the C-9 position of the Schizandrin core and evaluated for their cytotoxic activities against HeLa (cervical cancer), A549 (lung cancer), MCF-7 (breast cancer) and DU-145 (prostate cancer) cell lines. Among the synthesized series, 4e, 4f, 4g and 5 showed potent activities against tested cell lines. More significantly, compound 5 exhibited most potent cytotoxic activity against DU-145 with an IC50 value of 1.38 µM which is comparable to the standard agent, doxorubicin. Further, flow cytometry analysis indicated that 5 arrested cells in G2/M phase and consequently leading to apoptosis. Molecular docking analysis showed that 5 occupied the colchicine binding pocket of tubulin. Overall, the present study demonstrates that 5, as a mitotic-agent.

Modulation of Host miRNAs Transcriptome in Lung and Spleen of Peste des Petits Ruminants Virus Infected Sheep and Goats.

Peste des petits ruminants (PPR) is one of the highly contagious viral disease, characterized by fever, sore mouth, conjunctivitis, gastroenteritis, and pneumonia, primarily affecting sheep and goats. Reports suggested variable host response in goats and sheep and this host response vis-a-vis the expression of microRNAs (miRNAs) has not been investigated. Here, miRNAs were sequenced and proteomics data were generated to identify the role of differentially expressed miRNA (DEmiRNA) in PPR virus (PPRV) infected lung and spleen tissues of sheep and goats. In lungs, 67 and 37 DEmiRNAs have been identified in goats and sheep, respectively. Similarly, in spleen, 50 and 56 DEmiRNAs were identified in goats and sheep, respectively. A total of 20 and 11 miRNAs were found to be common differentially expressed in both the species in PPRV infected spleen and lung, respectively. Six DEmiRNAs-miR-21-3p, miR-1246, miR-27a-5p, miR-760-3p, miR-320a, and miR-363 were selected based on their role in viral infections, apoptosis, and fold change. The target prediction analysis of these six selected DEmiRNAs from the proteome data generated, revealed involvement of more number of genes in lung and spleen of goats than in sheep. On gene ontology analysis of host target genes these DEmiRNAs were found to regulate several immune response signaling pathways. It was observed that the pathways viz. T cell receptor signaling, Rap1 signaling, Toll-like receptor signaling, and B cell receptor signaling governed by DEmiRNAs were more perturbed in goats than in sheep. The data suggests that PPRV-induced miR-21-3p, miR-320a, and miR-363 might act cooperatively to enhance viral pathogenesis in the lung and spleen of sheep by downregulating several immune response genes. The study gives an important insight into the molecular pathogenesis of PPR by identifying that the PPRV-Izatnagar/94 isolate elicits a strong host response in goats than in sheep.

New cycloartane type-triterpenoids from the areal parts of Caragana sukiensis and their biological activities.

A comprehensive re-investigation of aerial parts of Caragana sukiensis resulted in the isolation of twelve compounds (1-12) including three new cycloartane type triterpenoids (3-5) respectively. Chemical structures of the isolated compounds were established by analysis of their IR, HRMSESI, 1D and 2D NMR spectroscopic data. In addition, these compounds were evaluated for their cytotoxic activity against cancer lines (HeLa, A549, MCF-7, DU-145) and Human embryonic kidney cell line (HEK-293). The results indicated that compound 8 showed potent cytotoxic activity against A549 with IC50 value of 1.54 µM which is comparable to standard drug, doxorubicin. Further, flow cytometric analysis showed that compound 8 arrested the cell cycle in the Go/G1 phase leading to apoptotic cell death. In addition, Hoechst 33258 staining, Annexin V-FITC assay and measurement of mitochondrial membrane potential also suggested that 8 induced cell death by apoptosis. Further, all the isolates were also screened for their antifeedant and insecticidal activity against tobacco caterpillar (Spodoptera litura), using no-choice leaf disk method. Among screened compounds 1, 3, 4, and 6 showed potent antifeedancy with ED50 values of 0.59, 1.19, 0.67, and 1.68 µg/cm2. Overall, this study identified a novel class of cycloartane tritepenoids as potent cyotoxic agents as well as antifeedants.

Comparative and temporal transcriptome analysis of peste des petits ruminants virus infected goat peripheral blood mononuclear cells.

Peste des petits ruminanats virus (PPRV), a morbillivirus causes an acute, highly contagious disease - peste des petits ruminants (PPR), affecting goats and sheep. Sungri/96 vaccine strain is widely used for mass vaccination programs in India against PPR and is considered the most potent vaccine providing long-term immunity. However, occurrence of outbreaks due to emerging PPR viruses may be a challenge. In this study, the temporal dynamics of immune response in goat peripheral blood mononuclear cells (PBMCs) infected with Sungri/96 vaccine virus was investigated by transcriptome analysis. Infected goat PBMCs at 48h and 120h post infection revealed 2540 and 2000 differentially expressed genes (DEGs), respectively, on comparison with respective controls. Comparison of the infected samples revealed 1416 DEGs to be altered across time points. Functional analysis of DEGs reflected enrichment of TLR signaling pathways, innate immune response, inflammatory response, positive regulation of signal transduction and cytokine production. The upregulation of innate immune genes during early phase (between 2-5 days) viz. interferon regulatory factors (IRFs), tripartite motifs (TRIM) and several interferon stimulated genes (ISGs) in infected PBMCs and interactome analysis indicated induction of broad-spectrum anti-viral state. Several Transcription factors - IRF3, FOXO3 and SP1 that govern immune regulatory pathways were identified to co-regulate the DEGs. The results from this study, highlighted the involvement of both innate and adaptive immune systems with the enrichment of complement cascade observed at 120h p.i., suggestive of a link between innate and adaptive immune response. Based on the transcriptome analysis and qRT-PCR validation, an in vitro mechanism for the induction of ISGs by IRFs in an interferon independent manner to trigger a robust immune response was predicted in PPRV infection.

Free radical scavenging, α-glucosidase inhibitory and anti-inflammatory constituents from Indian sedges, Cyperus scariosus R.Br and Cyperus rotundus L.

BACKGROUND: Cyperus scariosus R. Br and Cyperus rotundus L are widely used in ayurvedic preparation for the treatment of diabetes and other diseases. The early literature, so far, does not indicate the presence of any bioactive principle isolated from these plants. OBJECTIVE: To identify free radical scavenging, anti-diabetic and anti- inflammatory principles from these two species. MATERIALS AND METHODS: The bioassay guided fractionation and isolation of active constituents was done by chromatographic techniques. They also evaluated their anti-oxidant activity by DPPH and ABTS. The anti-diabetic activity was screened by α- glucosidase and α- amylase assays. Also, the further evaluation of in vitro anti-inflammatory activity using THP-1 monocytic cells and in vivo anti- inflammatory activity, was confirmed by carrageenan induced rat paw edema as model. RESULTS: The activity guided isolation led to isolation of twelve compounds Which are: Stigmasterol[1], ß- sitosterol[2], Lupeol[3], Gallic acid[4], Quercetin[5], ß- amyrin[6], Oleanolic acid[7], ß- amyrin acetate[8], 4- hydroxyl butyl cinnamate[9], 4- hydroxyl cinnamic acid[10], Caffeic acid,[11] and Kaempferol[12] respectively. Among the isolates, the compounds 4 and 5 displayed potent radical scavenging activity with an IC50 values of 0.43 and 0.067 ΅g/ml. The compounds 4, 5 and 10 showed significant anti-diabetic activities. while lupeol[3] showed potent IL-1 ß activity inhibition in THP-1 monocytic cells and also displayed significant (p<0.0025) in vivo anti-inflammatory activity. CONCLUSION: Inbrief, we isolated twelve compounds from both the species. Collectively, our results suggested that aromatic compounds showed good anti-oxidant and anti-diabetic activities. SUMMARY: The study investigates the free radical scavenging, α-glucosidase inhibitory and anti-inflammatory effects of constituents isolated from Indian sedges viz. C. scariosus and C. rotundus. The results indicated that phenolic compounds displayed potent fee radical scavenging activty and alpha-glucosidase inhibition activity. While terpene constituent, Lupeol[3] showed good IL-1ß activity inhibition in THP-1 monocytic cells and also displayed significant (p<0.0025) in vivo anti inflammatory activity in carrageenan induced rat paw edema. However, further studies are required to know the exact molecular mechanism. Abbreviations used: DPPH: 2,2- Diphenyl-1-1-picryl hydrazyl, ABTS: 2,2-Azinobis-3-ethylbenzo thiazoline-6-sulfonic acid, THP-1: Human leukaemia monocytic cell line, IL-1ß: Interleukin-1ß, IC50-Inhibitory concentration 50%.

HN protein of Newcastle disease virus sensitizes HeLa cells to TNF-α-induced apoptosis by downregulating NF-κB expression.

Hemagglutinin neuraminidase (HN) is a membrane protein of Newcastle disease virus (NDV) with the ability to induce apoptosis in many transformed cell lines. TNF-α is a multi-factorial protein that regulates cell survival, differentiation and apoptosis. In a previous study, we reported that HN protein induces apoptosis by downregulating NF-κB expression. Further, we speculated that downregulation of NF-κB expression might sensitize HeLa cells to TNF-α-mediated apoptosis. Therefore, the present study was undertaken to investigate if HN protein could sensitize HeLa cells to TNF-α and to examine the apoptotic potential of the HN protein and TNF-α in combination. The results revealed that the pro-apoptotic effects were more pronounced with the combination of HN and TNF-α than with HN or TNF-α alone, which indicates that the HN protein indeed sensitized the HeLa cells to TNF-α-induced cell death. The results of the study provide a mechanistic insight into the apoptotic action of HN protein along with TNF-α, which could be valuable in treating tumor types that are naturally resistant to TNF-α.

Production and characterization of monoclonal antibodies to Newcastle Disease Virus.

Newcastle Disease (ND) is one of the major causes of economic loss in the poultry industry. Newcastle Disease Virus (NDV) is a single-stranded, negative-sense enveloped RNA virus (Fam. Paramyxoviridae; Order Mononegavirales). In the present study three monoclonal antibodies (MAbs) were produced by polyethyleneglycol (PEG)-mediated fusion of lymphocytes sensitized to NDV Bareilly strain and myeloma cells. NDV possesses ability to agglutinate erythrocytes of avian species. All the three MAbs designated as 2H7, 3E9 and 3G6 caused hemagglutination inhibition of NDV by specifically binding to NDV. The reactivity for all the 3 MAbs on indirect ELISA was found to be significantly higher than the antibody and antigen controls. On flowcytometry of HeLa cells infected with NDV using the MAbs as primary antibodies, there was a significant difference in the percentage of cells showing positive fluorescence compared to the mock control. One of the MAbs (3E9) was found to react with hemagglutinin-neuraminidase (HN) protein on western blot.