Interferon-Inducing Activity of Acidic Polysaccharides

Readily available polysaccharides, amylopectin, amylose, dextrin, and yeast mannan, were chemically phosphorylated using polyphosphoric acid in the presence of a tertiary amine, and the resultant phosphates were examined for their interferon-inducing activity in rabbits employing an assay system consisting of a primary culture of rabbit kidney cells and vesicular stomatitis virus. All the phosphates were shown to be active as interferon inducer, and, especially, the activity of those containing more than 2% phosphorus were quite strong. Interferons evoked by the above phosphates resembled those induced by bacterial endotoxin, e.g., the viral inhibiting activity was susceptible to heat treatment, low pH and tryptic digestion. Since all the parent polysaccharides showed no interferon-inducing activity, it is reasonable to assume that the active center of these inducers might reside or be due to the anionic phosphate groups.     

Bluetongue virus RNA binding protein NS2 is a modulator of viral replication and assembly

Background  

Bluetongue virus (BTV) particles consist of seven structural proteins that are organized into two capsids. In addition, BTV also encodes three non-structural (NS) proteins of which protein 2 (NS2) is the RNA binding protein and is also the major component of virus encoded inclusion bodies (VIBs), which are believed to be virus assembly sites. To investigate the contribution of NS2 in virus replication and assembly we have constructed inducible mammalian cell lines expressing full-length NS2. In addition, truncated NS2 fragments were also generated in an attempt to create dominant negative mutants for NS2 function.     

Novel expression of kallikreins, kallikrein-related peptidases and kinin receptors in human pleural mesothelioma

Malignant mesothelioma is an aggressive cancer of the pleura that is causally related to exposure to asbestos fibres. The kallikrein serine proteases [tissue (hK1) and plasma (hKB1) kallikreins, and kallikrein-related peptidases (KRP/hK2-15)] and the mitogenic kinin peptides may have a role in tumourigenesis. However, it is not known whether hK1, hKB1, KRP/hK proteins or kinin receptors are expressed in pleural mesotheliomas. The expression of hK1, hKB1, KRP/hK2, 5, 6, 7, 8 and 9, and kinin B(1) and B(2) receptors was assessed in archived selected normal tissue and mesothelioma tumour sections by immunoperoxidase and immunofluorescence labelling. hK1, hKB1 and kinin B(1) and B(2) receptors were expressed in malignant cells of the epithelioid and sarcomatoid components of biphasic mesothelioma tumour cells. The percentage of cells with cytoplasmic and nuclear labelling and the intensity of labelling were similar for hK1, hKB1 and the kinin receptors. KRP/hK2, 6, 8 and 9 were also expressed in the cytoplasm and nuclei of mesothelioma cells, whereas KRP/hK5 and hK7 showed predominantly cytoplasmic localisation. This is a first report, but further studies are required to determine whether these proteins have a functional role in the pathogenesis of mesothelioma and/or may be potential biomarkers for pleural mesothelioma.     

Rewiring of sIgM-Mediated Intracellular Signaling through the CD180 Toll-like Receptor

Chronic lymphocytic leukemia (CLL) development and progression are thought to be driven by unknown antigens/autoantigens through the B cell receptor (BCR) and environmental signals for survival and expansion including toll-like receptor (TLR) ligands. CD180/RP105, a membrane-associated orphan receptor of the TLR family, induces normal B cell activation and proliferation and is expressed by approximately 60% of CLL samples. Half of these respond to ligation with anti-CD180 antibody by increased activation/phosphorylation of protein kinases associated with BCR signaling. Hence CLL cells expressing both CD180 and the BCR could receive signals via both receptors. Here we investigated cross-talk between BCR and CD180-mediated signaling on CLL cell survival and apoptosis. Our data indicate that ligation of CD180 on responsive CLL cells leads to activation of either prosurvival Bruton tyrosine kinase (BTK)/phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/AKT-mediated, or proapoptotic p38 mitogen-activated protein kinase (p38MAPK)-mediated signaling pathways, while selective immunoglobulin M (sIgM) ligation predominantly engages the BTK/PI3K/AKT pathway. Furthermore, pretreatment of CLL cells with anti-CD180 redirects IgM-mediated signaling from the prosurvival BTK/PI3K/AKT toward the proapoptotic p38MAPK pathway. Thus preengaging CD180 could prevent further prosurvival signaling mediated via the BCR and, instead, induce CLL cell apoptosis, opening the door to therapeutic profiling and new strategies for the treatment of a substantial cohort of CLL patients.     

Degradation of bacterial DNA by a natural antimicrobial agent with the help of biomimetic membrane system

The antimicrobial efficacy of methylglyoxal (MG) against several gram-negative bacteria including Escherichia coli has been reported. To determine the mechanism of action of MG, molecular interactions between lipid and MG within the liposomal membrane were also investigated. Multilamellar and unilamellar vesicles were prepared from 1, 2-dipalmitoyl-snglycero-3-phosphocholine (DPPC). The effect of MG on DPPC liposomal membrane was studied by fluorescence spectroscopy and differential scanning calorimetry. The results indicate that MG interacts mainly with the DPPC head group that produces a significant increase in the fluidity of liposomal vesicles, which could be the cause of a fusion/aggregation effect in microbial cells. The agarose gel electrophoresis study with the genomic DNA extracted from E. coli ATCC 25922 revealed that addition of MG could completely degrade this DNA within 1 h, pointing out to their distinctly high degree of sensitivity towards MG. Further, the drug was able to cross the cell membranes, penetrating into the interior of the cell and interacting with DNA for demonstrating antibacterial activity of MG.     

Utilization of an amphipathic leucine zipper sequence to design antibacterial peptides with simultaneous modulation of toxic activity against human red blood cells

The toxicity of naturally occurring or designed antimicrobial peptides is a major barrier for converting them into drugs. To synthesize antimicrobial peptides with reduced toxicity, several amphipathic peptides were designed based on the leucine zipper sequence. The first one was a leucine zipper peptide (LZP); in others, leucine residues at the a- and/or d-position were substituted with single or double alanine residues. The results showed that LZP and its analogs exhibited appreciable and similar antibacterial activity against the tested gram-positive and gram-negative bacteria. However, the substitution of alanine progressively lowered the toxicity of LZP against human red blood cells (hRBCs). The substitution of leucine with alanine impaired the binding and localization of LZP to hRBCs, but had little effect on the peptide-induced damage of Escherichia coli cells. Although LZP and its analogs exhibited similar permeability, secondary structures, and localization in negatively charged membranes, significant differences were observed among these peptides in zwitterionic membranes. The results suggest a novel approach for designing antibacterial peptides with modulation of toxicity against hRBCs by employing the leucine zipper sequence. Also, to the best of our knowledge, the results demonstrate that this sequence could be utilized to design novel cell-selective molecules for the first time.     

The Effect of Inbreeding on Mortality and Morbidity among Telugu-speaking Populations of Kharagpur, West Bengal, India

Increased mortality and morbidity including congenital malformations among the offspring of consanguineous marriages have been widely reported in human populations from different parts of the world. However, there are few studies on the effect of the intensity of inbreeding and different degrees of inbreeding on mortality and morbidity. The present study is an attempt to examine the effects of inbreeding on mortality and morbidity including congenital disorders in different levels of inbreeding among Telugu-speaking populations of Kharagpur, West Bengal, India, based on data collected through extensive pedigrees. The study reveals that the frequency of spontaneous abortions and stillbirths is higher in the offspring of consanguineous marriages than in that of non-consanguineous marriages. A similar effect is also observed in the infant mortality rate, which is known to have a genetic component, but is not seen in the mortality rate of children and juveniles. The rate of morbidity is consistently higher in the offspring of consanguineous marriages with a sex bias in favour of inbred females. The increased morbidity rates in inbred individuals tend to be inversely correlated with the increase in average autosomal inbreeding coefficient. This appears to strengthen Sanghvi’s hypothesis of a decline in the frequency of deleterious genes with intensification of inbreeding through generations. The present study also confirms an increase in genetic disorders with an increase in inbreeding in almost all populations.     

Detection of alginate oligosaccharides from mollusks

We attempted in this study to detect alginate oligosaccharides (AO) from mollusks. The samples used were digestive organs taken from turban shells and abalones which commonly ate brown algae. High-performance liquid chromatography (HPLC) and negative-ion electrospray ionization (ESI) quadrupole time-of-flight (Q-TOF) mass spectrometry (MS) analyses were used to confirm the presence of AO. Samples spiked with AO resulted in observable peaks where the HPLC area was increased. The highest content was estimated to be 401.8 mg/100 g of digestive organ. The product-ion data derived from AO molecular weight were detected at a constant interval by Q-TOF MS/MS analysis. These findings indicate that AO was present in the digestive organs of mollusks.     

Isolation and partial characterization of ribonuclease inhibitor from goat liver

Ribonuclease inhibitor (RI), a 50 kDa protein, has been found both in mammalian and nonmammalian tissues. We have isolated RI from goat liver and partial characterization has been accomplished. For the isolation of RI, DEAE cellulose column chromatography followed by affinity chromatography using CNBr activated Sepharose 4B was performed. The inhibition of ribonucleolytic activity of Ribonuclease A has been checked by an agarose gel based assay. The antiangiogenic property of the protein was tested by the chorioallantoic membrane (CAM) assay. Results indicate inhibition of angiogenesis.