Alteration in the in vitro activity of milk leukocytes during different parity in high yielding cross-bred cows

In vitro activity of milk leukocytes (viz. neutrophils, lymphocytes and macrophages) was evaluated in forty-eight (48) clinically healthy high-yielding cross-bred cows of mid-lactation stage (100–200 days of lactation), divided into four groups namely 1st parity (n = 12), 2nd parity (n = 12), 3rd parity (n = 12) and 4th and above parity (n = 12). Milk samples were taken (250 ml/cow) were taken. Milk somatic cell counts (SCC) and differential leukocyte counts (DLC) were performed microscopically. In vitro phagocytic index (PI) of milk neutrophils and macrophages was evaluated by colorimetric nitro blue tetrazolium reductive assay. Mitogen-induced milk lymphocyte blastogenic response was measured by colorimetric MTT (tetrazolium) assay after isolation of the milk leukocytes by density gradient centrifugation. Milk SCC differed significantly (p < 0.01) between different parity. Cows of 4 and above parity showed significantly (p < 0.01) higher milk SCC compared to primiparous cows. There was no significant difference in milk DLC during different parities in high-yielding cross-bred cows. There was a significant (p < 0.01) variation in lymphocyte blastogenesis amongst parity. The highest value of lymphocyte blastogenesis was seen at 3rd parity, whereas lowest value was obtained in the cows of both 1st and 4th or above parity. PI of milk neutrophils did not differ significantly between parity. PI of milk macrophages was significantly (p < 0.01) higher in 3rd parity and lower (p < 0.01) in 1st and 4th parities. The study indicated that depressed activity of milk lymphocytes and macropages was lower and SCC was higher in the cows of 4th and above parity indicating more mammary stress and hence susceptible to udder infection and mastitis. Therefore, better care and managemental interventions should be taken around these periods.     

A Simple Multi-band Metamaterial Absorber with Combined Polarization Sensitive and Polarization Insensitive Characteristics for Terahertz Applications

This paper presents a simple multi-band metamaterial absorber for terahertz applications. The unit cell of the proposed structure consists of a single square ring having gaps at the centers on three of its sides. The proposed absorber produces three absorption bands for all polarizations and hence the design can be considered as insensitive to polarization variation. It provides an average absorption of 96.92% for the TE polarization with a peak absorption of 99.44% at 3.87 THz and for the TM polarization, it provides an average absorption of 98.4% with a peak absorption of 99.86% at 3.87 THz. An additional absorption peak is observed for the TE polarization at 1.055 THz that gradually diminishes with the increase in polarization angle and completely vanishes for the TM polarization. Thus, the structure displays a hybrid polarization response with polarization insensitivity in three bands and polarization sensitivity in one band. Parametric analysis has been carried out validating the optimal selection of the design parameters. The simplicity of the design and its combined polarization sensitive and polarization insensitive absorption characteristics can find tremendous applications in the field of terahertz imaging and sensing.

     

Virulence and pathogenicity of a Candida albicans mutant with reduced filamentation

Deletion of DNA polymerase eta (Rad30/Polη) in pathogenic yeast Candida albicans is known to reduce filamentation induced by serum, ultraviolet, and cisplatin. Because nonfilamentous C. albicans is widely accepted as avirulent form, here we explored the virulence and pathogenicity of a rad30Δ strain of C. albicans in cell‐based and animal systems. Flow cytometry of cocultured fungal and differentiated macrophage cells revealed that comparatively higher percentage of macrophages was associated with the wild‐type than rad30Δ cells. In contrast, higher number of Polη‐deficient C. albicans adhered per macrophage membrane. Imaging flow cytometry showed that the wild‐type C. albicans developed hyphae after phagocytosis that caused necrotic death of macrophages to evade their clearance. Conversely, phagosomes kill the fungal cells as estimated by increased metacaspase activity in wild‐type C. albicans. Despite the morphological differences, both wild‐type and rad30? C. albicans were virulent with a varying degree of pathogenicity in mice models. Notably, mice with Th1 immunity were comparatively less susceptible to systemic fungal infection than Th2 type. Thus, our study clearly suggests that the modes of interaction of morphologically different C. albicans strains with the host immune cells are diverged, and host genetic background and several other attributing factors of the fungus could additionally determine their virulence.     

Fabrication of agar-gelatin hybrid scaffolds using a novel entrapment method for in vitro tissue engineering applications

Scaffolds of agar and gelatin were developed using a novel entrapment method where agar and gelatin molecules mutually entrapped one another forming stable cell adhesive matrices. Glutaraldehyde was used as a crosslinking agent for gelatin. Three types of hybrid matrices were prepared using agar and gelatin in different proportions in the weight ratio of 1:1, 2:1, and 3:1. Surface characterization of dry scaffolds was carried out by scanning electron microscope. Swelling studies were carried out in phosphate buffer saline (PBS) at physiological pH 7.4. The integral stability of the scaffolds was evaluated by estimating the released disintegrated gelatin from them in PBS at pH 7.4. The attachment kinetics of the cells was evaluated by culturing mouse fibroblast cell line NIH 3T3 on films. The cytocompatibility of these matrices was determined by studying growth kinetics of NIH 3T3 cells on them and morphology of cells was observed through optical photographs taken at various days of culture. It was found that the matrices containing agar and gelatin in 2:1 weight ratio exhibited best growth kinetics. The results obtained from these studies have suggested that the above-described method is a cheap and easy way to fabricate agar-gelatin hybrid scaffolds to grow cells which can be used in various in vitro tissue engineering applications like screening of drugs.     

Structural characterization and oligomerization of PB1-F2, a proapoptotic influenza A virus protein

Recently, a novel 87-amino acid influenza A virus protein with proapoptotic properties, PB1-F2, has been reported that originates from an alternative reading frame in the PB1 polymerase gene and is encoded in most known human influenza A virus isolates. Here we characterize the molecular structure of a biologically active synthetic version of the protein (sPB1-F2). Western blot analysis, chemical cross-linking, and NMR spectroscopy afforded direct evidence of the inherent tendency of sPB1-F2 to undergo oligomerization mediated by two distinct domains located in the N and C termini, respectively. CD and (1)H NMR spectroscopic analyses indicate that the stability of structured regions in the molecule clearly depends upon the hydrophobicity of the solvent. In aqueous solutions, the behavior of sPB1-F2 is typical of a largely random coil peptide that, however, adopts alpha-helical structure upon the addition of membrane mimetics. (1)H NMR analysis of three overlapping peptides afforded, for the first time, direct experimental evidence of the presence of a C-terminal region with strong alpha-helical propensity comprising amino acid residues Ile(55)-Lys(85) connected via an essentially random coil structure to a much weaker helix-like region, located in the N terminus between residues Trp(9) and Lys(20). The C-terminal helix is not a true amphipathic helix and is more compact than previously predicted. It corresponds to a positively charged region previously shown to include the mitochondrial targeting sequence of PB1-F2. The consequences of the strong oligomerization and helical propensities of the molecule are discussed and used to formulate a hypothetical model of its interaction with the mitochondrial membrane.     

Effect of immunization with lipid associated polysaccharide antigen and anti CD-2 antibodies on class II MHC expression and cellular immune response in BALB/C mice infected with Leishmania donovani

In a bid to characterize the antigens and immunization mechanisms which may be used to produce a protective response against L. donovani, role of lipid associated polysaccharide (LPS) antigen and whole antigen was evaluated. BALB/C mice were immunized with whole or LPS antigen in combination with one of three putative adjuvents (anti CD-2 antibody/FIA/0.85% Saline). LPS antigen emulsified in anti CD-2 antibody was found to induce significant antibodies in mice on day 28 against challenge with lethal dose of L. donovani. Immunoprophylactic properties of LPS and whole antigen was investigated on day 40 through cytokine elicitation (IL-2), MIF) in culture supernatants of spleen cells, but before that MHC-II expressed on macrophage was studied. The LPS antigen in combination with anti CD-2 antibody was found to be most immuno-reactive inducing higher MHC-II expression on macrophages which was associated with substantial rise in the level of MIF and IL-2. It coincided with decline in antibody titre in 100% mice immunized with LPS antigen while Leishmania injected as whole antigen failed to induce specific macrophage and T-cell response with all the above formulations. We surmise from our data that lipid associated polysaccharide antigen linked to anti CD-2 antibody has potential for eliciting protective immunity against Leishmania.