Oldest known non-marine diatoms (Aulacoseira) from the uppermost Cretaceous Deccan Intertrappean beds and Lameta Formation of India

Non-marine diatoms occur in the Deccan Intertrappean beds (Upper Cretaceous) of Mohgaon-Kalan, Chhindwara District, Madhya Pradesh and Pisdura, Lameta Formation (Upper Cretaceous), Maharashtra, India. This represents the oldest record of non-marine diatoms yet reported and the oldest from the Indian subcontinent. The diatoms were recovered from thin sections of chert and dinosaur coprolites by random fracturing. Solitary forms are the most common but colonial filaments up to five cells were also observed. Based on the morphological characters, the diatoms are identified as Aulacoseira Thwaites. The Lower Cretaceous marine diatom genus Archepyrgus Gersonde and Harwood also resembles Aulacoseira in general morphological characters and it seems that Aulacoseira evolved from Archepyrgus and migrated to the non-marine realm.     

Rolling adhesion kinematics of yeast engineered to express selectins

Selectins are cell adhesion molecules that mediate capture of leukocytes on vascular endothelium as an essential component of the inflammatory response. Here we describe a method for yeast surface display of selectins, together with a functional assay that measures rolling adhesion of selectin-expressing yeast on a ligand-coated surface. E-selectin-expressing yeast roll specifically on surfaces bearing sialyl-Lewis-x ligands. Observation of yeast rolling dynamics at various stages of their life cycle indicates that the kinematics of yeast motion depends on the ratio of the bud radius to the parent radius (B/P). Large-budded yeast "walk" across the surface, alternately pivoting about bud and parent. Small-budded yeast "wobble" across the surface, with bud pivoting about parent. Tracking the bud location of budding yeast allows measurement of the angular velocity of the yeast particle. Comparison of translational and angular velocities of budding yeast demonstrates that selectin-expressing cells are rolling rather than slipping across ligand-coated surfaces.     

Evaluation of toxic potential of captan: Induction of hsp70 and tissue damage in transgenic Drosophila melanogaster (hsp70-lacZ) Bg9

The study investigated the working hypothesis that a widely used fungicide captan exerts toxic effects on nontarget organisms. Transgenic Drosophila melanogaster (hsp70-lacZ) was used as a model by assaying stress gene expression as an endpoint for cytotoxicity and also to evaluate whether stress gene expression is sufficient enough to protect and to prevent tissue damage against toxic insult of the chemical. The study was further extended to understand the effect of the pesticide on development, life cycle, and reproduction of the organism and finally to evaluate a concentration of the chemical to be nontoxic to the organism. The study showed that (i) captan causes cytotoxicity at and above 0.015 ppm; (ii) at 0.0015 ppm captan, absence of hsp70 expression in the exposed organism was evaluated as the concentration referred to as no observed adverse effect level (NOAEL) for Drosophila; (iii) emergence pattern of flies was affected only at the highest concentration of captan by 4 days, while hatching and survivorship were unaffected even at this concentration; (iv) reproductive performance was significantly affected only at 125.0 and 1250.0 ppm captan, while in the lower dietary concentrations no such deleterious effects were observed; (v) at 1250.0 ppm, hsp70 failed to protect the cells from toxicant assault after 48 h exposure, thus leading to tissue damage as revealed by Trypan Blue staining. The present study shows the cytotoxic potential of captan and further reveals the application of stress genes in determining NOAEL and its expression as bioindicator of exposure to environmental contaminants.     

The immunologically protective P-4 antigen of Leishmania amastigotes. A developmentally regulated single strand-specific nuclease associated with the endoplasmic reticulum

The purified membrane-associated Leishmania pifanoi amastigote protein P-4 has been shown to induce protective immunity against infection and to elicit preferentially a T helper 1-like response in peripheral blood mononuclear cells of patients with American cutaneous leishmaniasis. As this molecule is potentially important for future vaccine studies, the L. pifanoi gene encoding the P-4 membrane protein was cloned and sequenced. Southern blot analyses indicate the presence of six tandemly arrayed copies of the P-4 gene in L. pifanoi; homologues of the P-4 gene are found in all other species of the genus Leishmania examined. DNA-derived protein sequence data indicated an identity to the P1 zinc-dependent nuclease of Penicillium citrinum (20.8%) and the C-terminal domain of the 3' nucleotidase of Leishmania donovani (33.7%). Consistent with these sequence analyses, purified L. pifanoi P-4 protein possesses single strand nuclease (DNA and RNA) and phosphomonoesterase activity, with a preference for UMP > TMP > AMP > CMP. Double-labeling immunofluorescence microscopic analyses employing anti-binding protein antibodies revealed that the P-4 protein is localized in the endoplasmic reticulum of the amastigote. Northern blot analyses indicated that the gene is selectively expressed in the intracellular amastigote stage (mammalian host) but not in the promastigote stage (insect) of the parasite. Based upon its subcellular localization and single-stranded specific nuclease activity, possible roles of the P-4 nuclease in the amastigote in RNA stability (gene expression) or DNA repair are discussed.     

Stimulation of protein kinase C modulates insulin-like growth factor-1-induced akt activation in PC12 cells

Activation of protein kinase C (PKC) plays an important role in the negative regulation of receptor signaling, but its effect on insulin-like growth factor-1 (IGF-1) receptor signaling remains unclear. In this study, we characterized the intracellular pathways involved in IGF-1-induced activation of Akt and evaluated the effects of the PKC activator phorbol 12-myristate 13-acetate (PMA) on the Akt activation by IGF-1. IGF-1 induced a time- and concentration-dependent activation of Akt. The effect of IGF-1 was blocked by the phosphatidylinositide 3-kinase (PI3K) inhibitors LY294002 (50 micrometer) and wortmannin (0.5 micrometer), but not by the MEK inhibitor PD98059 (50 micrometer) or the p70 S6 kinase pathway inhibitor rapamycin (50 nm), suggesting that the stimulation of Akt by IGF-1 is mediated by the PI3K pathway. Interestingly, cotreatment with PMA (400 nm) attenuated IGF-1-induced activation of Akt. The attenuation was blocked completely by the PKC inhibitor GO6983 (0.5 micrometer), but only partially by the MEK inhibitor PD98059 (50 micrometer), indicating that MAPK-dependent and -independent pathways are involved. PMA induced the activation of PKC in PC12 cells, and this induction was blocked by GO6983. These data further support the role of PKC in the effect of PMA. Moreover, PKCdelta is likely involved in the action of PMA on the basis of data obtained using isoform-specific inhibitors such as rottlerin. PMA also decreased IGF-1-induced tyrosine phosphorylation of insulin receptor substrate-1 and its association with PI3K. Taken together, these results suggest, for the first time, that stimulation of PKC modulates IGF-1-induced activation of Akt.     

Propagation of Dalbergia sissoo Roxb. through in vitro shoot proliferation from cotyledonary nodes

A protocol is presented for micropropagation of an economically important timber-yielding forest tree, Dalbergia sissoo Roxb. (Sissoo). Multiple shoots were induced from cotyledonary nodes derived from 1-week-old axenic seedlings on Murashige and Skoog's medium containing either N ⁶-benzyladenine (BA), kinetin (Kn), isopentenyladenine (2iP) or thidiazuron (TDZ), with BA being the most effective growth regulator. High-frequency shoot proliferation (99%) and maximum number of shoots per explant (7.9 shoots) were recorded with BA at an optimum level of 8.9 μM. Concentrations of all cytokinins tested above the optimum level markedly reduced the frequency of shoot proliferation. A proliferating shoot culture was established by repeatedly subculturing the original cotyledonary node on shoot multiplication medium after each harvest of the newly formed shoots. Primary shoots were multiplied as nodal explants, and from each stem node 2 or 3 shoots developed. Thus, 60–70 shoots were obtained in 3 months from a single cotyledonary node. About 91% of the shoots developed roots following transfer to half-strength MS medium containing a combination of 5.7 μM indole-3-acetic acid, 4.9 μM indole-3-butyric acid and 5.3 μM indole-3-propionic acid. Eighty percent of the plantlets were successfully acclimatized and established in soil. Received: 1 October 1997 / Revision received: 31 March 1998 / Accepted: 7 April 1998     

I n Vivo Evaluation of 5-ASA Colon-Specific Tablets Using Experimental-Induced Colitis Rat Animal Model

Colonic drug delivery is intended not only for local treatment in inflammatory bowel disease (IBD) but also for systemic delivery of therapeutics. Intestinal myeloperoxidase (MPO) determination could be used to estimate the average level of inflammation in colon as well as to determine the efficacy of drugs to be used in the treatment of inflammatory bowel diseases or study the specificity of dosage forms to be used for colonic targeting of anti-inflammatory drugs. Colonic prodrug sulfasalazine (SASP) gets metabolized to give 5-aminosalicylic acid (5-ASA), which is the active portion of SASP. However, when given orally, 5-ASA is absorbed in upper part of gastrointestinal tract (GIT) and not made available in colon. In the present study, colon-targeted delivery of 5-ASA was achieved by formulating tablets with two natural polymers namely guar gum and pectin using compression coating method. Colonic specificity of 5-ASA tablets (prepared using guar gum and pectin as polymers) was evaluated in vitro using simulated fluids mimicking in vivo environment as well as in vivo method using chemically (2,4,6-trinitrobenzenesulfonic acid and acetic acid)-induced colitis rat model. Both colon-specific formulations of 5-ASA (guar gum and pectin) were observed to be more effective in reducing inflammation in chemically induced colitis rat models when compared to colon-specific prodrug sulfasalazine as well as conventional 5-ASA administered orally.KEY WORDS: colitis, colon-specific drug delivery, myeloperoxidase     

Tregs Modulate Lymphocyte Proliferation,Activation, and Resident-Memory T-Cell Accumulation within the Brain during MCMV Infection

Accumulation and retention of regulatory T-cells (Tregs) has been reported within post viral-encephalitic brains, however, the full extent to which these cells modulate neuroinflammation is yet to be elucidated. Here, we used Foxp3-DTR (diphtheria toxin receptor) knock-in transgenic mice, which upon administration of low dose diphtheria toxin (DTx) results in specific deletion of Tregs. We investigated the proliferation status of various immune cell subtypes within inflamed central nervous system (CNS) tissue. Depletion of Tregs resulted in increased proliferation of both CD8⁺ and CD4⁺ T-cell subsets within the brain at 14 d post infection (dpi) when compared to Treg-sufficient animals. At 30 dpi, while proliferation of CD8⁺ T-cells was controlled within brains of both Treg-depleted and undepleted mice, proliferation of CD4⁺ T-cells remained significantly enhanced with DTx-treatment. Previous studies have demonstrated that Treg numbers within the brain rebound following DTx treatment to even higher numbers than in untreated animals. Despite this rebound, CD8⁺ and CD4⁺ T-cells proliferated at a higher rate when compared to that of Treg-sufficient mice, thus maintaining sustained neuroinflammation. Furthermore, at 30 dpi we found the majority of CD8⁺ T-cells were CD127^hi KLRG1^- indicating that the cells were long lived memory precursor cells. These cells showed marked elevation of CD103 expression, a marker of tissue resident-memory T-cells (T_RM) in the CNS, in untreated animals when compared to DTx-treated animals suggesting that generation of T_RM is impaired upon Treg depletion. Moreover, the effector function of T_RM as indicated by granzyme B production in response to peptide re-stimulation was found to be more potent in Treg-sufficient animals. Taken together, our findings demonstrate that Tregs limit neuroinflammatory responses to viral infection by controlling cell proliferation and may direct a larger proportion of lymphocytes within the brain to be maintained as T_RM cells.