Buffalo (Bubalus bubalis) epiphyseal proteins give protection from arsenic and fluoride-induced adverse changes in acetylcholinesterase activity in rats

The objective of this study was to determine the effect of fluoride (F) and arsenic (As) on the activity of acetylcholinesterase (AChE), a critically important nervous system enzyme, and to test the protective role of buffalo epiphyseal (pineal) proteins (BEP) in rats. Arsenic (20 mg/kg BW, intraperitoneally) and F (150 ppm, perorally) were exposed, and BEP was administered intraperitoneally (100 μ g/kg BW) along with F and As to rats for 7 days. As and F exposure significantly (p < 0.05) increased their levels in plasma and decreased the activity of AChE in plasma, RBCs, heart, and brain of rats. Interestingly, As- and F-induced inhibition of AChE activities increased As and F levels in plasma, and organs were significantly (p < 0.05) counteracted by BEP administration. These findings indicate the protective role of buffalo (Bubalus bubalis) epiphyseal proteins on F- and As-induced adverse changes in AChE activity as a candidate biomarker for neurotoxicity in female rats.     

Phage-displayed antibody libraries of synthetic heavy chain complementarity determining regions

A structure-based approach was used to design libraries of synthetic heavy chain complementarity determining regions (CDRs). The CDR libraries were displayed as either monovalent or bivalent single-chain variable fragments (scFvs) with a single heavy chain variable domain scaffold and a fixed light chain variable domain. Using the structure of a parent antibody as a guide, we restricted library diversity to CDR positions with significant exposure to solvent. We introduced diversity with tailored degenerate codons that ideally only encoded for amino acids commonly observed in natural antibody CDRs. With these design principles, we reasoned that we would produce libraries of diverse solvent-exposed surfaces displayed on stable scaffolds with minimal structural perturbations. The libraries were sorted against a panel of proteins and yielded multiple unique binding clones against all six antigens tested. The bivalent library yielded numerous unique sequences, while the monovalent library yielded fewer unique clones. Selected scFvs were converted to the Fab format, and the purified Fab proteins retained high affinity for antigen. The results support the view that synthetic heavy chain diversity alone may be sufficient for the generation of high-affinity antibodies from phage-displayed libraries; thus, it may be possible to dispense with the light chain altogether, as is the case in natural camelid immunoglobulins.     

Effect of simazine and atrazine on the mineralization of fertilizer and manure nitrogen

Summary Laboratory experiments were carried out with alluvial sandy loam soil to study the effect of simazine and atrazine herbicides at four levels (0.5, 1.0, 1.5 and 2.0 kg/ha) on the mineralization of nitrogen (ammoniacal and nitrate production) from fertilizer urea and sludge sources. The herbicides stimulated nitrate production. No specific trend in total mineralized nitrogen, ammoniacal and nitrate nitrogen was observed by varying the levels of herbicides. Mineralization of total nitrogen (ammoniacal and nitrate nitrogen) in presence of simazine and atrazine from the different sources in the descending order was:Urea > Sludge + Urea > Sludge > No Nitrogen.     

Up regulation of the GRP-78 and GADD-153 and down regulation of Bcl-2 proteins in primary glomerular diseases: a possible involvement of the ER stress pathway in glomerulonephritis

Alpha-lipoic acid (LA) has recently been reported to afford protection against neurodegenerative disorders in humans and experimental animals. However, the mechanisms underlying LA-mediated neuroprotection remain an enigma. Because peroxynitrite has been extensively implicated in the pathogenesis of various forms of neurodegenerative disorders, this study was undertaken to investigate the effects of LA in peroxynitrite-induced DNA strand breaks, a critical event leading to peroxynitrite-elicited cytotoxicity. Incubation of φX-174 plasmid DNA with the 3-morpholinosydnonimine (SIN-1), a peroxynitrite generator, led to the formation of both single- and double-stranded DNA breaks in a concentration- and time-dependent fashion. The presence of LA at 100–1,600 μM was found to significantly inhibit SIN-1-induced DNA strand breaks in a concentration-dependent manner. The consumption of oxygen induced by 250 μM SIN-1 was found to be decreased in the presence of high concentrations of LA (400–1,600 μM), indicating that LA at these concentrations may affect the generation of peroxynitrite from auto-oxidation of SIN-1. It is observed that incubation of the plasmid DNA with authentic peroxynitrite resulted in a significant formation of DNA strand breaks, which could also be dramatically inhibited by the presence of LA (100–1,600 μM). EPR spectroscopy in combination with spin-trapping experiments, using 5,5-dimethylpyrroline-N-oxide (DMPO) as spin trap, resulted in the formation of DMPO-hydroxyl radical adduct (DMPO-OH) from authentic peroxynitrite and LA at 50–1,600 μM inhibited the adduct signal. Taken together, these studies demonstrate for the first time that LA can potently inhibit peroxynitrite-mediated DNA strand breakage and hydroxyl radical formation. In view of the critical involvement of peroxynitrite in the pathogenesis of various neurodegenerative diseases, the inhibition of peroxynitrite-mediated DNA damage by LA may be responsible, at least partially, for its neuroprotective activities.     

Metamorphosis of the central nervous system ofDrosophila melanogaster Meigen (Diptera: Drosophilidae) during pupation

We studied the metamorphosis of the central nervous system (CNS) and neighbouring muscles ofDrosophila melanogaster (Diptera: Drosophilidae) during pupation by transmission electron microscopy (TEM). The age of white pupa was assumed to be 0 h and the process of metamorphosis was monitored, onward between 6 and 96 h at 25°C. The profiles in the neuropil showed degeneration at 6 h and its extent increased by 12 h. The presence of glycogen in some of these profiles indicated their larval character. Between 12–18 h, the neuronal profiles became separated from one another, the intervening space was filled with extracellular fluid, and some of the larval synapses degenerated. Synaptic vesicles started reappearing around 18 h and synapses were detectable by 24 h. Neuronal processes compactly filled the neuropil by 65 h and the maturation of synapses continued until 86 h. The degeneration of profiles in the neuropil was found to be bimodal, peaking at 12 and 42 h, and that of cortical cells was unimodal with a peak at 42 h. The number of neuronal profiles increased with the development time, indicating that more branching of neuronal profiles occurs in neuropils as the metamorphosis progresses. Average number of synapses per unit area (or volume) is minimum at 18 h and maximum at 72 h, when the average number of synapse per axon profile is 0.54. Because 2 axon profiles share one synapse, a value close to 0.5 for monad synapses shows that, on an average, each axon profile at least makes one synapse at this stage of development. Subsequently, there is more than 75% of reduction in the number of synapses during 73 and 78 h. In muscles, vacuoles suggesting histolysis appeared by 6 h. Their ultrastructure became deranged between 12–18 h and myoblasts were found to be present since 8 h. Except for a few muscles in the thorax, such as larval oblique muscles and pharyngeal muscles, most of the muscles in the head and thorax lost all the ultrastructural details and histolyzed by 18 h. Around 38 h, imaginai muscles were detectable, and well-developed muscles were found by 55 h. However, myofibrils continued to be added laterally to the preformed muscles even at 96 h. Electron-dense mitochondria (EDMITs) were found in the neuropil, cortex and muscles of pupa, along with mitochondria of characteristic shape and normal appearance. These EDMITs often occurred in large clusters of more than 100, at times near the surface of the tissue. A few of these were enclosed in vacuoles and were darker than the rest of the EDMITs and normal looking mitochondria. Histochemistry with diaminobenzidine showed the presence of cytochrome c and marker enzyme cytochrome oxidase, both in EDMITs and normal mitochondria. EDMITs were not found to be present in any tissue of the adultDrosophila. A preliminary report of the work was presented at the International Conference on Neurobiology at Goa in 1991 and appeared inNervous Systems, Principles of Design and Function (ed.) R N Singh (New Delhi: Wiley Eastern) pp 91–105 (1992).