Co-transplantation of carotid body and ventral mesencephalic cells as an alternative approach towards functional restoration in 6-hydroxydopamine-lesioned rats: implications for Parkinson's disease

Exogenous administration of various neurotrophic factors has been shown to protect neurons in animal model of Parkinson's disease (PD). Several attempts are being made to search a tissue source simultaneously expressing many of these neurotrophic factors. Carotid body (CB) contains oxygen-sensitive glomus cells rich in dopamine (DA) and expresses glial cell line-derived neurotrophic factor, brain-derived neurotrophic factor and neurotrophin-3. We have attempted to study the functional restoration following co-transplantation of CB cells and ventral mesencephalic cells (VMC) in a 6-hydroxydopamine-lesioned rat model of PD. A significant recovery (p < 0.001) in d-amphetamine-induced circling behavior (80%) and spontaneous locomotor activity (85%) was evident in co-transplanted animals at 12 weeks post-transplantation as compared to lesioned animals. Similarly, a significant (p < 0.001) restoration was observed in DA-D(2) receptor binding (77%), striatal DA (87%) and 3,4-dihydroxyphenylacetic acid (DOPAC) (85%) levels and nigral DA (75%) and DOPAC (74%) levels. Functional recovery was accompanied by tyrosine hydroxylase (TH) expression and quantification of TH-positive cells by image analysis revealed a significant restoration in TH-immunoreactive (IR) fiber density in striatum, as well as TH-IR neurons in substantia nigra pars compacta in co-transplanted animals over VMC-transplanted animals. The result suggests that co-transplantation of CB cells along with VMC provides better and long-term functional restoration in the rat model of PD, possibly by supporting the survival of newly grafted cells as well as remaining host DA neurons.     

Quantitative structure-activity relationship analysis of a series of 2,3-diaryl benzopyran analogues as novel selective cyclooxygenase-2 inhibitors

A series of recently synthesized 2,3-diaryl benzopyrans reported as novel selective cycloxygenase-2 inhibitors was subjected to quantitative structure-activity relationship (QSAR) analysis. Our attempt in correlating the derived physicochemical properties with the COX-2 inhibitory activity resulted in some statistically significant QSAR models with good predictive ability. The QSAR results and the probable pharmacophore features investigated through our study explored some interesting findings for the design of potent new class of selective COX-2 inhibitors.     

Molecular cloning, phylogenetic analysis and three-dimensional modeling of Cu,Zn superoxide dismutase (CnSOD1) from three varieties of Cryptococcus neoformans

Cryptococcus neoformans (Cn), causal agent of fungal meningoencephalitis, has three varieties with variable host predilection. To explore mechanisms for these pathogenic differences, we have characterized Cu,Zn SOD gene (CnSOD1). A Saccharomyces cerevisiae sod1Delta mutant was complemented with Cn var. grubii yeast expression library. The complementing clone had an ORF of 462 bp and the deduced 154 aa sequence showed 61% identity with S. cerevisiae SOD1 and 53-65% with other eukaryotic SOD1s. Cn var. grubii CnSOD1 cDNA was used to clone corresponding cDNAs from var. neoformans and var. gattii. ORFs from three varieties revealed 20-29% differences in deduced aa (s) with a significant 6% non-synonymous aa substitution between Cn var. grubii and Cn var. gattii. Cosmid library screening and PCR cloning were used to obtain genomic SOD1, which was split by five introns with identical placements and a typical 5' splice junction sequence, GTNNGY. These introns also showed a large nt variation among the three Cn varieties. Phylogenetic analyses revealed CnSOD1 to be in a group distinct from other eukaryotic SOD1s and with a significant divergence of the var. grubii from var. gattii. The CnSOD1 -deduced protein was modeled based on the crystal structure of S. cerevisiae SOD1, which showed an excellent fit. Most of the non-synonymous aa substitutions occurred on the outside of the molecule and these may contribute to differences in antigenicity among the three varieties. Notably, Cn var. neoformans and var. gattii Cu,Zn SOD had three substitutions of glycine (Gly26, Gly92 and Gly123 for Asn26, Ser92 and Ser123) that may contribute to the observed lower thermostability of this enzyme vis-a-vis Cn var. grubii. This is the first nucleotide and structural comparison of a protein-encoding gene from the three Cn varieties, which may provide a framework for future studies on the role of Cu,Zn SOD in Cn pathogenesis.     

Structure and regulation of opioid receptors

Significant advances have been made in understanding the structure, function, and regulation of opioid receptors and endogenous opioid peptides since their discovery approximately 25 years ago. This review summarizes recent studies aimed at identifying key amino acids that confer ligand selectivity to the opioid receptors and that are critical constituents of the ligand binding sites. A molecular model of the delta receptor based on the crystal structure of rhodopsin is presented. Agonist-induced down regulation of opioid receptors is discussed, highlighting recent evidence for the involvement of the ubiquitin/proteasome system in this process.     

Antioxidant property of Smilex china Linn

Smilex china Linn. (Smilacaceae Syn Liliaceae) has special pharmacognostic feature as its root is multiseriate, with sclerenchymatic pericycle. The alcoholic extract of rhizome of S. china shows significant protection against FeSO4 induced lipid peroxidation in rat liver homogenate, but has no effect on the rate of oxidation of reduced glutathione. This fraction scavenges the superoxide and hydroxyl radicals, but the effect was more towards the removal of superoxide than that of hydroxyl radicals. Thus it could be concluded that rhizome of S. china has strong antioxidant property.     

In vitro high frequency regeneration of plantlets of Vigna mungo and their ex vitro growth

Of the five explants of V. mungo var. T9 used, the excised shoot tips gave best response with regard to offshoot formation followed by the embryonal axis explants. While a treatment comprising 0.5 mgL(-1) BAP, 0.5 mgL(-1) 2iP and 0.1 mgL(-1) NAA induced differentiation of an average 10 offshoots in shoot tip explants, only 3 offshoots were formed in the explants of embryonal axis in a treatment containing 0.5 mgL(-1) BAP and 0.1 mgL(-1) NAA, found optimum for them. Multiple shoots differentiated when explants with earlier regenerated and growing offshoots were first cultured in a treatment containing 0.1 mgL(-1) BAP, 0.25 mgL(-1) IAA and 5 mgL(-1) CCC and then subcultured in the same treatment but having only 1 mgL(-1) CCC. The isolated shoots rooted in 0.5 mgL(-1) IAA resulted in the formation of complete plantlets of an average height of 15 cm in 20 days. The in vitro-regenerated plants grew normally under field conditions and came to flowering as well.     

Toxoplasma gondii ADP-ribosylation factor 1 mediates enhanced release of constitutively secreted dense granule proteins

Toxoplasma gondii dense granules are morphologically similar to dense matrix granules in specialized secretory cells, yet are secreted in a constitutive, calcium-independent fashion. We previously demonstrated that secretion of dense granule proteins in permeabilized parasites was augmented by the non-hydrolyzable GTP analogue guanosine 5'-3-O-(thio)triphosphate (GTPgammaS) (Chaturvedi, S., Qi, H., Coleman, D. L., Hanson, P., Rodriguez, A., and Joiner, K. A. (1998) J. Biol. Chem. 274, 2424-2431). As now demonstrated by pharmacological and electron microscopic approaches, GTPgammaS enhanced release of dense granule proteins in the permeabilized cell system. To investigate the role of ADP-ribosylation factor 1 (ARF1) in this process, a cDNA encoding T. gondii ARF1 (TgARF1) was isolated. Endogenous and transgenic TgARF1 localized to the Golgi of T. gondii, but not to dense granules. An epitope-tagged mutant of TgARF1 predicted to be impaired in GTP hydrolysis (Q71L) partially dispersed the Golgi signal, with localization to scattered vesicles, whereas a mutant impaired in nucleotide binding (T31N) was cytosolic in location. Both mutants caused partial dispersion of a Golgi/trans-Golgi network marker. TgARF1 mutants inhibited delivery of the secretory reporter, Escherichia coli alkaline phosphatase, to dense granules, precluding an in vivo assessment of the role of TgARF1 in release of intact dense granules. To circumvent this limitation, recombinant TgARF1 was purified using two separate approaches, and used in the permeabilized cell assay. TgARF1 protein purified on a Cibacron G3 column and able to bind GTP stimulated dense granule secretion in the permeabilized cell secretion assay. These results are the first to show that ARF1 can augment release of constitutively secreted vesicles at the target membrane.     

Role of interleukin-12 in patients with dengue hemorrhagic fever

Interleukin (IL)-12 has a broad range of activities including regulation of cytokine synthesis and selective promotion of Th1-type cell development. A shift from a Th1-type response to Th2-type has been suggested to be important in the pathogenesis of dengue hemorrhagic fever (DHF). This study was undertaken to investigate the possible role of IL-12 in this shift. A total of 76 patients with various grades of dengue illness and 21 normal healthy controls were tested for IL-12 levels in serum samples and IL-12 mRNA in their peripheral blood mononuclear cells. The results showed that the levels of IL-12 were the highest in patients with dengue fever (270+/-102 pg ml(-1)) followed by decreasing levels in the patients with DHF grade I (198+/-86 pg ml(-1); P<0.05) and DHF grade II (84+/-52 pg ml(-1); P<0.001). Neither IL-12 nor its mRNA could be detected in the patients with DHF grades III and IV. The cytokine appeared and reached peak levels during the first 4 days of illness, started to decline by day 5-8 and disappeared by day 9 onwards. The absence of IL-12 during severe illness and late phases of the disease may be responsible for the shift to a Th2-type response and thus for the pathogenesis of DHF.     

Phytostabilization of iron ore tailings through Calophyllum inophyllum L

The phytostabilization of waste material generated during mining and processing of iron ore through Calophyllum inophyllum L. have been investigated. Iron ore tailings and its varying composition with garden soil were taken to study plant growth, chlorophyll content and metal uptake pattern of Calophyllum inophyllum L. These studies indicate that 100% survival of plant species was noted in all the treatments without any toxicity symptoms. The increase in growth parameters and chlorophyll content along with the high metal accumulation in plant tissues suggests that Calophyllum inophyllum L. may be a potential tool for phytoremediation. The accumulation of Pb (1662 microgm/gm) and Fe (2313 microgm/gm) was observed to be maximum in the plant tissues followed by Cu, Zn, Cr, and Ni. The TF values for most of the heavy metals was observed to be > 1 which indicates that the plant can efficiently translocate these toxic metals to its above ground parts. Removal of more than 30% of the most of the heavy metal like Fe, Pb, and Cu & Zn has been observed in all the treatments during one year of observation. The overall study clearly suggests that the plant can be used as an efficient tool for restoration of mining wastes and other similarly contaminated sites.     

Evaluation of nutrient uptake and physical parameters on cell biomass growth and production of spilanthol in suspension cultures of Spilanthes acmella Murr.

Spilanthes acmella Murr. has a plethora of highly valuable biologically active compounds and has been listed as one of the important medicinal plants of the world. However, no perceptible biotechnological advances have been made for this genus to exploit or enhance its utility. To nullify the effect of seasonal variations, the present report is the first attempt to establish in vitro cell suspension cultures and to evaluate the production of spilanthol from them, a key component of the plant responsible for most of its pharmaceutical activities. The study examined the biomass growth in relation to the consumption of major nutrients and sucrose, agitation speed and dynamic change in pH. Results indicated that the consumption of phosphate resulted in the onset of decline phase in cultures. Spilanthol production was observed to be growth associated and maximum production occurred on the 15th day. Among the carbon sources, the highest production of spilanthol as 91.4 μg g(-1) DW was recorded in the medium supplemented with sucrose, followed by glucose which produced 56.8 μg g(-1) DW of spilanthol. Spilanthol could not be detected in fructose containing medium. Maximum viable cultures were obtained at a rotation speed of cells at 120 rpm. This study signifies the utility of Spilanthes suspension cultures for biosynthesis and constant production of spilanthol, throughout the year. The results of present study are useful for further scale-up process.