Immunomodulatory studies of a bioactive fraction from the fruit of Prunus cerasus in BALB/c mice

In order to evaluate the role of ethyl acetate fraction (PNRS-EtOAC) obtained from the Prunus cerasus fruit in the modulation of immune responses, detailed studies were carried out using a panel of in vivo assays. Oral administration of PNRS-EtOAC (25-100 mg/kg) stimulated the IgM and IgG titre expressed in the form of hemagglutination antibody (HA) titre. Further, it elicited a dose related increase in the delayed type hypersensitivity reaction (DTH) after 24 and 48 h in BALB/c mice. Besides augmenting the humoral and cell mediated immune response, the concentration of cytokines (IFN-γ, IL-4, and TNF-α) in serum with respect to T cell interactions, i.e. the proliferation of lymphocytes were significantly increased at 50 mg/kg compared with the control. The results in these studies demonstrated the immunostimulatory effect of PNRS-EtOAC in a dose-dependent manner with respect to the macrophage activation possibly expressing the phagocytosis and nitrite production by the enhancement of TNF-α production as a mode of action.     

Brain tissue energy dependence of CaM kinase IV cascade activation during hypoxia in the cerebral cortex of newborn piglets

Using an anti-cue keypress task, we examined executive control in Parkinson's disease (PD) patients treated with deep brain stimulation (DBS) of the subthalamic nucleus (STN) and dopaminergic medication. Across sessions, we varied stimulation (on, off) and dopaminergic medication (on, off). Reaction time (RT) results of the PD patients and their age-matched controls showed a consistent pattern of RT costs and benefits generated by anti-cues with short and long preparation intervals, respectively. This pattern was evident in all sessions, except when DBS stimulation and medication were off. In this condition PD patients showed no RT benefits. These findings are discussed in terms of an executive control process that suppresses the automatic but inappropriate response activation generated by anti-cues. In PD this mechanism is severely compromised but it can be remediated by dopaminergic medication and DBS, suggesting an essential role of the basal ganglia in the selection and suppression of competing responses.     

Identification of a major pathogenicity determinant and suppressors of RNA silencing encoded by a South Pacific isolate of <Emphasis Type="Italic">Banana bunchy top virus</Emphasis> originating from Pakistan

Five genes encoded by Banana bunchy top virus (BBTV) originating from Pakistan were expressed in Nicotiana benthamiana using a Potato virus X (PVX) vector. Expression of the master replication-associated protein (mRep) and movement protein (MP) resulted in necrotic cell death of inoculated tissues, as well as leaf curling and necrosis along the veins in newly emerging leaves. The systemic necrosis induced by the expression of MP was discolored (dark) in comparison to that induced by mRep. Expression of the cell-cycle link protein (Clink), the coat protein (CP), and the nuclear shuttle protein from the PVX vector induced somewhat milder symptoms, consisting of mild leaf curling and mosaic, although expression of the CP caused a necrotic response in inoculated leaf. The accumulation of viral RNA was enhanced by MP, Clink, and CP. Of the five BBTV-encoded gene products two, the MP and Clink, stabilized GFP-specific mRNA and reduced GFP-specific small interfering RNA in N. benthamiana line 16c when expressed under the control of the 35S promoter and co-inoculated with a construct for the expression of GFP hairpin RNA construct. These results identified MP and Clink as suppressors of RNA silencing. Taken together the ability of MP to induce severe symptoms in plants and suppress RNA silencing implicates this product as a major pathogenicity determinant of BBTV.     

NO-mediated activation of Src kinase during hypoxia in the cerebral cortex of newborn piglets

The present study aims to investigate the mechanism of Src kinase activation during hypoxia and tests the hypothesis that the hypoxia-induced activation of Src kinase, as determined by Src kinase phosphorylation, in the cerebral cortical membranes of newborn piglets is mediated by NO derived from neuronal nitric oxide synthase (nNOS). Fifteen piglets were divided into normoxic (Nx, n = 5), hypoxic (Hx, n = 5) and hypoxic-treated with nNOS inhibitor I (Hx-nNOSi) groups. Hypoxia was induced by decreasing FiO₂ to 0.06 for 1 h. nNOS inhibitor I (selectivity >2500 vs eNOS and >500 vs iNOS) was administered (0.4 mg/kg, i.v.) 30 min prior to hypoxia. Cortical membranes were isolated and phosphorylation of Src kinase was determined by Western blot analysis. Src kinase activity was determined by radioactive assay using immunopurified enzyme. Membrane proteins were separated by 12% SDS–PAGE and probed with anti-phospho (pTyr⁴¹⁸)-Src kinase antibody. Protein bands were detected, analyzed by densitometry and expressed as absorbance (OD × mm²). Density (OD × mm²) of phosphorylated Src kinase was 111.7 ± 21.1 in Nx, 234.5 ± 23.8 in Hx (p < 0.05 vs Nx) and 104.7 ± 18.1 in Hx-nNOSi (p < 0.05 vs Hx, p = NS vs Nx). Src kinase activity (pmol/mg protein/ h) was 2472 ± 75 in Nx, 4556 ± 358 in Hx (p < 0.05 vs Nx) and 2259 ± 207 in Hx-nNOSi (p < 0.05 vs Hx, p = NS vs Nx). The data show that pretreatment with nNOS inhibitor prevents the hypoxia-induced increase in tyrosine phosphorylation and the activity of Src kinase. We conclude that the mechanism of hypoxia-induced increased activation of Src kinase is mediated by nNOS derived NO. We propose that NO mediated inhibition of protein tyrosine phosphatases SH-PTP-1 and SH-PTP-2 leads to increased tyrosine phosphorylation and activation of Src kinase in the cerebral cortex of newborn piglets.     

Effect of altitude on picroside content in core collections of Picrorhiza kurrooa from the north western Himalayas

Picrorhiza kurrooa Royle ex Benth (Scrophulariaceae), commonly known as Kutki, is a major ingredient of many ayurvedic preparations prescribed in the treatment of various diseases. Picrosides I and II are the active agents responsible for the medicinal effects of Kutki, and the variation in content of these compounds in plants at different altitudes is a major question to be addressed. The picroside I and II content in various plant parts of P. kurrooa collected from different altitudes, viz. Sonemarg (2,740 m a.s.l.), Tangmarg (2,690 m a.s.l.), and Pulwama (1,630 m a.s.l.) in the north-western Kashmir Himalayas was analyzed by HPLC. A considerable degree of variation in picroside content was observed. Picroside I and II was highest in populations collected from Sonemarg followed by Tangmarg, suggesting that picroside accumulation is directly correlated with altitudinal change. More picroside I was found in the rhizome and roots of the Pulwama population as compared to Tangmarg samples, whereas the quantity of Picroside II was reduced in plants from Pulwama compared to the Tangmarg population, suggesting that cultivation of P. kurroa at lower altitude reduces the picroside content. The quantities of picrosides also varied spatially, being highest in rhizome followed by roots, inflorescence and leaves in the populations from all three locations. The study concludes that picroside I and II accumulation depends on altitude, which could help in the selection and collection of superior genotypes with uniform effects for utilization by the pharmaceutical industry.     

The atrophy and changes in the cellular compositions of the thymus and spleen observed in mice subjected to short-term exposure to perfluorooctanesulfonate are high-dose phenomena mediated in part by peroxisome proliferator-activated receptor-alpha (PPARα)

We have previously shown that short-term, high-dose exposure of mice to the environmentally persistent perfluorooctanoate (PFOA) results in thymic and splenic atrophy and the attenuation of specific humoral immune responses. Here we characterize the effects of a 10-day treatment with different dietary doses (1–0.001%, w/w) of perfluorooctanesulfonate (PFOS), a similar fluorochemical, on the immune system of male C57BL/6 mice. At doses greater than 0.02%, PFOS induced clinical signs of toxicity in the animals, whereas at the concentration of 0.02%, this compound caused weight loss, hepatomegaly and atrophy of the thymus, spleen and adipose tissue without toxicity. With this latter dose, histopathological and flow-cytometric analysis revealed that (i) the thymic cortex was virtually depleted of cells; (ii) the total numbers of thymocytes and splenocytes were reduced by 84 and 43%, respectively; (iii) although all populations of thymocytes and splenocytes were smaller, the thymic CD4⁺CD8⁺ cells and the splenic B-lymphocytes were most decreased. These alterations resembled those evoked by analogous exposure to PFOA, but were less pronounced. At lower doses (less than 0.02%), PFOS induced hepatomegaly without affecting the thymus or spleen. Finally, comparison of male wild-type 129/Sv mice and the corresponding knock-outs lacking peroxisome proliferator-activated receptor-alpha (PPARα) indicated that these effects of PFOS are not strain-dependent. More importantly, hepatomegaly is independent of PPARα, the thymic changes are partially dependent on this receptor, and splenic responses are largely eliminated in its absence. Thus, immunomodulation caused by PFOS is a high-dose phenomenon partially dependent on PPARα.     

Bladder flap posterior urethroplasty (Faseeh's technique)

Posterior urethral distraction defects are common nowadays because of increased incidence of road traffic accidents (RTA) and FALL. Approximately 90% are associated with pelvic fracture. Initial suprapubic cystostomy followed by delayed single stage end-to-end perineal urethroplasty, in most of these cases, is current mode of treatment. We feel the most important factor in determination of outcome is the length of distraction defect of the posterior urethra. In many cases, the gap between the two stumps exceeds 05 cms. This, coupled with the distorted anatomy of pelvis, can render end-to-end anastomoses impossible, despite all the claims made in literature. We have used an attached bladder flap to overcome this difficult situation. The results of the procedure evaluated by Uroflowmetry (UFM) micturating cystourethrogram and urethroscopy are very encouraging. We report this new technique for further evaluation.