In vitro multiplication of Quercus leucotrichophora and Q. glauca: Important Himalayan oaks

Multiple shoots of Quercus leucotrichophora L. and Q. glauca Thunb. were induced from the intact embryos (decoated seeds) as well as from the cotyledonary nodes (with attached cotyledons but without radicle and primary shoot) of 3-weeks old in vitro grown seedlings on Woody Plant (WP; Lloyd and McCown, 1980) and Murashige and Skoog (MS; 1962) media supplemented with 6-benzyladenine (BA), either alone or in combination with gibberellic acid (GA₃)/ indole-3-butyric acid (IBA). BA (22.19 M) was effective for induction of multiple shoots and addition of GA₃ to the medium further enhanced the shoot number and shoot height but resulted in shoot thinness. High frequency shoot multiplication was achieved using cotyledonary nodes. Shoots were further multiplied from the original explant on WP medium supplemented with BA (22.19 M). Nearly 78% and 67% rooting was obtained in Q. leucotrichophora and Q. glauca microshoots (3–4 cm high), respectively on 1/2 strength WP medium supplemented with IBA (14.76 M). However, this was associated with basal callus formation. Treatment with IBA (25–100 M) for 24 or 48 h followed by transfer to PGR free 1/2 strength WP medium not only improved the rooting percentage but also avoided basal callus formation. IBA at 100 M for 24 h was most effective (90% and 100% rooting in Q. leucotrichophora and Q. glauca, respectively). In vitro rooted plants were hardened and established in garden soil.Growth performance of 6-month-old in vitro raised plants was compared with ex vitro plants (seedlings) of the same age. The photosynthesis and transpiration rates of eight months old in vitro and ex vitro raised plants of both species were measured under different light (0, 600, 900, 1200, 1500 and 2000 mol m^–2s^–1) and temperature (20, 25, 30, 35 and 40 °C). Light optimum for photosynthesis was around 2000 mol m^–2s^–1 in Q. leucotrichophora and around 1500 mol m^–2s^–1 in Q. glauca whereas optimum temperature for photosynthesis was 25 °C in Q. leucotrichophora and 30 °C in Q. glauca. The rate of transpiration at different temperatures (20–40 °C), in the two species, increased with increase in the light intensity up to the highest level, i.e., 2000 mol m^–2s^–1. Temperatures beyond 35 °C adversely affected the rate of transpiration in in vitro raised as well as ex vitro plants of both the species. In vitro raised and hardened plants of both the species were comparable to ex vitro plants in terms of gas and water vapour exchange characteristics, within the limits of this study.     

Effect of nimesulide on acetic acid- and leukotriene-induced inflammatory bowel disease in rats

Inflammatory bowel disease (IBD) is a relapsing inflammation of intestine, which is mediated by release of inflammatory mediators. Both cyclo-oxygenase product prostaglandin (PGE2) and lipo-oxygenase product leukotriene (LTB4), may contribute to the pathogenesis of the inflammatory response. Nimesulide, a preferential COX-2 inhibitor was evaluated for its efficacy against experimental colitis in two different models (acetic acid- and LTB4-induced IBD) in rats. Inflammatory response was induced by intrarectal single administration of acetic acid or LTB4. Nimesulide (9 and 18 mg/kg, p.o.) significantly prevented development of inflammatory changes, decreased myeloperoxidase (MPO) activity, and also restored the altered contractility response of the isolated colon segment to KCl. The results suggested the involvement of both cyclo-oxygenase (COX) and lipo-oxygenase-mediated proinflammatory agents in colonic inflammatory process associated with IBD. Further, this study suggests that such therapeutic interventions may be of value in the treatment of IBD.     

Entrainment of circadian locomotor activity rhythm of the nocturnal field mouse Mus booduga using daily injections of melatonin

In this paper, we report the effects of daily injections of melatonin on the locomotor activity rhythm of the nocturnal field mouse Mus booduga. The locomotor activity rhythm of 45 animals was first monitored in constant darkness (DD) of the laboratory for about 15 days. The animals were then divided into three groups (experimental, vehicle-treated control, and the nontreated control groups) and subjected to three different treatments. The animals from the experimental group (n=19) were administered daily a single subcutaneous (s.c.) injection of melatonin (1 mg/kg) for about 45 days. The vehicle treated controls (n=13) were administered daily injections of 50% dimethyl sulfoxide (DMSO) for about 45 days, and the nontreated controls (n=13) were handled similar to the other two groups without being administered injections. Following the treatments, the animals were maintained in DD for about 20 days, after which the experiments were terminated. A significantly larger percentage of animals from the experimental group either entrained or showed phase control to daily treatments, compared to the animals from the two control groups. These results suggest that externally administered melatonin can influence the phase of the circadian locomotor activity rhythm of M. booduga. The fact that none of the nontreated controls showed any sign of phase control to daily handling, clearly demonstrates that the entrainment or phase control in the melatonin treated group of animals is caused by melatonin alone and not due to handling.     

Validation of the Hsp150 polypeptide carrier and HSP150 promoter in expression of rat alpha2,3-sialyltransferase in yeasts

Heterologous glycoproteins usually do not fold properly in yeast cells and fail to leave the endoplasmic reticulum. Here we show that the Hsp150Delta polypeptide carrier promoted proper folding and secretion of the catalytic ectodomain of rat alpha2,3-sialyltransferase (ST3Ne) in Pichia pastoris. The efficiency of the Hsp150Delta carrier in P. pastoris and Saccharomyces cerevisiae was at least as high as that of the MFalpha carrier. Most of Hsp150Delta-ST3Ne and MFalpha-ST3Ne remained noncovalently attached to the cell wall via the ST3Ne portion. The strength of the HSP150 promoter was found to be comparable to that of the GAL1 promoter.     

Degradation of 2,4,6-Trichlorophenol by Phanerochaete chrysosporium: Involvement of Reductive Dechlorination

Under secondary metabolic conditions, the lignin-degrading basidiomycete Phanerochaete chrysosporium mineralizes 2,4,6-trichlorophenol. The pathway for the degradation of 2,4,6-trichlorophenol has been elucidated by the characterization of fungal metabolites and oxidation products generated by purified lignin peroxidase (LiP) and manganese peroxidase (MnP). The multistep pathway is initiated by a LiP- or MnP-catalyzed oxidative dechlorination reaction to produce 2,6-dichloro-1,4-benzoquinone. The quinone is reduced to 2,6-dichloro-1,4-dihydroxybenzene, which is reductively dechlorinated to yield 2-chloro-1,4-dihydroxybenzene. The latter is degraded further by one of two parallel pathways: it either undergoes further reductive dechlorination to yield 1,4-hydroquinone, which is ortho-hydroxylated to produce 1,2,4-trihydroxybenzene, or is hydroxylated to yield 5-chloro-1,2,4-trihydroxybenzene, which is reductively dechlorinated to produce the common key metabolite 1,2,4-trihydroxybenzene. Presumably, the latter is ring cleaved with subsequent degradation to CO₂. In this pathway, the chlorine at C-4 is oxidatively dechlorinated, whereas the other chlorines are removed by a reductive process in which chlorine is replaced by hydrogen. Apparently, all three chlorine atoms are removed prior to ring cleavage. To our knowledge, this is the first reported example of aromatic reductive dechlorination by a eukaryote.     

Phospholipids of wheat chloroplast and its membranes under water stress

Phospholipids showed a differential change in the chloroplast membranes in two cultivars under water stress. Amongst the individual phospholipids, phosphatidyl choline (PC) increased under stress in the low water requiring cultivar C-306 but it decreased in high water requiring cultivar S-308. PC of chloroplast envelope and chloroplast thylakoids showed similar response. Increase in PC content in chloroplasts and its membranes of resistant cultivar may suggest a basis for stress resistance.     

Vasoactive Intestinal Polypeptide Promotes Intestinal Barrier Homeostasis and Protection Against Colitis in Mice

Inflammatory bowel disease is a chronic gastrointestinal inflammatory disorder associated with changes in neuropeptide expression and function, including vasoactive intestinal peptide (VIP). VIP regulates intestinal vasomotor and secretomotor function and motility; however, VIP’s role in development and maintenance of colonic epithelial barrier homeostasis is unclear. Using VIP deficient (VIPKO) mice, we investigated VIP’s role in epithelial barrier homeostasis, and susceptibility to colitis. Colonic crypt morphology and epithelial barrier homeostasis were assessed in wildtype (WT) and VIPKO mice, at baseline. Colitic responses were evaluated following dinitrobenzene sulfonic acid (DNBS) or dextran-sodium sulfate (DSS) exposure. Mice were also treated with exogenous VIP. At baseline, VIPKO mice exhibited distorted colonic crypts, defects in epithelial cell proliferation and migration, increased apoptosis, and altered permeability. VIPKO mice also displayed reduced goblet cell numbers, and reduced expression of secreted goblet cell factors mucin 2 and trefoil factor 3. These changes were associated with reduced expression of caudal type homeobox 2 (Cdx2), a master regulator of intestinal function and homeostasis. DNBS and DSS-induced colitis were more severe in VIPKO than WT mice. VIP treatment rescued the phenotype, protecting VIPKO mice against DSS colitis, with results comparable to WT mice. In conclusion, VIP plays a crucial role in the development and maintenance of colonic epithelial barrier integrity under physiological conditions and promotes epithelial repair and homeostasis during colitis.     

Quality of Artemisinin-Based Combination Formulations for Malaria Treatment: Prevalence and Risk Factors for Poor Quality Medicines in Public Facilities and Private Sector Drug Outlets in Enugu,Nigeria

BackgroundArtemisinin-based combination therapies are recommended by the World Health Organisation (WHO) as first-line treatment for Plasmodium falciparum malaria, yet medication must be of good quality for efficacious treatment. A recent meta-analysis reported 35% (796/2,296) of antimalarial drug samples from 21 Sub-Saharan African countries, purchased from outlets predominantly using convenience sampling, failed chemical content analysis. We used three sampling strategies to purchase artemisinin-containing antimalarials (ACAs) in Enugu metropolis, Nigeria, and compared the resulting quality estimates.MethodsACAs were purchased using three sampling approaches - convenience, mystery clients and overt, within a defined area and sampling frame in Enugu metropolis. The active pharmaceutical ingredients were assessed using high-performance liquid chromatography and confirmed by mass spectrometry at three independent laboratories. Results were expressed as percentage of APIs stated on the packaging and used to categorise each sample as acceptable quality, substandard, degraded, or falsified.ResultsContent analysis of 3024 samples purchased from 421 outlets using convenience (n=200), mystery (n=1,919) and overt (n=905) approaches, showed overall 90.8% ACAs to be of acceptable quality, 6.8% substandard, 1.3% degraded and 1.2% falsified. Convenience sampling yielded a significantly higher prevalence of poor quality ACAs, but was not evident by the mystery and overt sampling strategies both of which yielded results that were comparable between each other. Artesunate (n=135; 4 falsified) and dihydroartemisinin (n=14) monotherapy tablets, not recommended by WHO, were also identified.ConclusionRandomised sampling identified fewer falsified ACAs than previously reported by convenience approaches. Our findings emphasise the need for specific consideration to be given to sampling frame and sampling approach if representative information on drug quality is to be obtained.     

Sumoylation Influences DNA Break Repair Partly by Increasing the Solubility of a Conserved End Resection Protein

Protein modifications regulate both DNA repair levels and pathway choice. How each modification achieves regulatory effects and how different modifications collaborate with each other are important questions to be answered. Here, we show that sumoylation regulates double-strand break repair partly by modifying the end resection factor Sae2. This modification is conserved from yeast to humans, and is induced by DNA damage. We mapped the sumoylation site of Sae2 to a single lysine in its self-association domain. Abolishing Sae2 sumoylation by mutating this lysine to arginine impaired Sae2 function in the processing and repair of multiple types of DNA breaks. We found that Sae2 sumoylation occurs independently of its phosphorylation, and the two modifications act in synergy to increase soluble forms of Sae2. We also provide evidence that sumoylation of the Sae2-binding nuclease, the Mre11-Rad50-Xrs2 complex, further increases end resection. These findings reveal a novel role for sumoylation in DNA repair by regulating the solubility of an end resection factor. They also show that collaboration between different modifications and among multiple substrates leads to a stronger biological effect.