Efficient plant regeneration of the endangered medicinal orchid, <Emphasis Type="Italic">Coelogyne cristata</Emphasis> using protocorm-like bodies

An efficient method of Coelogyne cristata mass propagation was developed using segment of protocorm-like bodies (PLBs) (3 mm² in size). It was observed that ½ MS medium showed to be more effective to induce shoots through PLBs segment. The explants when cultured on ½ MS media containing TDZ and CP showed relatively superior effect on shoot regeneration as compared to the media containing TDZ alone or in combination with BP. Addition of BP and CP to the medium containing NAA and BA combinations proved distinctly better for shoot multiplication than that of the medium with NAA and BA combinations alone. The highest percentage of explants producing shoots, with a maximum average of 8.1 per explant, was induced on the medium supplemented with 1.0 mg l^?1 NAA and 0.5 mg l^?1 BA with CP. Shoots produced an average of 15 roots per explant on ½ MS medium supplemented with 2.0 mg l^?1 IBA and BP. The 4 cm height plantlets with well-developed roots were successfully acclimatized. The results suggest that CP and BP can be used effectively to initiate shooting and rooting of Coelogyne cristata. Ploidy analysis of regenerated plants using flow cytometry revealed the same ploidy level (diploid). This efficient and reliable protocol could be useful for mass multiplication and germplasm conservation of the wild medicinal orchid.     

Knock Out of S1P3 Receptor Signaling Attenuates Inflammation and Fibrosis in Bleomycin-Induced Lung Injury Mice Model

Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid metabolite involved in many critical cellular processes, including proliferation, migration, and angiogenesis, through interaction with a family of five G protein–coupled receptors (S1P1–5). Some reports have implicated S1P as an important inflammatory mediator of the pathogenesis of airway inflammation, but the role of S1P3 in the pathogenesis of lung diseases is not completely understood. We used S1P3-deficient (knockout (KO)) mice to clarify the role of S1P3 receptor signaling in the pathogenesis of pulmonary inflammation and fibrosis using a bleomycin-induced model of lung injury. On the seventh day after bleomycin administration, S1P3 KO mice exhibited significantly less body weight loss and pulmonary inflammation than wild-type (WT) mice. On the 28th day, there was less pulmonary fibrosis in S1P3 KO mice than in WT mice. S1P3 KO mice demonstrated a 56% reduction in total cell count in bronchoalveolar lavage fluid (BALF) collected on the seventh day compared with WT mice; however, the differential white blood cell profiles were similar. BALF analysis on the seventh day showed that connective tissue growth factor (CTGF) levels were significantly decreased in S1P3 KO mice compared with WT mice, although no differences were observed in monocyte chemotactic protein-1 (MCP-1) or transforming growth factor β1 (TGF-β1) levels. Finally, S1P levels in BALF collected on the 7th day after treatment were not significantly different between WT and S1P3 KO mice. Our results indicate that S1P3 receptor signaling plays an important role in pulmonary inflammation and fibrosis and that this signaling occurs via CTGF expression. This suggests that this pathway might be a therapeutic target for pulmonary fibrosis.     

Akt-mediated phosphorylation of the G protein-coupled receptor EDG-1 is required for endothelial cell chemotaxis

The role of the protein kinase Akt in cell migration is incompletely understood. Here we show that sphingosine-1-phosphate (S1P)-induced endothelial cell migration requires the Akt-mediated phosphorylation of the G protein-coupled receptor (GPCR) EDG-1. Activated Akt binds to EDG-1 and phosphorylates the third intracellular loop at the T(236) residue. Transactivation of EDG-1 by Akt is not required for G(i)-dependent signaling but is indispensable for Rac activation, cortical actin assembly, and chemotaxis. Indeed, T236AEDG-1 mutant sequestered Akt and acted as a dominant-negative GPCR to inhibit S1P-induced Rac activation, chemotaxis, and angiogenesis. Transactivation of GPCRs by Akt may constitute a specificity switch to integrate rapid G protein-dependent signals into long-term cellular phenomena such as cell migration.     

Cyclooxygenase is an immediate-early gene induced by interleukin-1 in human endothelial cells.

The monokine interleukin-1 (IL-1) inhibits endothelial cell growth and induces prostacyclin production in human endothelial cells. Since cyclooxygenase (Cox) is the rate-limiting enzyme in the synthesis of prostanoids, we evaluated the ability of IL-1 to stimulate Cox expression by human umbilical vein endothelial cells (HUVEC) in vitro. Our data demonstrate that 1) the Cox mRNA is expressed at low levels in untreated cells; 2) IL-1 alpha induces the Cox mRNA within 2 h, and this induction is sustained for more than 24 h; 3) IL-1 alpha induction is dose-dependent; 4) cycloheximide potentiates the induction of the Cox mRNA by IL-1 alpha while actinomycin D prevents the induction, and 5) IL-1 alpha also stimulates Cox production in a time-dependent fashion which correlates with the increase in prostacyclin synthesis. These data suggest that Cox is an immediate-early gene induced by IL-1 in HUVEC and may contribute to the regulation of the endothelial cell differentiation pathway in vitro.     

Elevated Fis expression enhances recombinant protein production in Escherichia coli

A genetic strategy to enhance recombinant protein production is discussed. A small DNA bending protein, Fis, which has been shown to activate rRNA synthesis upon a nutrient upshift, was overexpressed in E. coli strain W3110 carrying vector pUCR1. Overexpression of Fis during exponential growth was shown to activate rrn promoters to different extents. A 5-fold improvement in chloramphenicol acetyltransferase (CAT) production in cultures with elevated Fis level was observed in shake-flask cultivations. A similar improvement in the culture performance was also observed during fed-batch fermentation; the specific CAT activity increased by more than 50% during the fed-batch phase for cultures with elevated Fis expression. In contrast, no increase in specific CAT activity was detected for cultures carrying pUCR2, expressing a frame-shift Fis mutant. Expression of Fis from a complementary vector, pKFIS, restored CAT production from W3110:pUCR2 to approximately the same level as cultures carrying pUCR1, indicating that the enhancement in CAT production was indeed Fis-dependent. The framework presented here suggests that differential activation in recombinant protein production may be achieved with differential Fis overexpression. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 56: 138-144, 1997.     

Prevalence of Intestinal Protozoans among Schoolchildren in Suburban Areas near Yangon,Myanmar

Although intestinal protozoans are common etiologies of diarrhea, few studies have been conducted in Myanmar. This study planned to investigate the prevalence of Giardia lamblia, Entamoeba coli, Entamoeba histolytica, and Endolimax nana among schoolchildren and their guardians in suburban areas near Yangon, Myanmar. We performed a cross-sectional survey among schoolchildren and their guardians from 7 primary schools in South Dagon and Hlaing Thar Yar districts, Yangon, Myanmar. Stool samples were observed with a microscope after concentration technique and iodine staining. Total 821 stool samples, including 556 from schoolchildren and 265 from guardians, were examined. The median age was 6 years old for schoolchildren and 36 years old for guardians. A 53.1% of the school children and 14.6 % of the guardians were males. The overall prevalence of each intestinal protozoan species was as follows: 3.4% (28/821) for G. lamblia; 3.5% (29/821) for E. coli; 1.2% (10/821) for E. histoytica, and 3.0% for E. nana. This study showed that intestinal protozoans are common in primary schoolchildren and their guardians in suburban areas near Yangon, Myanmar. Health interventions, such as hand washing education, improvement of sanitation, and establishment of water purification systems are urgently needed in this area.