Hairy root culture of <Emphasis Type="Italic">Plumbago indica</Emphasis> as a potential source for plumbagin

Hairy roots of Plumbago indica were established at high frequency (90 %) by infecting leaf explants with Agrobacterium rhizogenes strain ATCC 15834. The axenic root cultures were established under darkness in hormone-free liquid Murashige and Skoog medium containing 3 % sucrose. The highest plumbagin content was found to accumulate in roots at their exponential phase of growth. A low pH (4.6) and a low concentration of sucrose (1 %) were beneficial for root growth in darkness, while pH 5.6 and 3 % sucrose under continuous irradiance enhanced plumbagin accumulation in roots up to 7.8 mg g⁻¹(d.m.). Direct shoot regeneration from hairy root culture was also achieved under continuous irradiance, thus indicated an easy way of obtaining transformed P. indica plants.     

An ergonomic study on teenage girls working in the manual brick manufacturing units in the unorganized sectors in West Bengal, India

There are laws which protect and prevent the teenage girls from working. But still in the developing countries like India, Bangladesh, etc. teenage girls coming from low socio-economic background work, to add to the family income. There is dearth of data regarding the ergonomic studies of the teenage girls working in the manual brick manufacturing units of the unorganized sectors in India. Therefore, a field study was conducted in about four brick manufacturing units of West Bengal, India. Their job in these fields was to carry 8-10 unbaked and sun-dried bricks (49.11 +/- 2.13 Kg) from the brick field to the kiln-top and again carry the baked bricks back from the kiln to some other corner of the brick field, under the hot sun. Thus, this study aimed at evaluating their work load and their consequent work-related health effects. Results show that 86 % of them suffered from work-related pain in different body parts and there was an increase in their physiological and psychological parameters after work. It is concluded that the frequent use of awkward postures while at work (OWAS) and performing heavy to moderate types of tasks in the field are deteriorating their health conditions.     

Regeneration of transformed plants from hairy roots of <Emphasis Type="Italic">Plumbago indica</Emphasis>

Spontaneous shoot regeneration was observed from Agrobacterium rhizogenes-induced hairy roots of Plumbago indica when these were incubated in liquid MS medium for a period of 3 weeks under continuous light. Insertion of the rolB gene in putative transformed plants was confirmed by PCR and sequencing. Transformed plants grown for a period of 1 week on solid MS medium containing 0.5 mg l⁻¹ 6-benzyladenine and then transplanted to growth regulator-free medium showed better overall growth than control plants. Transformed plants had a higher root bio-biomass and an increased plumbagin content relative to non-transformed plants.     

In vitro splicing of erythropoietin by the Mycobacterium tuberculosis RecA intein without substituting amino acids at the splice junctions

Protein splicing is a self-catalyzed process involving the excision of an intervening polypeptide sequence, the intein, and joining of the flanking polypeptide sequences, the extein, by a peptide bond. We have studied the in vitro splicing of erythropoietin (EPO) using a truncated form of the Mycobacterium tuberculosis RecA mini-intein in which the homing endonuclease domain was replaced with a hexahistidine sequence (His-tag). The intein was inserted adjacent to cysteine residues to assure that the spliced product had the natural amino acid sequence. When expressed in Escherichia coli, intein-containing EPO was found entirely as inclusion bodies but could be refolded in soluble form in the presence of 0.5 M arginine. Protein splicing of the refolded protein could be induced with a reducing agent such as DTT or tris(2-carboxyethyl)phosphine and led to the formation of EPO and mini-intein along with some cleavage products. Protein splicing mediated by the RecA intein requires the presence of a cysteine residue adjacent to the intein insertion site. We compared the efficiencies of protein splicing adjacent to three of the four cysteine residues of EPO (Cys29, Cys33 and Cys161) and found that insertion of intein adjacent to Cys29 allowed far more efficient protein splicing than insertion adjacent to Cys33 or Cys161. For ease of purification, our experiments involved a His-tagged EPO fusion protein and a His-tagged intein and the spliced products (25 kDa EPO and 24 kDa mini-intein) were identified by Western blotting using anti-EPO and anti-His-tag antibodies and by mass spectroscopy. The optimal splicing yield at Cys29 (40%) occurred at pH 7.0 after refolding at 4 degrees C and splicing for 18 h at 25 degrees C in the presence of 1 mM DTT.     

Fluorescence probe study of Ca2+-dependent interactions of calmodulin with calmodulin-binding peptides of the ryanodine receptor

We have used a highly environment-sensitive fluorescent probe 6-bromoacetyl-2-dimethylaminonaphthalene (badan) to study the interaction between calmodulin (CaM) and a CaM-binding peptide of the ryanodine receptor (CaMBP) and its sub-fragments F1 and F4. Badan was attached to the Thr34Cys mutant of CaM (CaM-badan). Ca(2+) increase in a physiological range of Ca(2+) (0.1-2 microM) produced about 40 times increase in the badan fluorescence. Upon binding to CaMBP, the badan fluorescence of apo-CaM showed a small increase at a slow rate; whereas that of Ca-CaM showed a large decrease at a very fast rate. Upon binding of CaM to the badan-labeled CaMBP, the badan fluorescence showed a small and slow increase at low Ca(2+), and a large and fast increase at high Ca(2+). Thus, the badan probe attached to CaM Cys(34) can be used to monitor conformational changes occurring not only in CaM, but also those in the CaM-CaMBP interface. Based on our results we propose that both the interaction interface and the global conformation of the CaM-CaMBP complex are altered by calcium.     

Swertia (Gentianaceae): chemical and pharmacological aspects

A compilation of the constituents isolated from Swertia species covering the literature up to December 2003 is presented. The botanical classification and ethno-pharmacology of Swertia plants, as well as the biological activities and pharmacological applications of both distinct phytochemicals and medicinally active plant materials (formulations, extracts, etc.) are discussed in detail.     

Norepinephrine induces alveolar epithelial apoptosis mediated by alpha-, beta-, and angiotensin receptor activation

Norepinephrine (NE) induces apoptosis in cardiac myocytes, and autocrine production of angiotensin (ANG) II is required for apoptosis of alveolar epithelial cells (AECs) (Wang R, Zagariya A, Ang E, Ibarra-Sunga O, and Uhal BD. Am J Physiol Lung Cell Mol Physiol 277: L1245--L1250, 1999; Wang R, Alam G, Zagariya A, Gidea C, Pinillos H, Lalude O, Choudhary G, and Uhal BD. J Cell Physiol 185: 253--259, 2000). On this basis, we hypothesized that NE might induce apoptosis of AECs in a manner inhibitable by ANG system antagonists. Purified NE induced apoptosis in the human A549 AEC-derived cell line or in primary cultures of rat AECs, with EC(50) values of 200 and 20 nM, respectively. Neither the alpha-agonist phenylephrine nor the beta-agonist isoproterenol could mimic NE when tested alone but when applied together could induce apoptosis with potency equal to NE. Apoptosis and net cell loss (47--59% in 40 h) in response to NE was completely abrogated by the ANG-converting enzyme inhibitor lisinopril or the ANG II receptor antagonist saralasin, each at concentrations capable of blocking Fas- or tumor necrosis factor-alpha-induced apoptosis. These data suggest that NE induces apoptosis of human and rat AECs through a mechanism involving the combination of alpha- and beta-adrenoceptor activation followed by autocrine generation of ANG II.     

Early expression of WUSCHEL is a marker for in vitro shoot morphogenesis in tobacco and <Emphasis Type="Italic">Beta palonga</Emphasis>

We have studied the role of growth regulators behind in vitro shoot organogenesis and somatic embryogenesis in two plant systems, viz. tobacco (Nicotiana tabacum L. var. Jayasri) and Beta palonga R.K. Basu & K.K. Mukh. We have also correlated the phenomena of de differentiation with the relative expression of WUS (WUSCHEL) gene in a time-dependent manner. The results indicated that early WUS gene expression is a definite marker for in vitro shoot organogenesis in tobacco and Beta both in direct and indirect modes of regeneration. Additionally, we have performed a comparative homology modeling and in silico structural analysis of WUSCHEL proteins of B. palonga, B. vulgaris, and Arabidopsis to find out the commonality of the ligand binding site. The amino acids of the binding sites were identical (Arginine, Tryptophan, Proline, Asparagine, and Tyrosine) in the three materials under study; except two additional amino acids (Isoleucine and Alanine) in B. vulgaris.     

Predictors of chemoradiation related febrile neutropenia prophylaxis in older adults – Experience from a limited resource setting

Aim

To identify risk factors that lower efficacy of antibiotic prophylaxis of febrile neutropenia among older patients on chemoradiation.