Identification of an early endosomal protein regulated by phosphatidylinositol 3-kinase

Phosphatidylinositol 3-kinases (PI 3-kinases) have been implicated in membrane trafficking in the secretory and endocytic pathways of yeast and mammalian cells, but the molecular mechanisms by which these lipid kinases operate are not known. Here we identify a protein of 170 kDa that is rapidly released from cell membranes in response to wortmannin, a potent inhibitor of mammalian PI 3-kinases. The amino acid sequence of peptides from p170 reveal its identity to early endosomal antigen (EEA) 1, an endosomal antigen with homology to several yeast proteins genetically implicated in membrane trafficking. Immunofluorescence analysis of 3T3-L1 adipocytes with antisera against p170/EEA1 reveal a punctate peripheral pattern that becomes diffuse in response to wortmannin. In vitro, p170/EEA1 binds specifically to liposomes containing PIns(3)P, suggesting that the effect of wortmannin on cells is due to inhibition of PIns(3)P production. Thus, p170/EEA1 may define a family of proteins that mediate the regulatory effects of 3′-phosphoinositides on membrane trafficking in yeast and mammalian cells.     

Direct calibration of a reference standard against the air kerma strength primary standard, at 192Ir HDR energy

The primary standard of low air kerma rate sources or beams, maintained at the Radiological Standards Laboratory (RSL) of the Bhabha Atomic Research Centre (BARC), is a 60 cm3 spherical graphite ionization chamber. A 192Ir HDR source was standardized at the hospital site in units of air kerma strength (AKS) using this primary standard. A 400 cm3 bakelite chamber, functioning as a reference standard at the RSL for a long period, at low air kerma rates (compared to external beam dose rates), was calibrated against the primary standard. It was seen that the primary standard and the reference standard, both being of low Z, showed roughly the same scatter response and yielded the same calibration factor for the 400 cm3 reference chamber, with or without room scatter. However, any likelihood of change in the reference chamber calibration factor would necessitate the re-transport of the primary standard to the hospital site for re-calibration. Frequent transport of the primary standard can affect the long-term stability of the primary standard, due to its movement or other extraneous causes. The calibration of the reference standard against the primary standard at the RSL, for an industrial type 192Ir source maintained at the laboratory, showed excellent agreement with the hospital calibration, making it possible to check the reference chamber calibration at RSL itself. Further calibration procedures have been developed to offer traceable calibration of the hospital well ionization chambers.     

In vitro metabolism of midazolam, triazolam, nifedipine, and testosterone by human liver microsomes and recombinant cytochromes p450: role of cyp3a4 and cyp3a5.

Midazolam, triazolam (TRZ), testosterone, and nifedipine have all been widely used as probes for in vitro metabolism of CYP3A. We used these four substrates to assess the contributions of CYP3A4 and CYP3A5 to in vitro biotransformation in human liver microsomes (HLMs) and in recombinant enzymes. Recombinant CYP3A4 and CYP3A5 (rCYP3A4 and rCYP3A5) both produced 1-OH and 4-OH metabolites from midazolam and triazolam, 6 beta-hydroxytestosterone from testosterone, and oxidized nifedipine from nifedipine. Overall, the metabolic activity of CYP3A5 was less than that of CYP3A4. Ketoconazole potently inhibited midazolam, triazolam, testosterone, and nifedipine metabolite formation in HLMs and in rCYP3A4. The inhibitory potency of ketoconazole in rCYP3A5 was about 5- to 19-fold less than rCYP3A4 for all four substrates. In testosterone interaction studies, testosterone inhibited 1-OH-TRZ formation, but significantly activated 4-OH-TRZ formation in HLMs and rCYP3A4 but not in rCYP3A5. Oxidized nifedipine formation was inhibited by testosterone in rCYP3A4. However, in rCYP3A5, testosterone slightly activated oxidized nifedipine formation at lower concentrations, followed by inhibition. Thus, CYP3A4 and CYP3A5 both contribute to midazolam, triazolam, testosterone, and nifedipine biotransformation in HLMs, with CYP3A5 being metabolically less active than CYP3A4 in general. Because the inhibitory potency of ketoconazole in rCYP3A5 is substantially less than in rCYP3A4 and HLMs, CYP3A5 is probably less important than CYP3A4 in drug-drug interactions involving ketoconazole and CYP3A substrates.     

Cyclodextrin complexes of valdecoxib: properties and anti-inflammatory activity in rat.

The influence of natural beta-cyclodextrin and its hydrophilic derivatives (HPbetaCd and SBE7betaCd) on the in vitro dissolution rate, in vivo absorption and oral bioavailability of a poorly water soluble anti-inflammatory agent, valdecoxib (VALD) was studied. Equimolar drug-cyclodextrin solid complexes were prepared by kneading and coevaporation methods and characterized by infrared spectroscopy, differential scanning calorimetry, X-ray diffraction. In the liquid state, the cyclodextrin complexes were studied using phase solubility analysis, (1)H nuclear magnetic resonance and circular dichroism spectroscopy. Drug solubility and dissolution rate in distilled water were notably improved by employing the betaCds. The DP(15) (i.e. percent of dissolved VALD at 15 min) was 10.5% for the pure drug and 50, 91 and 93% for VALD-betaCd, VALD-HPbetaCd and VALD-SBE7betaCd complexes, respectively. Moreover, it was found that in the, the cyclodextrin complexes of drug showed significant improvement in the anti-inflammatory activity.     

Poly-Ingredient Formulation Bresol? Ameliorates Experimental Chronic Obstructive Pulmonary Disease (COPD) in Rats

In the present study, the protective effect of Bresol^® – a polyherbal formulation – was evaluated in an experimental model of cigarette smoke (CS)-induced COPD in rats. Ten minutes daily exposure to CS for 7 weeks caused significant elevation of TNF-α (p<0.01) and total protein (p<0.01) in the bronchoalveolar lavage fluid (BALF) of positive untreated control animals, indicating ongoing inflammatory process in the lungs. Further, histopathological findings have confirmed the presence of pathological lesions in the trachea and lungs. Five weeks of post-treatment with Bresol^® (250 and 500 mg/kg, p.o.) showed significant and dose-dependent anti-inflammatory effects against CS-induced lung abnormalities by maintaining the TNF-α and total protein levels within the normal range. Additionally, Bresol^®-treated animals showed normal cyto-architecture of the trachea and lungs. In conclusion, Bresol^® showed dose-dependent protection against CS-induced lung and tracheal injury in rats, which further indicates, Bresol^® is a useful healing agent, may help to decelerate the progression of COPD, and reduce the exacerbations in patients.     

Efficacy and safety of beating heart coronary revascularization coupled with ascending aorta-bifemoral grafting: Analysis of short term results

Introduction  

Coronary Artery Disease (CAD) frequently coexists with Peripheral Vascular Disease (PVD) and poses management issues. When there is a concomitant infrarenal aortic occlusive disease, abdominal aorta is the traditional donor of bypass inflow to the lower limbs. However, the ascending aorta may also be used as the source of inflow to both the femoral and coronary arteries in patients who present with combined CAD and PVD. Here, 5 year follow up results of simultaneous off-pump coronary artery bypass grafting (OPCAB) and Ascending Aorto-Bifemoral Grafting [AABG] are presented and merits of the procedure are discussed.     

Necrotising fasciitis of the neck

Necrotising fasciitis is a fulminant soft tissue infection that causes necrosis of fascia and subcutaneous tissue while sparing skin and muscle initially. It is most commonly seen in adults involving the perineum, extremities and and minal wall. Immunncompromised patients are at an increased risk of developing this infection. These infections require early diagnosis, aggressive surgical debridaient and appropriate antibiotic therapy. Mortality rates have been reported to be as high as 52 and 73% in general surgery literature.(Freuschtag et al, 1985., Rouse et at 1982). Necrotising fasciitis of the head and neck is rare. The commonest cause is secondary to denial infections. We report a case of necrotising fasciitis of the neck secondary to a peritonsillar abscess in a previously healthy individual. The pathogenesis and treatment of this fulminant infection are also discussed.     

Encapsulation of water-insoluble drug by a cross-linking technique: Effect of process and formulation variables on encapsulation efficiency, particle size, and in vitro dissolution rate

Ibuprofen-gelatin micropellets were prepared by the cross-linking technique using formaldehyde. Spherical micropellets having an entrapment efficiency of 65% to 85% were obtained. The effect of core to coat ratio, speed of agitation, temperature, and volume of oil phase was studied with respect to entrapment efficiency, micropellet size, and surface characteristics. Fourier transform infrared spectroscopy and differential scanning calorimetric analysis confirmed the absence of any drug-polymer interaction. X-ray diffraction patterns showed that there is a decrease in crystallinity of the drug. The micromeritic properties of micropellets were found to be slightly changed by changing various processing parameters to give micropellets of good flow property. The in vitro release profile could be altered significantly by changing various processing parameters to give a controlled release of drug from the micropellets. The stability studies of the drug-loaded micropellets showed that the drug was stable at storage conditions of room temperature, 37 degrees C, 25 degrees /60% relative humidity (RH) and 45 degrees /60% RH, for 12 weeks.