Attachment staining: a novel method for flow cytometric quantitation of protein expression in limited numbers of adherent cells

Malignant tumors are known to exhibit high rates of glycolytic activity leading to high production of lactic acid. Hence, neoplastic cells have elevated activity of enzymes responsible for glycolysis. Echitamine chloride, an indole alkaloid extracted from the bark of Alstonia scholaris, has been reported to have a highly promising anticancer activity against fibrosarcoma in rats. In the present study, the effect of echitamine chloride on energy metabolism of S-180 cells is investigated to have a better understanding on the mode of action of echitamine chloride. The effect of echitamine chloride on the mitochondrial and cellular respiration of S-180 cells was studied. Also, the effects on glucose utilization, pyruvate utilization and lactate formation were studied on whole S-180 cells and S-180 cell-free homogenate. The levels of glycolytic enzymes such as hexokinase and lactate dehydrogenase were estimated in which particular emphasis has been laid on hexokinase which occurs both in cytosolic and particulate forms in neoplastic cells. Hence the differential effect of echitamine chloride on the levels of total, cytosolic and particulate hexokinase has been investigated. In conclusion, echitamine chloride affects both cellular and mitochondrial respiration, leading to reduction of the cellular energy pool and thereby resulting in the loss of viability of S-180 cells.     

Blood and tissue compatibility of modified polyester: thrombosis, inflammation, and healing

OBJECTIVE: The effects of BRL-32872, azimilide and a selective blocker of the delayed rectifier potassium current, E-4031, were measured at two different basic cycle lengths (BCL), 300 and 1000 ms. Calcium channel antagonists of sarcolemmal (verapamil and nitrendipine) and sarcoplasmic reticulum (ryanodine) membranes were used to investigate whether the inhibition of the calcium current or the calcium release from the sarcoplasmic reticulum could alter the reverse-rate dependence of E-4031 on action potential duration (APD). METHODS: Guinea pig isolated papillary muscles were superfused with a Tyrode solution maintained at 37 degrees C and stimulated at a BCL of 300 or 1000 ms. The standard microelectrode technique was used to record action potential parameters and to study the effects of azimilide, BRL-32872 and E-4031. E-4031 was superfused at increasing concentrations (0.01, 0.03, 0.1 and 0.3 microM) in the absence or in the presence of verapamil (0.3 microM), nitrendipine (0.03 microM) or ryanodine (0.1 microM). RESULTS: BRL-32872 and azimilide induced a self-limited concentration-dependent increase in APD. The effect of BRL-32872 was not dependent on the stimulation frequency whereas the effect of azimilide was significantly reduced at the shorter BCL. E-4031 induced a concentration-dependent increase in APD at both stimulation BCL. The increase in APD was significantly more pronounced in fibres stimulated at a BCL of 1000 ms than in fibres stimulated at a BCL of 300 ms, characterising the reverse-frequency dependent effect of class III antiarrhythmic agents. The reverse-frequency dependence in action potential prolongation induced by E-4031 was significantly reduced in the presence of a low concentration of verapamil (0.3 microM), nitrendipine (0.03 microM), or ryanodine (0.1 microM. CONCLUSION: The results show that BRL-32872, in contrast to azimilide, does not induce the reverse-rate dependency of action potential prolongation typically produced by class III antiarrhythmic agents such as E-4031. Our results also show that reverse-rate dependency induced by E-4031 can be reduced by the simultaneous administration of a low concentration of a calcium channel antagonist or an inhibitor of the release of calcium from the sarcoplasmic reticulum. It is thus suggested that compounds with a suitable balance of potassium and calcium antagonistic activities may have less adverse effects than purely selective potassium channel blockers.     

Occupational skin diseases in Washington State, 1989 through 1993: using workers' compensation data to identify cutaneous hazards

OBJECTIVES: This study sought to characterize occupational dermatoses and cutaneous hazards. METHODS: Workers' compensation claims filed for skin disease in the Washington State Fund were analyzed for 1989 through 1993; incidence rates for industries and employers were calculated, and cutaneous hazards associated with the highest rates were identified. RESULTS: A total of 7445 claims were filed for skin disorders, principally contact dermatitis; 675 (9.1%) involved more than 3 missed work-days. The rate of accepted skin disorder claims was 1.0 per 1000 full-time employee-years. The highest incidence rates (4.6 to 30.7 accepted claims per 1000 full-time employee-years) were in certain manufacturing industries (plastics related, concrete products, aircraft parts, sporting goods, and boat building), wholesale farm product raw materials, automotive glass replacement, and beauty shops. Seven of the 10 employers with the highest incidence rates (19.6 to 85.5 accepted claims per 1000 full-time employee-years) used fiber-reinforced plastics (composites) and exposed workers to epoxy and other resin systems associated with contact dermatitis. CONCLUSIONS: Workers' compensation data identify known and emerging workplace cutaneous hazards and show promise for targeting prevention efforts.     

Non-peptidic HIV protease inhibitors: C2-symmetry-based design of bis-sulfonamide dihydropyrones

Potent, non-peptidic, dihydropyrone sulfonamide HIV protease inhibitors have been previously described. Crystallographic analysis of dihydropyrone sulfonamide inhibitor/HIV protease complexes suggested incorporation of a second, C2 symmetry-related sulfonamide group. Selected bis-sulfonamide dihydropyrone analogues display high HIV protease inhibitory activity.     

Common pediatric esophageal disorders

Esophageal disorders in children can result in significant morbidity. The most common esophageal disorder seen in children is gastroesophageal reflux. Other common disorders affecting the esophagus include peptic esophageal strictures, esophageal atresia with or without tracheoesophageal fistula, caustic and foreign body ingestions, achalasia, and cricopharyngeal achalasia. We discuss what is currently known about these common pediatric esophageal disorders with regard to pathophysiology, clinical presentation, and diagnostic and treatment strategies.     

Expression of vimentin and cytokeratin in eutopic and ectopic endometrium of women with adenomyosis and ovarian endometrioma

Tuberculosis is a continuous threat for health in all parts of the world. In industrialized centers an emerging increase of tuberculosis is observed due to low health standards, poor health education and control, increasing migration and the HIV epidemic. This should alert the pediatrician, since most tuberculosis infections in childhood start with contacts of infected adults with children. There is still a good response to most drugs against tuberculosis, but in some parts of the world the resistance to conventional treatment is an emerging problem. In general incidence, morbidity and mortality of tbc in children of the western world is low and has good prognosis. However, the diagnostic principles and the education of pediatricians for recognizing tbc is still of utmost importance.     

Chemokine receptor CCR2 expression and monocyte chemoattractant protein-1-mediated chemotaxis in human monocytes. A regulatory role for plasma LDL

The subendothelial accumulation of macrophage-derived foam cells is one of the hallmarks of atherosclerosis. The recruitment of monocytes to the intima requires the interaction of locally produced chemokines with specific cell surface receptors, including the receptor (CCR2) for monocyte chemoattractant protein-1 (MCP-1). We have previously reported that monocyte CCR2 gene expression and function are effectively downregulated by proinflammatory cytokines. In this study we identified low density lipoprotein (LDL) as a positive regulator of CCR2 expression. Monocyte CCR2 expression was dramatically increased in hypercholesterolemic patients compared with normocholesterolemic controls. Similarly, incubation of human THP-1 monocytes with LDL induced a rapid increase in CCR2 mRNA and protein. By 24 hours the number of cell surface receptors was doubled, causing a 3-fold increase in the chemotactic response to MCP-1. The increase in CCR2 expression and chemotaxis was promoted by native LDL but not by oxidized LDL. Oxidized LDL rapidly downregulated CCR2 expression, whereas reductively methylated LDL, which does not bind to the LDL receptor, had only modest effects on CCR2 expression. A neutralizing anti-LDL receptor antibody prevented the effect of LDL, suggesting that binding and internalization of LDL were essential for CCR2 upregulation. The induction of CCR2 expression appeared to be mediated by LDL-derived cholesterol, because cells treated with free cholesterol also showed increased CCR2 expression. These data suggest that elevated plasma LDL levels in conditions such as hypercholesterolemia enhance monocyte CCR2 expression and chemotactic response and potentially contribute to increased monocyte recruitment to the vessel wall in chronic inflammation and atherogenesis.     

Increased MPTP neurotoxicity in vesicular monoamine transporter 2 heterozygote knockout mice

The neurotoxic action of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) has been proposed to be attenuated by sequestration into intracellular vesicles by the vesicular monoamine transporter (VMAT2). The purpose of this study was to determine if mice with genetically reduced levels of VMAT2 (heterozygote knockout; VMAT2 +/-) were more vulnerable to MPTP. Striatal dopamine (DA) content, the levels of DA transporter (DAT) protein, and the expression of glial fibrillary acidic protein (GFAP) mRNA, a marker of gliosis, were assessed as markers of MPTP neurotoxicity. In all parameters measured VMAT2 +/- mice were more sensitive than their wild-type littermates (VMAT2 +/+). Administration of MPTP (7.5, 15, or 30 mg/kg, b.i.d.) resulted in dose-dependent reductions in striatal DA levels in both VMAT2 +/- and VMAT2 +/+ animals, but the neurotoxic potency of MPTP was approximately doubled in the VMAT2 +/- mice: 59 versus 23% DA loss 7 days after 7.5 mg/kg dose for VMAT2 +/- and VMAT2 +/+ mice, respectively. Dopaminergic nerve terminal integrity, as assessed by DAT protein expression, also revealed more drastic reductions in the VMAT2 +/- mice: 59 versus 35% loss at 7.5 mg/kg and 95 versus 58% loss at 15 mg/kg for VMAT2 +/- and VMAT2 +/+ mice, respectively. Expression of GFAP mRNA 2 days after MPTP was higher in the VMAT2 +/- mice than in the wild-type: 15.8- versus 7.8-fold increase at 7.5 mg/kg and 20.1- versus 9.6-fold at 15 mg/kg for VMAT2 +/- and VMAT2 +/+ mice, respectively. These observations clearly demonstrate that VMAT2 +/- mice are more susceptible to the neurotoxic effects of MPTP, suggesting that VMAT2-mediated sequestration of the neurotoxin into vesicles may play an important role in attenuating MPTP toxicity in vivo.     

Ataxic hemiparesis: critical appraisal of a lacunar syndrome

BACKGROUND and PURPOSE: Ataxic hemiparesis is a well-recognized lacunar syndrome involving homolateral ataxia with accompanying corticospinal tract impairment. Despite 30 years of clinical experience there continues to be some doubt as to the defining clinical characteristics, precise neuroanatomic localization of the syndrome, and etiologic mechanisms. METHODS: We now present 45 new cases that have been analyzed for clinico-radiologic correlation and etiology. Also, all published cases from the English literature known to the authors are reviewed. RESULTS: We found that the clinical syndrome of ataxic hemiparesis accurately predicts a small deep infarction, generally in the pons or internal capsule. Sensory loss is highly associated with a capsular localization. We found that 47% of the cases were attributed to small-vessel disease, 11% to cardioembolism, and only 7% to artery-to-artery embolism (all in the basilar artery); 1 case was attributed to thrombocytosis, 1 to multiple sclerosis, and the rest either had negative or incomplete evaluation. Approximately two thirds of the infarctions occurred in patients with neuroimaging evidence of other ischemic brain lesions. CONCLUSIONS: Ataxic hemiparesis is a distinct clinical syndrome that accurately predicts a small deep infarction, most commonly in the pons or internal capsule. Only sensory loss accurately predicts a capsular localization. Etiology in nearly half of the cases can be attributed to small-vessel disease. Furthermore, ataxic hemiparesis appears to be a good marker for generalized asymptomatic cerebrovascular disease.     

Importance of ribosomal frameshifting for human immunodeficiency virus type 1 particle assembly and replication

The recent development and use of protease inhibitors have demonstrated the essential role that combination therapy will play in the treatment of individuals infected with the human immunodeficiency virus type 1 (HIV-1). Past clinical experience suggests that due to the appearance of resistant HIV-1 variants, additional therapeutics will be required in the future. To identify new options for combination therapy, it is of paramount importance to pursue novel targets for drug development. Ribosomal frameshifting is one potential target that has not been fully explored. Data presented here demonstrate that small molecules can stimulate frameshifting, leading to an imbalance in the ratio of Gag to Gag-Pol and inhibiting HIV-1 replication at what appears to be the point of viral particle assembly. Thus, we propose that frameshifting represents a new target for the identification of novel anti-HIV-1 therapeutics.