Interaction of methotrexate with polycationic carriers

Binding parameters were measured for the $\text{in vitro}$ interaction of methotrexate with poly-L-lysine and diethylaminoethyl dextran. The complexes were found to have apparent affinity constants of 2590 and 440 M^?1 respectively at 37°, ionic strength 0.02. For hypotonic solutions containing therapeutic concentrations of methotrexate, this results in 87% binding of the drug to poly-L-lysine and 74% to diethylaminoethyl dextran. While the binding decreased to about 50% upon increasing ionic strength to physiologic levels, sufficient drug-carrying capacity was retained at isotonicity to support the potential utility of such complexes as tissue-specific drug carriers.     

Development of cardiac form and function in ectothermic sauropsids

Evolutionary morphologists and physiologists have long recognized the phylogenetic significance of the ectothermic sauropsids. Sauropids have been classically considered to bridge between early tetrapods, ectotherms, and the evolution of endotherms. This transition has been associated with many modifications in cardiovascular form and function, which have changed dramatically during the course of vertebrate evolution. Most cardiovascular studies have focused upon adults, leaving the development of this critical system largely unexplored. In this essay, we attempt a synthesis of sauropsid cardiovascular development based on the limited literature and indicate fertile regions for future studies. Early morphological cardiovascular development, i.e., the basic formation of the tube heart and the major pulmonary and systemic vessels, is similar across tetrapods. Subsequent cardiac chamber development, however, varies considerably between developing chelonians, squamates, crocodilians, and birds, reflected in the diversity of adult ventricular structure across these taxa. The details of how these differences in morphology develop, including the molecular regulation of cardiac and vascular growth and differentiation, are still poorly understood. In terms of the functional maturation of the cardiovascular system, reflected in physiological mechanisms for regulating heart rate and cardiac output, recent work has illustrated that changes during ontogeny in parameters such as heart rate and arterial blood pressure are somewhat species‐dependent. However, there are commonalities, such as a β‐adrenergic receptor tone on the embryonic heart appearing prior to 60% of development. Differential gross morphological responses to environmental stressors (oxygen, hydration, temperature) have been investigated interspecifically, revealing that cardiac development is relatively plastic, especially, with respect to change in heart growth. Collectively, the data assembled here reflects the current limited morphological and physiological understanding of cardiovascular development in sauropsids and identifies key areas for future studies of this diverse vertebrate lineage. J. Morphol., 2009. © 2009 Wiley‐Liss, Inc.     

Expression and characterization of delayed rectifying K+ channels in anterior rat taste buds

Delayed rectifying K⁺ (DRK) channels in taste cells have been implicated in the regulation of cell excitability and as potential targets for direct and indirect modulation by taste stimuli. In the present study, we have used patch-clamp recording to determine the biophysical properties and pharmacological sensitivity of DRK channels in isolated rat fungiform taste buds. Molecular biological assays at the taste bud and single-cell levels are consistent with the interpretation that taste cells express a variety of DRK channels, including members from each of the three major subfamilies: KCNA, KCNB, and KCNC. Real-time PCR assays were used to quantify expression of the nine DRK channel subtypes. While taste cells express a number of DRK channels, the electrophysiological and molecular biological assays indicate that the Shaker Kv1.5 channel (KCNA5) is the major functional DRK channel expressed in the anterior rat tongue. transduction     

Degradation of mutated bovine pancreatic trypsin inhibitor in the yeast vacuole suggests post-endoplasmic reticulum protein quality control

The rate-limiting step in protein secretion is folding, which occurs in the endoplasmic reticulum (ER) lumen, and almost all secreted proteins contain disulfide bonds that form in the ER and stabilize the native state. Secreted proteins unable to fold may aggregate or they may be subject to ER-associated protein degradation. To examine the fate of aberrant forms of a well characterized, disulfide-bonded secreted protein, we expressed bovine pancreatic trypsin inhibitor in yeast. Bovine pancreatic trypsin inhibitor is a single domain, 58-amino acid polypeptide containing three disulfide bonds, and yeast cells secrete the wild type protein. In contrast, the Y35L mutant, which folds rapidly but is unstable, remains soluble and is not secreted. Surprisingly, the proteolysis of Y35L is unaffected in yeast containing mutations in genes encoding factors required for ER-associated protein degradation and is stable if artificially retained in the ER. Rather, Y35L is diverted from the Golgi to the vacuole and degraded. Because only the mutant protein is quantitatively proteolyzed these data suggest that a post-ER quality control check-point diverts unstable proteins to the vacuole for degradation.     

Chargeability measurements of selected pharmaceutical dry powders to assess their electrostatic charge control capabilities

The purpose of this study was to develop an instrument (the Purdue instrument) and the corresponding methodologies to measure the electrostatic charge development (chargeability) of dry powders when they are in dynamic contact with stainless steel surfaces. The system used an inductive noncontact sensor located inside an aluminum Faraday cage and was optimized to measure the charging capabilities of a fixed volume of powder (0.5 cc). The chargeability of 5,5-diphenyl-hydantoin, calcium sulfate dihydrate, cimetidine, 3 grades of colloidal silicon dioxide, magnesium stearate, 4 grades of microcrystalline cellulose, salicylic acid, sodium carbonate, sodium salicylate, spray-dried lactose, and sulfin-pyrazone were tested at 4 linear velocities, and the particle size distribution effect was assessed for 3 different grades of colloidal silicon dioxide and 4 different grades of micro-crystalline cellulose. The chargeability values exhibited a linear relationship for the range of velocities studied, with colloidal silicon dioxide exhibiting the maximum negative chargeability and with spray-dried lactose being the only compound to exhibit positive chargeability. The instrument sensitivity was improved by a factor of 2 over the first generation version, and the electrostatic charge measurements were reproducible with relative standard deviations ranging from nondetectable to 33.7% (minimum of 3 replicates). These results demonstrate the feasibility of using the Purdue instrument to measure the electrostatic charge control capabilities of pharmaceutical dry powders with a reasonable level of precision.