Stereochemical influence on the stability of radio-metal complexes in vivo. Synthesis and evaluation of the four stereoisomers of 2-(p-nitrobenzyl)-trans-CyDTPA

Distinct differences in in vivo stability of the two diastereomeric C-Functionalized CyDTPA chelating agents, (CHX-A DTPA and CHX-B DTPA, both racemates), as recently reported prompted further investigation as to why differences in configuration produced striking effects on the in vivo stability of their yttrium complexes. To this end, the four individual component stereoisomers of CHX-A and CHX-B were synthesized and ability to bind yttrium was investigated both in vitro and in vivo.     

Effects of myosin light chain kinase inhibitors on carbachol-activated nonselective cationic current in guinea-pig gastric myocytes

The effects of myosin light chain kinase inhibitors on muscarinic stimulation-activated nonselective cationic current (ICCh) in guinea-pig gastric antral myocytes were studied using the whole-cell patch-clamp technique. ICCh was induced by carbachol (CCh, 50 microM) at a holding potential of -30 mV or -60 mV. ML-7, a chemical inhibitor of myosin light chain kinase (MLCK), inhibited ICCh concentration dependently in a reversible manner (53 +/- 8.6% at 1 microM, mean +/- SE, n = 11). In addition, amplitudes of ICCh were only 37 +/- 2.7% of the daily control values following the addition of a peptide inhibitor of MLCK to the pipette solution. On the other hand, ML-7 had an inhibitory effect on voltage-operated Ca2+ channel current. The peak value of Ba2+ current at 0 mV was reduced to 35 +/- 7.4% (n = 9) by 3 microM of ML-7. As ICCh is known to have an intracellular Ca2+ dependence, we tried to exclude the possibility that ML-7 inhibited ICCh indirectly via suppression of Ca2+ current and the similar inhibitory effects of ML-7 on ICCh were confirmed under the following conditions: (1) clamp of membrane potential at -60 mV; (2) clamp of intracellular [Ca2+] to 1 microM by 10 mM BAPTA; (3) pre-inhibition of Ca2+ channel by verapamil. Different from the effects on ICCh, ML-7 barely inhibited the same cationic current induced by guanosine 5'-O-(3-thiotriphosphate) (GTP[gammaS], 0.2 mM) in the pipette solution. These results suggest that a Ca2+/calmodulin-MLCK-dependent pathway can modulate the activation of ICCh in guinea-pig gastric antral myocytes.     

Comparison of calcium lactate with chlorine as a washing treatment for fresh‐cut lettuce and carrots: quality and nutritional parameters

The efficacies of calcium lactate and chlorine washing treatments of fresh‐cut lettuce and carrots were compared during storage at 4 °C over 10 days. The gas composition of packages, colour, enzyme activity, texture, sensory attributes, microflora and levels of ascorbic acid and carotenoids were evaluated at 1, 3, 7 and 10 days. Calcium lactate treatment was not significantly different to chlorine treatment (p < 0.05) in terms of maintaining colour, texture and acceptability of fresh‐cut lettuce and carrots during the entire storage period. The washing treatments did not affect levels of ascorbic acid of fresh‐cut lettuce or carrots. Carotenoid levels were higher in calcium lactate‐treated carrots than chlorine‐treated samples at the end of storage. Mesophilic, psychrotrophic and lactic acid bacteria counts were not significantly different between treatments for both vegetables. Copyright © 2005 Society of Chemical Industry     

Endothelial cell cytosolic free calcium regulates neutrophil migration across monolayers of endothelial cells

Polymorphonuclear leukocytes (PMN) traverse an endothelial cell (EC) barrier by crawling between neighboring EC. Whether EC regulate the integrity of their intercellular adhesive and junctional contacts in response to chemotaxing PMN is unresolved. EC respond to the binding of soluble mediators such as histamine by increasing their cytosolic free calcium concentration ([Ca++]i) (Rotrosen, D., and J.I. Gallin. 1986. J. Cell Biol. 103:2379-2387) and undergoing shape changes (Majno, G., S. M. Shea, and M. Leventhal. 1969. J. Cell Biol. 42:617-672). Substances such as leukotriene C4 (LTC4) and thrombin, which increased the permeability of EC monolayers to ions, as measured by the electrical resistance of the monolayers, transiently increased EC [Ca++]i. To determine whether chemotaxing PMN cause similar changes in EC [Ca++]i, human umbilical vein endothelial cells (HUVEC) maintained as monolayers were loaded with fura-2. [Ca++]i was measured in single EC during PMN adhesion to and migration across these monolayers. PMN-EC adhesion and transendothelial PMN migration in response to formyl-methionyl-leucyl-phenylalanine (fMLP) as well as to interleukin 1 (IL-1) treated EC induced a transient increase in EC [Ca++]i which temporally corresponded with the time course of PMN-EC interactions. When EC [Ca++]i was clamped at resting levels with a cell permeant calcium buffer, PMN migration across EC monolayers and PMN induced changes in EC monolayer permeability were inhibited. However, clamping of EC [Ca++]i did not inhibit PMN-EC adhesion. These studies provide evidence that EC respond to stimulated PMN by increasing their [Ca++]i and that this increase in [Ca++]i causes an increase in EC monolayer permeability. Such [Ca++]i increases are required for PMN transit across an EC barrier. We suggest EC [Ca++]i regulates transendothelial migration of PMN by participating in a signal cascade which stimulates EC to open their intercellular junctions to allow transendothelial passage of leukocytes.     

A multifactorial analysis of melanoma: prognostic histopathological features comparing Clark's and Breslow's staging methods

A multifactorial analysis was used to identify the dominant prognostic variables affecting survival from a computerized data base of 339 melanoma patients treated at this institution during the past 17 years. Five of the 13 parameters examined simultaneously were found to independently influence five year survival rates: 1) pathological stage (I vs II, p = 0.0014), 2) lesion ulceration (present vs absent, p = 0.006), 3) surgical treatment (wide excision vs wide excision plus lymphadenectomy, p = 0.024), 4) melanoma thickness (p = 0.032), and 5) location (upper extremity vs lower extremity vs trunk vs head and neck, p = 0.038). Additional factors considered that had either indirect or no influence on survival rates were clinical stage of disease, age, sex, level of invasion, pigmentation, lymphocyte infiltration, growth pattern, and regression. Most of these latter variables derived their prognostic value from correlation with melanoma thickness, except sex which correlated with location (extremity lesions were more frequent on females, trunk lesions on males). This statistical analysis enabled us to derive a mathematical equation for predicting an individual patient's probability of five year survival. Three categories of risk were delineated by measuring tumor thickness (Breslow microstaging) in Stage I patients: 1) thin melanomas (<0.76 mm) were associated with localized disease and a 100% cure rate: 2) intermediate thickness melanomas (0.76-4.00 mm) had an increasing risk (up to 80%) of harboring regional and/or distant metastases and 3) thick melanomas (>/=4.00 mm) had a 80% risk of occult distant metastases at the time of initial presentation. The level of invasion (Clark's microstaging) correlated with survival, but was less predictive than measuring tumor thickness. Within each of Clark's Level II, III and IV groups, there were gradations of thickness with statistically different survival rates. Both microstaging methods (Breslow and Clark) were less predictive factors in patients with lymph node or distant metastases. Clinical trials evaluating alternative surgical treatments or adjunctive therapy modalities for melanoma patients should incorporate these parameters into their assessment, especially in Stage I (localized) disease where tumor thickness and the anatomical site of the primary melanoma are dominant prognostic factors.