CD44 ligation induces apoptosis via caspase- and serine protease-dependent pathways in acute promyelocytic leukemia cells.

We have recently reported that ligation of the CD44 cell surface antigen with A3D8 monoclonal antibody (mAb) triggers incomplete differentiation and apoptosis of the acute promyelocytic leukemia (APL)-derived NB4 cells. The present study characterizes the mechanisms underlying the apoptotic effect of A3D8 in NB4 cells. We show that A3D8 induces activation of both initiator caspase-8 and -9 and effector caspase-3 and -7 but only inhibition of caspase-3/7 and caspase-8 reduces A3D8-induced apoptosis. Moreover, A3D8 induces mitochondrial alterations (decrease in mitochondrial membrane potential DeltaPsi m and cytochrome c release), which are reduced by caspase-8 inhibitor, suggesting that caspase-8 is primarily involved in A3D8-induced apoptosis of NB4 cells. However, the apoptotic process is independent of TNF-family death receptor signalling. Interestingly, the general serine protease inhibitor 4-(2-aminoethyl)-benzenesulfonyl fluoride (AEBSF) decreases A3D8-induced apoptosis and when combined with general caspase inhibitor displays an additive effect resulting in complete prevention of apoptosis. These results suggest that both caspase-dependent and serine protease-dependent pathways contribute to A3D8-induced apoptosis. Finally, A3D8 induces apoptosis in all-trans-retinoic acid-resistant NB4-derived cells and in APL primary blasts, characterizing the A3D8 anti-CD44 mAb as a novel class of apoptosis-inducing agent in APL.     

The in vitro permeability of human skin to benzene, ethylene glycol, formaldehyde, and n-hexane

The permeability of human skin to benzene, ethylene glycol, formaldehyde, and n-hexane was studied using excised skin in a flow-through diffusion cell. The rate of resorption was determined by measuring the amount of substance found in the receptor fluid beneath the skin at steady-state. The rates of resorption (microgram X cm-2 X hr-1) were: benzene 99. ethylene glycol 118, formaldehyde from a concentrated solution of formalin 319, formaldehyde from a solution of 10% formalin in phosphate buffer 16.7, and n-hexane 0.83. The amount of substance in the skin at steady-state and after 0.5 hr of exposure was also determined. For all substances, the sum of the amount in the receptor medium and in the skin at steady-state, were larger than the amount obtained by multiplying the resorption rate by the time of exposure. For benzene, ethylene glycol and n-hexane the amount absorbed during the first half-hour of exposure was considerable larger than the amount resorbed during a same unit of time at steady-state. These data call attention to the fact that the absorption rate is higher before steady state is attained.     

Effect of oxygen withdrawal on active and passive electrical properties of arterially perfused rabbit ventricular muscle

Oxygen withdrawal from myocardial cells leads to changes of the transmembrane action potential (mainly action potential shortening), to cellular uncoupling, and to changes of vascular permeability. This study was aimed at the simultaneous measurement of electrical activity and passive electrical properties (extracellular and intracellular longitudinal resistance) in arterially perfused rabbit papillary muscles under different conditions of changed oxygen supply. These included 1) complete anoxia (erythrocyte-free perfusate), 2) hypoxia (PO2 between 23-28 mm Hg, erythrocytes present) in the presence and absence of glucose, and 3) normoxia with erythrocyte-free perfusate. Similarly to myocardial ischemia, rapid cellular uncoupling occurred only after an initial stable period of approximately 17 minutes, and it required complete anoxia. The marked shortening of the action potential developed before cellular uncoupling. In six out of eight experiments, the fibers were inexcitable when uncoupling started. In severe hypoxia, no significant change of internal longitudinal resistance was observed over 35-40 minutes. The time course of the extracellular longitudinal resistance was different from the change in intracellular resistance: A marked decrease occurred almost immediately after the onset of oxygen withdrawal. This decrease was followed by a small increase in conduction velocity, which was most likely due to a change in the interstitial compartment (edema). It was observed during anoxic as well as during hypoxic perfusion. We conclude that 1) cellular uncoupling in arterially perfused tissue requires almost complete oxygen lack and occurs with a delay of more than 10 minutes, 2) marked action potential shortening precedes uncoupling, and therefore can not simply be attributed to an increase in free, intracellular calcium, and 3) vascular endothelial function is more sensitive to oxygen withdrawal than the myocyte.     

The concept of culture: A core issue in health disparities

Contemporary discourse contains numerous examples of use of the concept of culture by social and behavioral scientists. Simple reification, where the speaker makes culture into a thing capable of action exemplifies one usage in public discourse. Some quantitative social scientists attempt to characterize people’s cultural identities by means of a single categorical variable, which often “lumps” people into categories such as “Hispanic” or “Black” that in fact have numerous culturally bounded subcategories. Approaches that emphasize cultural process are preferable to those who attempt to categorize; more complex measures of acculturation help investigators to make convincing analyses of circumstances in which health disparities occur. Examples in which investigators make appropriate use of cultural characterizations demonstrate their utility in investigating health disparities in Haitian American women, injecting and noninjecting drug users, Hispanic youth, and adult Hispanics at risk of HIV infection. Focus on culture in the study of health disparities can identify entanglements between structural factors such as poverty and lack of education and cultural factors such as beliefs about health. Qualitative methods coupled with quantitative methods have great potential to improve investigators’ grasp of cultural nuance while capturing the distribution of qualitatively derived behaviors.     

A new transthyretin variant (Glu61Gly) associated with cardiomyopathy.

We report the identification of a new transthyretin (TTR) gene mutation and variant protein, Glu61Gly, in a 55-year-old man with progressive cardiomyopathy, mild peripheral neuropathy and bilateral carpal tunnel syndrome. A diagnosis of TTR-associated familial amyloidosis (ATTR) was considered after an endomyocardial biopsy revealed amyloid deposits in the heart of a patient who had no family history of amyloidosis and no evidence of a plasma cell dyscrasia. Serum screening for a TTR variant by isoelectric focusing (IEF) was positive and prompted further studies to identify the genetic abnormality and to characterize the amyloidogenic protein. Direct DNA sequence analysis of all four coding regions in the TTR gene demonstrated heterozygosity in exon 3. Near equal amounts of guanine (G) and adenine (A) were observed at the second base position of codon 61. The wild-type (GAG) and mutated (GGG) sequences found in codon 61 correspond to glutamic acid (Glu) and glycine (Gly) residues, amino acids which differ in mass by -72 Da. Mass spectrometric analyses of TTR immunoprecipitated from serum showed the presence of both wild-type and variant proteins. The observed mass results for the wild-type and variant proteins were consistent with the predicted values calculated from the genetic analysis data.     

Discussing options between patients and health care professionals in genetic diagnosis: ethical and legal criteria

The specific characteristics of genetic data lead to ethical-legal conflicts in the framework of genetic diagnosis. Several international organisations, including UNESCO and the Council of Europe, have enacted rules referring to the use of genetic information. This paper discusses possible legal and ethical criteria that could be used in genetic testing.     

Influence of pregnancy, lactation, litter size and diet energy density on the stomach and intestine of sows

In the first experiment, 52 sows, each having raised one litter, were randomly assigned to the five following groups: control (nongravid) for pregnancy (CP), 110 d pregnancy (P110), control (nongravid) for lactation (CL), 4-wk lactation with 8 (L8) and with 12 (L12) piglets. In a second experiment, 36 sows, each having raised three litters, were randomly assigned to the following groups: control group (nongravid) fed a low-energy-density, 1% tallow diet (CLED) and two lactating groups, one fed the low-energy-density diet (LLED) and one fed a high-energy-density, 10% tallow diet (LHED). At slaughter, the stomach, small and large intestine and cecum were excised, emptied and freed from fat. Lengths and pre- and post-defatting weights were measured. Portions of tissues were homogenized and analyzed for protein, pepsin, maltase, RNA and DNA. Pregnancy had no effect on the weights of the different components of the gastrointestinal tract. Liver and small intestine weights were larger in lactating sows than in the CL group. Sows nursing 12 piglets had heavier livers than those nursing 8. The fundic mucosa of the latter had higher total pepsin activity and total protein and RNA contents than that of L12 sows. LHED sows had heavier small intestine and lower total pepsin content of the fundic mucosa than LLED sows.     

Gallibacterium anatis-secreted metalloproteases degrade chicken IgG.

Gallibacterium anatis (previously named Pasteurella haemolytica-like) is considered a normal inhabitant of genital and upper respiratory tracts of healthy chickens, but it is also associated with different pathological conditions. Secreted metalloproteases from field and reference G. anatis cultures were obtained by methanol precipitation and were characterized. Proteins of molecular mass higher than 100 kDa showing proteolytic activity were observed in 10% polyacrylamide gels copolymerized with 1% bovine casein. They were active at alkaline pH, and inhibited by ethylenediamine tetraacetic acid. Their activity was stable at 50 degrees C, but partially inhibited at 60 degrees C, and totally inhibited at higher temperatures. Secreted proteins were able to degrade chicken IgG after 24 h of incubation, and cross-reacted with a polyclonal antibody against purified protease from Actinobacillus pleuropneumoniae. Secreted metalloproteases could play a role in infections caused by G. anatis.