CBL-B is required for leukemogenesis mediated by BCR-ABL through negative regulation of bone marrow homing.

BCR-ABL induces chronic myeloid leukemia (CML) through the aberrant regulation of multiple signaling substrates. Previous research has shown that BCR-ABL mediates down-modulation of CBL-B protein levels. A murine bone marrow transplantation (BMT) study was performed to assess the contribution of Cbl-b to BCR-ABL-induced disease. The predominant phenotype in the Cbl-b(-/-) recipients was a CML-like myeloproliferative disease (MPD) similar to that observed in the wild-type animals, but with a longer latency, diminished circulating leukocyte numbers and reduced spleen weights. Despite the decreased leukemic burden in comparison to their wild-type counterparts, the Cbl-b(-/-) animals displayed enhanced numbers of Gr-1(+)/Mac-1(+) spleen cells and neutrophilia. On the basis of prior evidence of CBL-B-dependent motility toward SDF-1α, we hypothesized that Cbl-b deficiency might impair bone marrow localization during transplantation. Homing experiments showed reduced migration of Cbl-b(-/-) cells to the bone marrow. Intrafemoral transplantation of BCR-ABL-transduced Cbl-b(-/-) cells revealed equivalent latency of disease development to the wild-type transplants, supporting the conclusion that Cbl-b deficiency diminishes homing of leukemic cells to the bone marrow, and perturbs the proliferation of BCR-ABL-expressing malignant clones during CML development.

Functionally important residues in the peptidyl-prolyl isomerase Pin1 revealed by unigenic evolution.

Pin1 is a phosphorylation-dependent member of the parvulin family of peptidyl-prolyl isomerases exhibiting functional conservation between yeast and man. To perform an unbiased analysis of the regions of Pin1 essential for its functions, we generated libraries of randomly mutated forms of the human Pin1 cDNA and identified functional Pin1 alleles by their ability to complement the Pin1 homolog Ess1 in Saccharomyces cerevisiae. We isolated an extensive collection of functional mutant Pin1 clones harboring a total of 356 amino acid substitutions. Surprisingly, many residues previously thought to be critical in Pin1 were found to be altered in this collection of functional mutants. In fact, only 17 residues were completely conserved in these mutants and in Pin1 sequences from other eukaryotic organisms, with only two of these conserved residues located within the WW domain of Pin1. Examination of invariant residues provided new insights regarding a phosphate-binding loop that distinguishes a phosphorylation-dependent peptidyl-prolyl isomerase such as Pin1 from other parvulins. In addition, these studies led to an investigation of residues involved in catalysis including C113 that was previously implicated as the catalytic nucleophile. We demonstrate that substitution of C113 with D does not compromise Pin1 function in vivo nor does this substitution abolish catalytic activity in purified recombinant Pin1. These findings are consistent with the prospect that the function of residue 113 may not be that of a nucleophile, thus raising questions about the model of nucleophilic catalysis. Accordingly, an alternative catalytic mechanism for Pin1 is postulated.

Development of a field test for upper-body power.

The purpose of this study was to develop a field test capable of measuring upper-body power through the use of a common weight-training apparatus, a Smith machine (SM), set up for bench press (BP) movement. A small, battery-operated digital timing device was designed and constructed to allow a precise calculation of power (in conjunction with measures of distance and force) for this specific movement, which involved an explosive press from the chest to a position just short of full arm extension. In pilot work, 1 repetition maximums (1RM) were determined on the SM BP for 3 male subjects, and by subsequently testing power on the same subjects at varying resistances, an average relative percentage of the 1RM-producing peak power values was found by power curve analysis for test standardization. Reliability was assessed (using 11 men) by SM power measurements taken over 3 days on the SM fitted with the timer. An intraclass R (0.998) indicated a high correlation between the 3 separate field-test trials. Finally, 8 male subjects were used to compare SM scores with a criterion measure, the Linea Isokinetic BP station (Loredan Biomedical, Inc., Sacramento CA). A Pearson product moment coefficient found a high correlation between the field test (SM) and Linea power scores (r = 0.987). A 2-tailed dependent t-test between the field and criterion scores was not significant, suggesting that no consistent error variable was present. It can be concluded that this is a valid field test of power for this movement.

Care coordination in managed care. Creating a quality continuum for high risk elderly patients.

The portion of the American population 65 years and older is growing rapidly, and the group 85 years and older is the largest growing segment of our population. The country's largest health maintenance organization, Kaiser Permanente, in Fontana, California, experienced a 9% growth in its membership between 1996 and 1997. Following the Kaiser national Model of Care recommendations, Fontana's Kaiser Permanente created their department of Extended Care Services and implemented programs that coordinated patient care across the continuum to meet the needs of its members who were 65 years and older. The programs included new member screening and orientation, telephonic care coordination, and a volunteer-based care call program. They also developed a medical management model that included a consultative geriatric assessment clinic and a primary care clinic as well as provision of team-based care to members residing in skilled nursing facilities.

Effects of perturbations in hepatic free and esterified cholesterol pools on bile acid synthesis, cholesterol 7 alpha-hydroxylase, HMG-CoA reductase, acyl-CoA:cholesterol acyltransferase and cytosolic cholesteryl ester hydrolase.

Effects of expansion of the hepatic free cholesterol pool on bile acid and cholesterol metabolism and homeostasis were examined in rats fed cholesterol in high-fat diets or treated with oleyl-p-(n-decyl)-benzenesulfonate (ODS) or progesterone. Cholesterol feeding for 10-16 days, which increased free (33%) and esterified (6-fold) cholesterol, had no effect on cholate synthesis, total bile acid synthesis, or cholate turnover, whereas these activities were increased 60-80% by ODS and progesterone, which produced only small increases (19%) in free cholesterol. Cholesterol feeding reduced beta-hydroxy-beta-methylglutaryl (HMG)-CoA reductase (72%) and cholesteryl ester hydrolase (48%) and increased acyl-CoA:cholesterol acyltransferase (184%), whereas ODS and progesterone reversed these compensatory responses in cholesterol-fed rats. Cholesterol 7 alpha-hydroxylase was changed no more than 22% by any treatment. A bolus of ODS elevated biliary cholesterol output 41% and shifted biliary bile acid synthesis and composition toward 12-deoxy bile acids. These effects were not seen in ODS-fed or progesterone-treated rats, in which cholesteryl ester stores were depleted. It is concluded that effects of free cholesterol on bile acid synthesis and biliary cholesterol are probably mediated by specific precursor or regulatory pools which can be independently regulated and which represent a relatively small fraction of hepatic free cholesterol.

Regulation of bile acid synthesis. II. Effect of bile acid feeding on enzymes regulating hepatic cholesterol and bile acid synthesis in the rat.

Bile acid synthesis is believed to be regulated by bile salts returning to the liver via the portal vein and suppressing cholesterol 7 alpha-hydroxylase, the rate-limiting enzyme in the bile acid biosynthesis pathway. In order to characterize the relative effectiveness of bile salts in regulating bile acid synthesis, seven different bile acids were administered (1% w/w in chow) to rats over a 14-day period. Biliary bile salt composition was determined from bile samples obtained prior to killing; in all cases, the fed bile acid became the predominant bile salt in bile. The specific activities of microsomal cholesterol 7 alpha-hydroxylase, HMG-CoA reductase and acylconenzyme A:cholesterol acyltransferase were determined after killing. Hydrophilic bile salts (ursocholic, hyocholic, ursodeoxycholic and hyodeoxycholic) did not inhibit HMG-CoA reductase or cholesterol 7 alpha-hydroxylase activities. By contrast, more hydrophobic bile salts (cholic, chenodeoxycholic and deoxycholic) inhibited the activities of these two enzymes in order of increasing hydrophobicity. Neither hydrophobic nor hydrophilic bile salts inhibited acylcoenzyme A:cholesterol acyltransferase activity. No consistent effect of bile acid feeding on total microsomal cholesterol was observed. Based on the results of these studies, we propose that the hydrophilic-hydrophobic balance of the bile acid pool may play an important role in the regulation of bile acid synthesis. We postulate that the activities of cholesterol 7 alpha-hydroxylase and HMG-CoA reductase may be regulated by hydrophobic bile acid-induced changes in the lipid composition and physicochemical properties (fluidity) of the microsomal membranes to which both of these rate-limiting enzymes are attached.(ABSTRACT TRUNCATED AT 250 WORDS)

Randomized trial of pneumatic antishock garments in the prehospital management of penetrating abdominal injuries.

Experimental data have suggested that pneumatic external counterpressure improves outcome in intra-abdominal hemorrhage by either a tamponade effect and/or elevation in central systemic blood pressure. As a result, the empiric use of the pneumatic antishock garment (PASG) has become a standard of care, even to the point where the device has been legislated as required equipment on emergency medical rescue vehicles. However, the effect of the PASG on intra-abdominal hemorrhage has not been evaluated in randomized clinical trials. The purpose of this study was to evaluate the effect of the PASG on the survival of hypotensive patients with penetrating abdominal injuries. During a 2 1/2-year period, 201 consecutive patients presenting with penetrating anterior abdominal injuries and an initial prehospital systolic blood pressure of 90 mm Hg or less were entered into the study. All prehospital care was delivered by the same municipal emergency medical services system, and all patients subsequently were transported to the same regional trauma facility. The patients were randomized into control and pneumatic external counterpressure groups by an alternate-day assignment of PASG use. The resulting study populations (control, n = 104; PASG, n = 97) were found to be well matched for survival probability indices, prehospital response and transport times, and the volume of IV fluids received. The results demonstrated no significant difference in the survival rates of the control and PASG treatment groups (81 of 104 vs 67 of 97). From these data we conclude that, contrary to previous claims, the PASG provides no significant advantage in improving survival in the urban prehospital management of penetrating abdominal injuries.

The relationship between total prehospital time and outcome in hypotensive victims of penetrating injuries.

Most authorities in the field of trauma recommend that seriously injured patients be transported directly to a regional trauma center, even if it requires bypassing nearby community hospitals. The purpose of our study was to examine the relationship between the survival rates of patients with presumed hemorrhagic shock due to penetrating injuries and the total prehospital time required to manage and deliver those patients to a single regional trauma center in a large urban area. During a 30-month-period, 498 consecutive victims of penetrating injury, presenting in the field with a systolic blood pressure of 90 mm Hg or less and transported to a single regional trauma center, were prospectively evaluated in terms of age; initial prehospital trauma score; injury severity score (ISS); TRISS probability of survival; response, scene, transport, and total prehospital times; and survival (discharge from hospital). All patients were managed and transported by a single urban paramedic service that has a fairly uniform response time (5.3 +/- 3 min) throughout its entire service area. The response area is spread out over an approximately 1,000 square-mile region, and transport times to the regional trauma center can exceed a half hour. The total prehospital time (TPT) was calculated as the time elapsed from the receipt of the emergency call to the time of arrival at the regional trauma center. Patients arbitrarily were categorized into four subsets according to the initial prehospital trauma score (1, 2 to 6, 7 to 11, 12 to 15). Patients also were analyzed in terms of four incremental groups of increasing TPT (0-20, 21-30, 31-40, greater than 40 min).(ABSTRACT TRUNCATED AT 250 WORDS)

Protein kinase-mediated activation of temperature-labile and temperature-stable cholesteryl ester hydrolases in the rat testis.

Both temperature-stable and temperature-labile testicular cholesteryl ester hydrolases are shown to be regulated by an endogenous cAMP-dependent protein kinase activity. The temperature-stable form (Mr = 28,000) was activated 3-fold by the endogenous kinase. This activation was completely blocked by protein kinase inhibitor. Following purification by high performance gel permeation chromatography, the temperature-stable form could also be activated 2-fold by bovine heart protein kinase, type I. The partially purified endogenous protein kinase, type I, which was completely separated from hydrolase activity by ion exchange chromatography, increased hydrolase activity 2-fold in the presence of optimal concentrations of cAMP, ATP, and Mg2+. Cholesteryl ester hydrolase activity could be stabilized indefinitely at -10 degrees C with the addition of 0.1 mM thioglycolate, but not by other thiol reagents. In contrast, the endogenous protein kinase activity was lost from 104,000 X g supernatants after 14 days. However, the property of activation could be restored by addition of bovine heart protein kinase. The temperature-labile hydrolase (Mr = 72,000) could be totally inactivated by a Mg2+-dependent, fluoride-sensitive cytosolic factor and reactivated by cAMP-dependent protein kinase. These observations strongly suggest that the inactivating factor is a phosphoprotein phosphatase.

Acute nephrotoxicity of N-phenyl and N-(monochlorophenyl) succinimides in Fischer 344 and Sprague-Dawley rats.

The experimental fungicide N-(3,5-dichlorophenyl)succinimide (NDPS) has been shown to be nephrotoxic in Sprague-Dawley and Fischer 344 rats. The purpose of this study was to evaluate the role of the chlorine atoms in NDPS-induced nephropathy. Male Sprague-Dawley or Fischer 344 rats received a single intraperitoneal injection of a phenylsuccinimide (0.4 or 1.0 mmol/kg) or sesame oil (2.5 ml/kg), and renal function was monitored at 24 h and 48 h. In Sprague-Dawley rats urine volume was increased by NDPS and N-(3-chlorophenyl)succinimide (3-NCPS) (0.4 and 1.0 mmol/kg) at 24 h but only by NDPS at 48 h. Accumulation of both p-aminohippurate (PAH) and tetraethylammonium (TEA) was decreased only by NDPS (1.0 mmol/kg) administration. N-(2-chlorophenyl)succinimide (2-NCPS) or N-(4-chlorophenyl) succinimide (4-NCPS) (1.0 mmol/kg) administration reduced only basal and lactate-stimulated PAH accumulation. Only NDPS increased blood urea nitrogen (BUN) concentration and kidney weight. In Fischer 344 rats results were similar to those obtained in Sprague-Dawley rats, except that 3-NCPS was the only monochlorophenylsuccinimide which produced a decrease in PAH accumulation by renal cortical slices. N-Phenylsuccinimide had little effect on any renal parameter studied in either rat strain. The order of increasing nephrotoxicity generally paralleled the increasing partition. coefficients of the compounds. These results indicate that reducing the chlorine substitution of NDPS produces compounds with reduced nephrotoxic potential. In addition, lipophilic character might be a predictor for the nephrotoxic potential of N-(halophenyl)succinimides in Sprague-Dawley and Fischer 344 rats.