Dulaglutide decreases plasma aminotransferases in people with Type 2 diabetes in a pattern consistent with liver fat reduction: a post hoc analysis of the AWARD programme.

AIMS: To evaluate the effects of dulaglutide vs placebo on liver and glycaemic/metabolic measurements in a population with Type 2 diabetes and in a subgroup with non-alcoholic fatty liver/non-alcoholic steatohepatitis. METHODS: A total of 1499 participants from AWARD-1, AWARD-5, AWARD-8 and AWARD-9 clinical trials were included in this analysis (dulaglutide 1.5 mg, n=971 and placebo, n=528). Thresholds of alanine aminotransferase levels ≥30 IU/l in men and ≥19 IU/l in women were used to determine the subgroup who had non-alcoholic fatty liver/non-alcoholic steatohepatitis. Objectives included changes from baseline to 6 months in: (1) alanine aminotransferase, aspartate transaminase and gamma-glutamyl transpeptidase levels in the overall population and (2) alanine aminotransferase, aspartate transaminase, gamma-glutamyl transpeptidase and glycaemic/metabolic measurements (e.g. HbA1c , fasting serum glucose, body weight, lipids and homeostatic model assessment) in the non-alcoholic fatty liver/non-alcoholic steatohepatitis subgroup. RESULTS: In the overall population at 6 months, dulaglutide significantly reduced alanine aminotransferase, aspartate transaminase and gamma-glutamyl transpeptidase levels vs placebo [least squares mean treatment differences: -1.7 IU/l (95% CI -2.8, -0.6), P=0.003; -1.1 IU/l (95% CI -2.1, -0.1), P=0.037; -6.6 IU/l (95% CI -12.4, -0.8), P=0.025, respectively]. In the subgroup with non-alcoholic fatty liver/non-alcoholic steatohepatitis (alanine aminotransferase levels greater than or equal to the upper limit of normal), mean baseline liver enzyme values were 38.0 IU/l, 27.8 IU/l and 43.9 IU/l for alanine aminotransferase, aspartate transaminase and gamma-glutamyl transpeptidase, respectively. In this population, more pronounced reductions from baseline in alanine aminotransferase were observed with dulaglutide vs placebo (-8.8 IU/l vs -6.7 IU/l). In the subgroup of people with alanine aminotransferase levels less than the upper limit of normal, changes from baseline in alanine aminotransferase did not significantly differ between treatment groups (0.0 IU/l vs 0.7 IU/l). CONCLUSIONS: Once-weekly dulaglutide improved alanine aminotransferase, aspartate transaminase and gamma-glutamyl transpeptidase levels compared with placebo in a pattern consistent with liver fat reductions. Our results add further weight to the notion that glucagon-like peptide-1 receptor agonists may provide benefit in lowering liver fat in addition to their other metabolic actions.

[Safety and efficacy of growth hormone treatment: GeNeSIS study in Spain]. / Seguridad y efectividad del tratamiento con hormona de crecimiento: estudio GeNeSIS en España.

INTRODUCTION: Country-specific information on pediatric GH therapy is available from multi-national studies. METHODS: A total of 1294 children in Spain enrolled in the observational Genetics and Neuroendocrinology of Short-stature International Study (GeNeSIS). Adverse events were assessed in all GH-treated patients (n=1267) and effectiveness in those with GH deficiency (GHD, 78%). RESULTS: Mean age at time of entry to the study was 9.8 years. GH was initiated at a median (Q1-Q3) 0.22 (0.20-0.25) mg/kg/week and administered for 2.8 (1.6-4.4) years. For 262 patients with GHD and 4-year data, mean (95% CI) height velocity was 4.3 (4.1 - 4.6) cm/year at baseline, 9.0 (8.7 to 9.4) cm/year at 1-year, and 5.5 (5.2 to 5.8) cm/year at 4-years. Height standard deviation score (SDS) was -2.48 (-2.58 to -2.38) at baseline and -1.18 (-1.28 to -1.08) at 4 years. Final height SDS minus target height SDS (n=241) was -0.09 (-0.20 to 0.02). In 1143 GH-treated patients with ≥1 year follow-up, 93 (8.1%) reported treatment-emergent adverse events. Serious events were reported for 7 children, with 2 considered GH-related. CONCLUSION: These data confirm the benefit of GH replacement therapy on height gain for the patients in Spain. The safety profile was consistent with that already known for GH therapy.

Acute myelogenous leukemia in Down's syndrome: report of a single pediatric institution using a BFM treatment strategy.

Between July 1990 and December 1995, 111 new consecutive pediatric patients with acute myelogenous leukemia (AML) have been treated in our institution. Eleven of them (9.9%) had Down's syndrome (DS), 6 boys and 5 girls. The median age was 22.5 (range 10-40) months. FAB subtypes were the following: M7: 6, M4: 3, and M0: 2. Five of them had previously had myelodysplasia and in 3, all FAB M7, myelofibrosis was detected. This population was treated with two consecutive protocols. Nine patients were included in the AML-HPG-90 protocol and 2 patients in the AML-HPG-95 study, respectively. However, all DS patients in this series received the same treatment. Eight patients achieved complete remission: two patients received two cycles of intensification with high dose (HD) ara-C, and 1 patient, only one cycle; the other 5 were prevented from receiving such therapy because of unacceptable toxicity or death. At 45 months, event-free survival and overall survival estimates were 0.30, S.E. 0.16. Mortality was remarkably high. All deaths (7) were associated with sepsis (5) or pulmonary infection (2). Three deaths occurred before achieving complete remission, 3 patients died during the consolidation phase and 1 died whilst off treatment. No one presented leukemic relapse. We conclude that this AML-BFM treatment strategy is highly toxic to children with DS and AML in our setting. Efforts will be made to improve clinical support and to administer less intensive therapy to this particular pediatric AML subgroup, which, in fact, has a better prognosis than the same non-trisomic population.

Fertility following fimbriectomy and tubo-ovarian microsurgery in the rabbit.

It is generally believed that tubal fimbriae function as a highly specialized and indispensable component of the ovum pickup mechanism. The present study was undertaken to examine the feasibility of creating a functional tubo-ovarian relationship following fimbriectomy. Fourteen New Zealand White rabbits underwent unilateral microsurgical fimbriectomy followed by creation of a distal ampullary flap which was oversewn to the exposed ovarian cortex. Two weeks after surgery, the animals were inseminated and induced to ovulate with an intravenous injection of 100 IU of human chorionic gonadotropin. Thirteen of fourteen uterine horns on the control side and ten of fourteen on the operated side became pregnant. The ratios of ova ovulated to those which implanted were 73.2% on the control side and 37.8% on the operated side. Morphologic studies revealed a normal complement of ciliated and secretory cells lining the endosalpinx of the juxtaposed ampullary flap. The portion of ovarian cortex covered by the flap similarly exhibited no morphologic abnormalities. These findings indicate that fertility can be maintained in the absence of fimbriae and suggest a surgical technique for restoration of fertility following elective sterilization by fimbriectomy or following pathologically induced destruction of the fimbriae.

Effect of zaleplon on digoxin pharmacokinetics and pharmacodynamics.

The pharmacokinetics and pharmacodynamics of digoxin alone and digoxin plus zaleplon were studied. Healthy, nonsmoking men between 18 and 45 years of age were given a single oral dose of digoxin 0.375 mg daily on days 1 through 9. On days 10 through 14, the subjects received digoxin 0.375 mg plus oral zaleplon 10 mg daily. Blood samples were obtained on days 3, 5, 8, 9, and 14, and serum digoxin concentration data were analyzed by model-independent pharmacokinetic methods. Blood pressure, heart rate, PR interval, and QTc interval were recorded to determine the effect of zaleplon on digoxin pharmacodynamics. A total of 20 men completed the study. Maximum serum digoxin concentration and area under the serum digoxin concentration-versus-time curve from 0 to 24 hours met bioequivalence test criteria. There were no significant differences in QTc or PR interval between days 9 (digoxin alone) and 14 (digoxin plus zaleplon), and there were no clinically important changes from baseline to the study's end in vital signs, physical examination findings, or ECG results for individual subjects. Eighteen percent of the subjects who received digoxin alone and 35% of those who received digoxin plus zaleplon reported one or more adverse effects; all were mild and resolved quickly. Zaleplon had no significant effects on selected pharmacokinetic and pharmacodynamic properties of digoxin.

[Influence of HIV/AIDS infection on some biochemical indicators of the nutritional status]. / Influencia de la infección por VIH/SIDA sobre algunos indicadores bioquímicos del estado nutricional.

The main objective of this study was to analyze the influence of nutritional state among HIV-1 infected people, according to the different clinical stages referred by the CDC (Control Disease Center of the United States) in 1987, as well as the changes in the concentrations of some biochemical markers linked to nutritional state. A similar study was carried out in a control group with UltramicroELISA non-reagent healthy individuals, anthropometrically classified. Concentrations of total proteins, albumin, cholesterol, triglycerides, urea, uric acid and creatinine were analyzed by sex and clinical group, comparing the levels obtained through a variance study. When comparing HIV-1 asymptomatic infected patients to HIV-1 and HIV-2 non infected people, the results showed a non significant increase in the level of total proteins with a significant decrease of albumin and creatinine, the latter observed only in male patients. In stage IV patients, an important decrease of cholesterol and a significant increase of the triglycerides were found, as well as the lowest albumin levels. Urea and uric acid levels did not experience statistically significant changes. It was concluded that the study of biochemical markers is advisable, since it contributes to the detection by default of malnutrition marginal states in infected individuals.

Prenatal laboratory screening.

Expectations of the clinician are evolving in obstetric practice in regard to standards of performance, cost effectiveness, and outcomes for mother and child. Prenatal laboratory screening is shown to be an area where new developments are evident. The purpose of this article is to update information found in standard obstetrics textbooks.

Identification of vitellogenin in the little skate (Raja erinacea).

1. Vitellogenin was isolated from mature female skates by selective precipitation with MgCl2/EDTA followed by chromatography on DEAE-cellulose columns. 2. A single monomer of approximately 205 kDa was identified on 6.0% SDS-PAGE gels. 3. In addition, isolation of yolk proteins with ammonium sulfate yielded proteins of 94 and 38 kDa (putative phosvitins) and putative lipovitellins of ca 105, 91 and 67 kDa. 4. In vivo phosphate incorporation in female and male skates implanted with estradiol indicated that vitellogenin was phosphorylated. 5. Total protein phosphate incorporation was significantly higher in females than male skates. 6. In male skates treated with estradiol, phosphate incorporation increased from 2 days after implantation to a maximum at approximately 11 days after implantation. 7. Determination of the rate of disappearance of 32P-labeled protein suggests a half-life of ca 200 hr in normal female skate plasma.

Vitellogenin--homologs of mammalian apolipoproteins?

1. To determine if a relationship exists between vertebrate vitellogenins and mammalian plasma proteins the EMBL and NBRF computer databases were searched with two partial amino acid sequences from Xenopus laevis and Gallus gallus vitellogenin. 2. A significant relationship was found between vitellogenin and human apolipoprotein B-100 genes, and confirmed using homology-determination programs. 3. Further analysis shows that unique multiple proline consensus regions found in apolipoprotein B-100 are significantly similar to proline dominant regions in vitellogenin. 4. This work suggests that these proteins are functionally and structurally related and should be categorized as a functional group of hepatic lipid transport and metabolism proteins.

Putative apolipoprotein B-100 in the freshwater turtle Chrysemys picta: effects of estrogen and progesterone.

1. The isolation and purification of a putative apolipoprotein B-100 in the plasma of the freshwater turtle Chrysemys picta is described. 2. The protein was purified through differential ultracentrifugation and subsequent Sepharose 6B column chromatography. 3. The molecular weight of the protein determined by electrophoresis was approximately 350 kDa. 4. An antibody to chicken apolipoprotein B-100 specifically recognizes this 350 kDa protein in Western blots, suggesting its identity with apolipoprotein B-100. 5. An antibody to the putative Chrysemys apolipoprotein B-100-like protein was developed and used in an ELISA to quantitate protein levels in plasma. 6. Acute estrogen treatment increased levels of apolipoprotein B-100 (7.64 +/- 0.79 mg/ml plasma) over that of control animals (5.07 +/- 1.74 mg/ml plasma). 7. In contrast, chronic estrogen treatment reduced apolipoprotein B-100 significantly to 2.94 +/- 0.53 mg/ml plasma (P < 0.05).

A kinematic study of crawling behavior in the leech, Hirudo medicinalis.

The medicinal leech crawls along a solid substrate by repeated alternating extensions and shortenings of the body. Extension occurs with the posterior sucker attached and the head sucker free. The head sucker then attaches, followed by shortening and release of the tail sucker. The tail sucker is then pulled toward the head, where it reattaches to the substrate. The head sucker then releases, and another crawling cycle begins (Figs. 1, 5). There are two crawling variants: inchworm crawling, in which the head and tail suckers are closely apposed at the end of a cycle and the body forms a loop above the substrate, and vermiform crawling, in which the suckers are placed farther apart and the body remains fairly close to the substrate (Fig. 1). The cycle period and the distance traveled during a cycle are greater in inchworm than in vermiform crawling; however, the velocity of travel is the same for both (Fig. 2). For both variants, the interval between head sucker attachment and tail sucker release is similar at all cycle periods and has a value consistent with direct interneuronal conduction of a signal from head sucker sensory neurons to tail sucker motor neurons. The interval between tail sucker attachment and head sucker release, however, is longer and varies with the cycle period, suggesting a more complex interneuronal circuit in the pathway from tail sucker sensory neurons to head sucker motor neurons (Fig. 4). The onsets of the components of the crawling cycle (extension, post-extension pause, shortening, and post-shortening pause) show an anteroposterior lag (Figs. 5, 7). For both variants, the travel time between segments varies directly with the period (Fig. 8). For both crawl types, the durations of the cycle components vary directly with the period, with several exceptions (Figs. 9, 10). A model is presented that summarizes the coordination of the various motor events in a cycle of leech crawling (Figs. 11 and 12).

Leishmania donovani p36(LACK) DNA vaccine is highly immunogenic but not protective against experimental visceral leishmaniasis.

The acquisition of immunity following subclinical or resolved infection with the intracellular parasite Leishmania donovani suggests that vaccination could prevent visceral leishmaniasis (VL). The LACK (Leishmania homolog of receptors for activated C kinase) antigen is of interest as a vaccine candidate for the leishmaniases because of its immunopathogenic role in murine L. major infection. Immunization of mice with a truncated (24-kDa) version of the 36-kDa LACK antigen, delivered in either protein or DNA form, was found previously to protect against cutaneous L. major infection by redirecting the early T-cell response away from a pathogenic interleukin-4 (IL-4) response and toward a protective Th1 response. The amino acid sequence of the Leishmania p36(LACK) antigen is highly conserved, but the efficacy of this vaccine antigen in preventing disease caused by strains other than L. major has not been determined. We investigated the efficacy of a p36(LACK) DNA vaccine against VL because of the serious nature of this form of leishmaniasis and because it was unclear whether the LACK vaccine would be effective in a model where there was not a dominant pathogenic IL-4 response. We demonstrate here that although the LACK DNA vaccine induced a robust parasite-specific Th1 immune response (IFN-gamma but not IL-4 production) and primed for an in vivo T-cell response to inoculated parasites, it did not induce protection against cutaneous or systemic L. donovani challenge. Coadministration of IL-12 DNA with the vaccine did not enhance the strong vaccine-induced Th1 response or augment a protective effect.

Xenotransplantation of immunodeficient mice with mobilized human blood CD34+ cells provides an in vivo model for human megakaryocytopoiesis and platelet production.

The study of megakaryocytopoiesis has been based largely on in vitro assays. We characterize an in vivo model of megakaryocyte and platelet development in which human peripheral blood stem cells (PBSCs) differentiate along megakaryocytic as well as myeloid/lymphoid lineages in sublethally irradiated nonobese diabetic/severe combined immunodeficient (NOD-SCID) mice. Human hematopoiesis preferentially occurs in the bone marrow of the murine recipients, and engraftment is independent of exogenous cytokines. Human colony-forming units-megakaryocyte (CFU-MK) develop predominantly in the bone marrow, and their presence correlates with the overall degree of human cell engraftment. Using a sensitive and specific flow cytometric assay, human platelets are detected in the peripheral blood from weeks 1 to 8 after transplantation. The number of circulating human platelets peaks at week 3 with a mean of 20 x 10(9)/L. These human platelets are functional as assessed by CD62P expression in response to thrombin stimulation in vitro. Exogenous cytokines have a detrimental effect on CFU-MK production after 2 weeks, and animals treated with these cytokines have no circulating platelets 8 weeks after transplantation. Although cytokine stimulation of human PBSCs ex vivo led to a significant increase in CFU-MK, CD34+/41+, and CD41+ cells, these ex vivo expanded cells provided only delayed and transient platelet production in vivo, and no CFU-MK developed in vivo after transplantation. In conclusion, xenogeneic transplantation of human PBSCs into NOD/SCID mice provides an excellent in vivo model to study human megakaryocytopoiesis and platelet production.

Mechanical testing of bioresorbable implants for use in metacarpal fracture fixation.

The purpose of this study was to evaluate the mechanical properties of a bioresorbable plate and screw system that was developed for the treatment of unstable metacarpal fractures and to compare the strength and stiffness of this system with those of conventional titanium plates and screws. Using a 4-point bending test, we measured the strength and stiffness of these implants over a 12-week period of in vitro degradation. Our data suggest that these implants provide stable bending strength and stiffness for 8 weeks and gradually lose their strength over a period of 12 weeks. Further research is necessary to determine whether this level of fixation is adequate to stabilize unstable metacarpal fractures.

Malnutrition alters the innate immune response and increases early visceralization following Leishmania donovani infection.

Malnutrition is a risk factor for the development of visceral leishmaniasis. However, the immunological basis for this susceptibility is unknown. We have developed a mouse model to study the effect of malnutrition on innate immunity and early visceralization following Leishmania donovani infection. Three deficient diets were studied, including 6, 3, or 1% protein; these diets were also deficient in iron, zinc, and calories. The control diet contained 17% protein, was zinc and iron sufficient, and was provided ab libitum. Three days after infection with L. donovani promastigotes, the total extradermal (lymph nodes, liver, and spleen) and skin parasite burdens were equivalent in the malnourished (3% protein) and control mice, but in the malnourished group, a greater percentage (39.8 and 4.0%, respectively; P = 0.009) of the extradermal parasite burden was contained in the spleen and liver. The comparable levels of parasites in the footpads in the two diet groups and the higher lymph node parasite burdens in the well-nourished mice indicated that the higher visceral parasite burdens in the malnourished mice were not due to a deficit in local parasite killing but to a failure of lymph node barrier function. Lymph node cells from the malnourished, infected mice produced increased levels of prostaglandin E(2) (PGE(2)) and decreased levels of interleukin-10. Inducible nitric oxide synthase activity was significantly lower in the spleen and liver of the malnourished mice. Thus, malnutrition causes a failure of lymph node barrier function after L. donovani infection, which may be related to excessive production of PGE(2) and decreased levels of IL-10 and nitric oxide.

Separation between cytosolic calcium and secretion in chromaffin cells superfused with calcium ramps.

This paper describes experiments in which cytosolic Ca2+ concentrations ([Ca2+]i) and catecholamine release were measured in two populations of chromaffin cells stimulated with a solution enriched in K+ (100 mM). Once depolarized, external Ca2+ or Ba2+ ions were offered to cells either as a single 2.5 mM step or as a ramp that linearly increased the concentration from 0 to 2.5 mM over a 10-min period. A clear separation between the changes of the [Ca2+]i and the time course of secretion was observed. Specifically, secretion and [Ca2+]i rose in parallel when a Ca2+ step was used to reach a peak in a few seconds; however, while secretion declined to the basal level, [Ca2+]i remained elevated at a plateau of 400 nM. With a Ca2+ ramp, only a transient small peak of secretion was observed, yet the [Ca2+]i remained elevated throughout the 10-min stimulation period. The separation between secretion and [Ca2+]i was observed even when voltage-dependent Ca2+ channels were expected to remain open (mild depolarization in the presence of 1 microM Bay K 8644). By using Ba2+ steps or ramps, sustained noninactivating secretory responses were obtained. The results suggest that the rate and extent of secretion are not a simple function of the [Ca2+]i at a given time; they are compatible with the following conclusions: (i) A steep extracellular-to-cytosolic Ca2+ gradient is required to produce a sharp increase in the [Ca2+]i at exocytotic sites capable of evoking a fast but transient secretory response. (ii) As a result of Cai(2+)-dependent inactivation of Ca2+ channels, those high [Ca2+]i are possible only at early times after cell depolarization. (iii) The Cai(2+)-dependent supply of storage granules to the secretory machinery cooperates with the supply of Ca2+ through Ca2+ channels to regulate the rate and extent of secretion.

Effects of tyramine and calcium on the kinetics of secretion in intact and electroporated chromaffin cells superfused at high speed.

Fast superfusion of electroporated bovine adrenal chromaffin cells with a K+ glutamate-based solution containing 50 nM free Ca2+ and 2 mM adenosine 5'-triphosphate, dipotassium salt (K2ATP), produced a steady-state low catecholamine secretion, measured on-line with an electrochemical detector (about 20 nA). Rapid switching to electroporation solutions containing increasing Ca2+ concentrations ([Ca2+]) produced a rapid increase in the rate and peak secretion, followed by a decline. At intermediate [Ca2+] (3-100 microM), a fast peak and a slow secretory plateau were distinguished. The fast secretory peak identifies a readily releasable catecholamine pool consisting of about 200-400 vesicles per cell. Pretreatment of cells with tyramine (10 microM for 4 min before electroporation) supressed the initial fast secretory peak, leaving intact the slower phase of secretion. With [Ca2+] in the range of 0.1-3 microM, the activation rate of secretion increased from 2.3 to 35.3 nA.s-1, reached a plateau between 3-30 microM and rose again from 100 to 1000 microM [Ca2+] to a maximum of 91.9 nA.s-1. In contrast, total secretion first increased (0.1-1 microM Ca2+), then plateaud (1-100 microM Ca2+) and subsequently decreased (100-1000 microM Ca2+). At 30 and 1000 microM extracellular [Ca2+] or [Ca2+]o, the activation rates of secretion from intact cells depolarised with 70 mM K+ were close to those obtained in electroporated cells. However, secretion peaks were much lower in intact (93 nA at 30 microM Ca2+) than in electroporated cells (385 nA). On the other hand, inactivation of secretion was much faster in intact than in electroporated cells; as a consequence, total secretion in a 5-min period was considerably smaller in intact (10.6 microA.s at 1000 microM Ca2+) than in electroporated cells (42.4 microA.s at 1 microM Ca2+). Separation of the time-courses of changes in intracellular [Ca2+] or [Ca2+]i and secretion in intact chromaffin cells depolarised with 70 mM K+ was demonstrated at different [Ca2+]o. The increase in the rate of catecholamine release was substantially higher than the increase of the average [Ca2+]i. In contrast, the decline of secretion was faster than the decline of the peak [Ca2+]i. The results are compatible with the idea that the peak and the amount of catecholamine released from depolarised intact cells is determined essentially by plasmalemmal factors, rather than by vesicle supply from reserve pools. These plasmalemmal factors limit the supply of Ca2+ by the rates of opening and closing of voltage-dependent Ca2+ channels of the L- and Q-subtypes, which control the local [Ca2+]i near to exocytotic sites.