Histone acetyltransferase inhibitors block neuroblastoma cell growth in vivo.

We have previously described novel histone acetyltransferase (HAT) inhibitors that block neuroblastoma cell growth in vitro. Here we show that two selected pyridoisothiazolone HAT inhibitors, PU139 and PU141, induce cellular histone hypoacetylation and inhibit growth of several neoplastic cell lines originating from different tissues. Broader in vitro selectivity profiling shows that PU139 blocks the HATs Gcn5, p300/CBP-associated factor (PCAF), CREB (cAMP response element-binding) protein (CBP) and p300, whereas PU141 is selective toward CBP and p300. The pan-inhibitor PU139 triggers caspase-independent cell death in cell culture. Both inhibitors block growth of SK-N-SH neuroblastoma xenografts in mice and the PU139 was shown to synergize with doxorubicin in vivo. The latter also reduces histone lysine acetylation in vivo at concentrations that block neoplastic xenograft growth. This is one of the very few reports on hypoacetylating agents with in vivo anticancer activity.

Increased leukocyte survival and accelerated onset of lymphoma in the absence of MCL-1 S159-phosphorylation.

The antiapoptotic BCL-2 protein MCL-1, which opposes mitochondrial outer membrane permeabilization, was shown to have a crucial role in the survival of hematopoietic cells. We have previously shown that, upon loss of phosphatidylinositol 3-kinase signaling, S159 of MCL-1 is phosphorylated by glycogen synthase kinase-3 (GSK-3), earmarking MCL-1 for enhanced ubiquitylation and degradation. In this study, we introduced MCL-1(wt) or the phosphorylation-deficient mutant MCL-1(S159A) in mouse BM cells, followed by adoptive transfer to recipient mice. Mice expressing MCL-1(S159A) exhibited significantly elevated white blood cell and lymphocyte counts, whereas no effect was observed on the distribution of T and B lymphocyte subsets or the numbers of monocytes, red blood cells or platelets. Expression of MCL-1(S159A) in Eµ-Myc transgenic bone marrow significantly accelerated the onset of disease, and these mice displayed increased spleen weights compared with Eµ-Myc/MCL-1(wt) mice. Our data demonstrate that the absence of MCL-1 S159 phosphorylation provides a survival advantage for hematopoietic cells in vivo and facilitates oncogenesis.

[Keratoconus lenses: the small correction miracle]. / Keratokonuslinse: Das kleine Korrektionswunder.

Keratoconus is a non-inflammatory corneal disease associated with a cone-shaped protrusion and progressive corneal thinning. Apart from progression control and stabilizing interventions, correcting the optical error induced by a mostly highly irregular corneal surface is of paramount importance with respect to quality of life and ability to work. This goal can be achieved efficiently by contact lenses with only rare adverse conditions. This article provides a current overview on contact lens fitting in keratoconus and presents own associated results.

Characterization of four endogenous viral genes in semi-congenic lines of meat chickens.

Restriction fragment length polymorphism analyses were used to examine endogenous viral genes (ev genes or ALVE genes) of the avian leukosis viral (ALV) family in semi-congenic lines of meat chickens. The Generation 6 lines examined in this study were semi-congenic in that each contained birds with either zero or with one ALVE gene in hemizygous state plus some solitary long terminal repeat (LTR) elements. Using four restriction enzymes on chicken genomic DNA and two probes, one representing the entire ALV retroviral genome and one with only a small part plus the LTR, four ALVE genes were characterized. Each seemed to be complete with no detectable deletions. None appeared to be similar to known ALVE genes of White Leghorns, whereas two of the four may be the same as ALVE genes reported by others in White Plymouth Rock chickens.

Characterization of eight endogenous viral (ev) genes of meat chickens in semi-congenic lines.

Restriction fragment length polymorphism analysis was conducted for a set of eight different meat chicken-derived endogenous viral genes (ev genes) of the avian leukosis viral (ALV) family. Each viral element was first isolated into a separate single-element line by selective breeding. Genomic DNA from the founder male for each semi-congenic, single-element line was digested with each of four restriction enzymes, and the resulting Southern blots were each hybridized with up to four probes representing different portions of the ALV retroviral genome. Among the eight elements, there was one that represents the broiler equivalent of locus ev3 of White Leghorn chickens. A second broiler element showed a SacI-specific junction fragment similar to that of ev8. The remainder appeared to be different from any of the 21 ev genes previously described for White Leghorn chickens. Four of the eight elements examined were essentially complete, but the rest have sustained internal deletions.

Estimates of relatedness and inbreeding in goose strains from DNA fingerprints.

The purpose of this study was to determine if DNA fingerprints (DFPs) could be used to estimate relatedness and inbreeding of strains of geese and to compare three methods of calculating relatedness indices. Strains included a control and selected strain from each of the Chinese and Synthetic (Chinese, Hungarian and Pilgrim) breeds. DFP patterns for each strain were based on individual DNA samples from six females, or on pooled DNA from 15 females different from those used for individual samples. Three relatedness indices were used, namely, genetic distance, modified Rogers distance and band sharing. All relatedness indices showed a closer relationship of strains within than between breeds. Correlation coefficients among relatedness indices were higher based on pooled DNA (r > or = magnitude of 0.97) than those based on individual DNA (r > or = magnitude of 0.741). Inbreeding estimates were higher for selected compared with control strains. It appears that the use of DFPs to estimate relatedness, regardless of index used, and inbreeding can be valuable for studying geese where there is a limited breeding history.

Effect of dietary contents of cholecalciferol, 1 alpha,25-dihydroxycholecalciferol and 24,25-dihydroxycholecalciferol on blood concentrations of 25-hydroxycholecalciferol, 1 alpha,25-dihydroxycholecalciferol, total calcium and eggshell quality.

1. Withdrawal of cholecalciferol (D3) supplement from a layers diet drastically reduced blood 25-hydroxycholecalciferol (25-OH-D3), 1 alpha,25-dihydroxycholecalciferol (calcitriol) and egg specific gravity (SG) within two weeks, followed by a decrease in blood total calcium (Ca). 2. Doubling the D3 supplement in the control diet (27.5 micrograms or 1100 IU/kg) almost linearly increased the circulating concentration of 25-OH-D3 without raising the concentration of calcitriol, Ca, or egg SG. 3. Replacing D3 by the optimal concentration of calcitriol (5 micrograms/kg diet) improved egg SG after 21 weeks of treatment without increasing blood calcitriol or total Ca. 4. By itself, 24,25-dihydroxycholecalciferol [24,25-(OH)2D3] was unable to maintain normal blood levels of calcitriol, Ca or egg SG and, when added together with calcitriol in the diet, tended to elevate blood Ca but suppress the beneficial effect of calcitriol on shell quality, with little or no effect on blood calcitriol.

Expression of an amylase--alcohol dehydrogenase chimeric gene in transgenic strains of Drosophila melanogaster.

A chimeric gene, consisting of 428 bp of the promoter sequences of the alpha-amylase gene of Drosophila melanogaster, fused to the transcribed region of the alcohol dehydrogenase (Adh) gene, was introduced into the genome of an Adhnull stock of Drosophila via P element mediated transformation. DNA analysis (Southern blotting) of three transformant strains confirmed the insertion of either one or two copies of the chimeric gene per strain. A histochemical study of ADH enzyme activity in dissected tissues of the transgenic larvae revealed that the chimeric Amy-Adh gene was expressed only in the posterior larval midgut and that this expression was repressed by dietary glucose, thus representing an expression pattern characteristic of the Amy gene. This indicates that the Amy upstream promoter sequences contain signals mediating both tissue specificity and glucose repression of the Adh structural gene in the transgenic larvae. The level of ADH activity expressed in transgenic flies was relatively low. This was paralleled by a low level of Adh mRNA, indicating a reduction in the transcriptional rate of the chimeric gene.

Research note: avian leukosis retroviral genes are not detected in geese.

Genomic DNA from four strains of geese was analyzed for the presence of endogenous viral elements using a probe that can detect over 20 Rous-associated endogenous viral genes (ev genes) in chickens, as well as a probe and protocol that detects endogenous avian viruses (EAV). Southern blot analysis of genomic DNA did not reveal any ev genes in DNA of 15 geese from Chinese, Synthetic, or two Embden goose strains. Even under low stringency conditions, using a probe that covered most of the polymerase (pol) gene of the Rous-associated virus (RAV) and that revealed EAV elements in a chicken without ev genes, no viral loci were evident in goose DNA.

Endogenous viral gene distribution in populations of meat-type chickens.

The present study was designed to document the complexity of endogenous viral (ev) genes and seek evidence for their association with production traits in selected and control strains of meat-type chickens. Three populations were studied, each consisting of a control strain and one to three strains selected for various production traits. The ev genes were revealed by digesting genomic DNA with restriction enzymes and detecting DNA fragments on Southern blots using radioactive probes from nucleotide sequences of the avian leukosis virus genome. A total of 31 polymorphic ev loci were identified in these populations from a SacI digest, with an average of 7.3 ev genes per bird. There were no significant differences in ev genes per bird between strains within populations or between selected and control strains overall. Thirty of 62 comparisons in the three populations indicated ev gene frequency differences (P less than .05). Within populations, 13 of 93 comparisons of ev gene frequencies between control and selected strains and 8 of 62 between three selected strains of a sire population showed such differences (P less than .05). Selection for body weight and feed efficiency had been observed to reduce gene frequencies of the slow-feathering gene, which usually contains the ev21 locus; however, these effects were not detected (.05 less than P less than .06) between strains of the dam population in the current study. Such differences suggested possible associations between ev genes and production traits in meat-type chickens.

Comparison of procedures for collecting semen from ganders and inseminating geese.

Experiments were conducted to compare management of ganders and semen collection procedures with respect to semen and sperm yield, and two frequencies of artificial insemination were tested with respect to fertility. Housing ganders in groups, singly, or singly with the introduction of a female just before collection was the rank order of housing system from least to most successful collection of ejaculates. There were no significant differences among types of housing with respect to semen volume, but ejaculates from ganders housed singly had the greatest (P less than .05) spermatozoal concentrations. Ejaculates collected with an artificial vagina were of greater (P less than .05) volume and total spermatozoal yield but not spermatozoal concentration than those collected by aspiration. Interactions between collector and method were observed for spermatozoal traits. Geese inseminated on 2 consecutive days/wk showed greater (P less than .01) fertility than those inseminated once per week. Therefore, collection of semen with an artificial vagina from ganders housed singly, with insemination weekly but on consecutive days, should result in successful reproduction of geese.

Optimal dietary level of 1 alpha,25-dihydroxycholecalciferol for eggshell quality in laying hens.

The optimal dietary level of 1 alpha,25-dihydroxycholecalciferol [1,25-(OH)2D3] for eggshell quality was established. White Leghorn hens, 59 wk of age, were fed one of eight diets that contained the same basal ingredients, including 3.1% calcium, but different levels (microgram/kg) or forms of calciferol supplements: no calciferol supplement of any form (56 hens); 27.5 (control) or 55.0 micrograms of cholecalciferol (56 hens each); 3, 5, or 7 micrograms of 1,25-(OH)2D3 (28 hens each); 5 micrograms of 24,25-dihydroxycholecalciferol [24,25-(OH)2D3] with 28 hens; 5 micrograms each of 1,25-(OH)2D3 and 24,25-(OH)2D3 (28 hens). All groups were fed the control diet prior to the 21-wk treatment. The group fed 5 micrograms 1,25-(OH)2D3/kg diet ranked first in specific gravity (SG), e.g., 1.081 versus 1.077 for the control group at Week 21 (P less than .05). The group fed 7 micrograms 1,25-(OH)2D3/kg consumed 30% less feed and laid 20% fewer eggs than the control, but shell quality was not affected. The groups receiving no calciferol supplement or receiving only 24,25-(OH)2D3 laid eggs with significantly lower SG than the control after 2 wk of treatment (1.072 or less versus 1.082 at Week 2). The rest of the treatment groups mentioned were comparable to the control in eggshell quality and egg production. Groups fed the combination of 1,25-(OH)2D3 and 24,25-(OH)2D3 per kilogram of feed, or 1,25-(OH)2D3 alone at 5 micrograms/kg, had significantly higher tibial weights relative to the control group. All groups receiving the diets without cholecalciferol supplementation had markedly reduced hatchability. It was concluded that the optimal dietary level of 1,25-(OH)2D3 for improving eggshell quality without affecting egg production was approximately 5 micrograms/kg and the toxic level was 7 micrograms/kg.

Effects of vitamin D3 metabolites on physiological traits of White Leghorn hens.

This study was conducted to determine the effect of feeding vitamin D3(D3) metabolites on BW of hen, weight of uterus, plasma Ca, jejunal and uterine adenosine triphosphatase (ATPase), and carbonic anhydrase. At 416 days of age each of 7 groups of laying hens was fed the basal ration supplemented with one of 7 concentrations (micrograms per kg) of D3 or its metabolites as treatments: 0 micrograms of D3; 27.5 micrograms of D3; 3, 5, or 7 micrograms of 1,25(OH)2D3; 5 micrograms of 24,25(OH)2D3; and 5 micrograms of 24,25(OH)2D3 plus 5 micrograms of 1,25(OH)2D3. Treatment effects were compared at various periods after the start of the study. Hens fed the unsupplemented ration had lower (P less than .05) values for all traits than hens fed the D3-supplemented ration by 162 days after the start of treatment. In a comparison of all dietary treatments except the one involving 0 micrograms D3, from 154 to 161 days after the start of the experiment, treatment effects were significant (P less than or equal to .05) for BW, uterine ATPase, and carbonic anhydrase; hens fed 5 micrograms of 24,25(OH)2D3 per kg of ration ranked the lowest of all treatment groups for these traits. Hens fed 27.5 micrograms of D3 and those fed 5 micrograms of 1,25(OH)2D3 per kg of ration did not differ (P greater than .05) for any traits studied. The results suggest that 5 micrograms of 1,25(OH)2D3 per kg of ration can replace 27.5 micrograms of D3 per kg of ration but that 5 micrograms of 24,25(OH)2D3 per kg of ration tends to have a negative effect on physiological systems of the hen.

Effect of 1 alpha,25-dihydroxycholecalciferol on egg shell quality and egg production.

1. The effect of replacing dietary cholecalciferol (D3) by 1 alpha,25-dihydroxycholecalciferol (1,25-(OH)2D3) on egg shell quality and egg production was tested on 32-week-old White Leghorn laying hens over 9 weeks. 2. Hens fed on a diet supplemented with 5 micrograms 1,25-(OH)2D3/kg diet, tended to lay more eggs, and the eggs had significantly higher specific gravity and percentage shell than eggs from control hens fed on a diet supplemented with 27.5 micrograms D3/kg diet. 3. The effect became apparent after about 4 weeks of treatment and persisted until the end of the test. 4. Hens fed on a diet without D3 supplement started to lay very thin or soft shelled eggs within 4 weeks, suggesting that the birds' reserves of D3 or its metabolites were depleted within this period. 5. The results suggest that 1,25-(OH)2D3 can be substituted for D3 in layer diets to improve egg shell quality.

Effects of cholecalciferol or calcium deficiency on oestrogen metabolism in the laying hen.

1. Seventeen 32-week-old White Leghorn laying hens were induced to become deficient in calciferol or in calcium (laying thin or soft shelled eggs) by withdrawing either cholecalciferol (27.5 micrograms/kg diet) or calcium (31 g/kg diet) supplements from the control diet. 2. The metabolic fate and metabolic clearance rate (MCR) of intravenously injected 3H-oestradiol-17 beta were then monitored for 40 min. 3. In both the calciferol and calcium-deficient groups a major oestrogen sulphate pathway was particularly affected, resulting in a decreased conversion of oestradiol-17 beta-3-sulphate to oestradiol-17 alpha-3-sulphate, with a concomitant reduced MCR of oestradiol-17 beta from plasma. 4. The metabolic defect was corrected by feeding the control diet. 5. Because the metabolic defect observed in calciferol deficiency occurred in Ca deficiency in a more severe form, we conclude that the more immediate cause was calcium rather than calciferol deficiency. To our knowledge, this is the first observation of a calcium-deficient effect on oestrogen sulphate metabolism in vivo.

Genetic parameters of plasma very low density lipoproteins, abdominal fat lipase, and protein, fatness, and growth traits of broiler chickens.

Genetic parameters of physiological, growth, and fatness traits were investigated in one control and two selected dam strains of broiler chickens. Feed consumption and efficiency were measured between 28 and 42 days of age but were adjusted to estimate values of population average body weights at these ages. Birds were bled at 45 days of age for assay of plasma very low density lipoproteins (VLDL) and killed at 47 days of age for carcass and fat measurements. Abdominal fat was assayed for lipase activity expressed per milligram of protein (LIP/mgP) or per gram of fat (LIP/gF) and protein content of the enzyme preparation expressed as microgram protein/mL (P/mLEPrep). Absolute values of partial correlations corrected for sex and strain were low between production and physiological traits and between fatness and LIP/mgP but were moderate at .3 between fatness and plasma VLDL, LIP/gF, and P/mLEPrep. Heritabilities were moderate to high (greater than or equal to .32) for growth and fatness traits, moderate (.25) for plasma VLDL, and low (less than or equal to .16) for P/LEPrep, LIP/mgP, and LIP/gF. Genetic correlations involving plasma VLDL were as follows: .49 with body weight at 42 days, -.74 with feed consumption, .64 with feed efficiency, .24 with carcass weight at 47 days, 1.07 with abdominal fat weight, and .97 with abdominal fat percentage. Similarly, absolute values of genetic correlations involving P/mLEPrep tended to be as high or higher but genetic correlations involving LIP/mgP and LIP/gF tended to be lower than those involving plasma VLDL.(ABSTRACT TRUNCATED AT 250 WORDS)

Plasma very low density lipoproteins, abdominal fat lipase, and fatness during rearing in two strains of broiler chickens.

Experimental control Strains 30, representative of commercial broiler dam stocks in the late 1970's, and K, representative of commercial broiler stocks of 20 years earlier, were compared. Development of carcass fatness and related traits and physiological traits from 3 to 17 weeks of age were studied. About 12 chickens of mixed sex of each strain were bled and killed at 2-week intervals beginning at 3 weeks of age for measurement of traits. From 3 to 17 weeks of age, percentage abdominal fat of carcass, carcass fat (ether extract) and plasma very low density lipoproteins (VLDL) increased with age. Percentage carcass nitrogen, ash, and water, lipase activity of abdominal fat expressed per mg protein or per g of fat (LIP/g F), and protein concentration of the abdominal fat enzyme preparation (P/ml EPrep) decreased with age. Strain 30 had a higher percentage of abdominal fat and carcass fat, but less carcass nitrogen, ash, and water than Strain K, although there were interactions with sex and age. For physiological traits, Strain 30 had more LIP/g F and less P/ml EPrep than Strain K. Males had lower percentages of abdominal fat, carcass fat, and plasma VLDL, and higher percentages of carcass nitrogen, ash, and water than females. There were no significant differences between sexes for the other physiological traits. There were significant (P less than .01) partial correlations between plasma VLDL and percentage abdominal fat (.22), and between P/ml EPrep and percentages of abdominal fat (-.25), carcass fat (-.29), carcass nitrogen (.30), and carcass water (.30). These observations in conjunction with multiple regression analyses indicated that, in addition to plasma VLDL, protein concentration of adipose tissue expressed as P/ml EPrep might be a useful predictor of fatness in chickens.