Pre-eclampsia screening in Denmark (PRESIDE): national validation study.

OBJECTIVES: To investigate the predictive performance of the Fetal Medicine Foundation (FMF) first-trimester screening algorithm for pre-eclampsia in a Danish population and compare screening performance with that of the current Danish strategy, which is based on maternal risk factors. METHODS: This was a prospective study of women with a singleton pregnancy attending for their first-trimester ultrasound scan and screening for aneuploidies at six Danish university hospitals between May 2019 and December 2020. Prenatal data on maternal characteristics and medical history were recorded, and measurements of mean arterial pressure (MAP), uterine artery pulsatility index (UtA-PI), serum pregnancy-associated plasma protein-A (PAPP-A) and serum placental growth factor (PlGF) were collected without performing a risk assessment for pre-eclampsia. Information on acetylsalicylic acid use was recorded. After delivery, pregnancy outcome, including gestational age at delivery and pre-eclampsia diagnosis, was recorded. Pre-eclampsia risk assessment for each woman was calculated blinded to outcome using the FMF screening algorithm following adjustment to the Danish population. Detection rates (DRs) of the FMF algorithm were calculated for a fixed screen-positive rate (SPR) of 10% and for the SPR achieved in the current Danish screening. RESULTS: A total of 8783 pregnant women were included, with a median age of 30.8 (interquartile range (IQR), 28.1-33.9) years. The majority were white (95%), naturally conceiving (90%), non-smokers (97%) and had no family history of pre-eclampsia (96%). The median body mass index was 23.4 (IQR, 21.2-26.6) kg/m2 . A complete risk assessment including maternal characteristics, MAP, UtA-PI, PlGF and PAPP-A was available for 8156 women (92.9%). In these women, UtA-PI was measured bilaterally with a median value of 1.58 (IQR, 1.27-1.94) and the median resting MAP of 80.5 (IQR, 76.1-85.4) mmHg in two consecutive measurements. Among these, 303 (3.7%) developed pre-eclampsia, including 55 (0.7%) cases of pre-eclampsia with delivery < 37 weeks of gestation and 16 (0.2%) cases of pre-eclampsia with delivery < 34 weeks. At a SPR of 10%, combined screening using the FMF algorithm based on maternal characteristics, MAP, UtA-PI, PlGF and PAPP-A had a DR of 77.4% (95% CI, 57.6-97.2%) for pre-eclampsia with delivery < 34 weeks, 66.8% (95% CI, 54.4-79.1%) for pre-eclampsia with delivery < 37 weeks and 44.1% (95% CI, 38.5-49.7%) for pre-eclampsia with delivery at any gestational age. The current Danish screening strategy using maternal risk factors detected 25.0% of women with pre-eclampsia with delivery < 34 weeks and 19.6% of women with pre-eclampsia with delivery < 37 weeks at a SPR of 3.4%. When applying the FMF algorithm including maternal characteristics, MAP, UtA-PI and PlGF at the fixed SPR of 3.4%, the DRs were 60.5% (95% CI, 36.9-84.1%) for PE with delivery < 34 weeks and 45.2% (95% CI, 32.0-58.5%) for PE with delivery < 37 weeks. CONCLUSION: In this large Danish multicenter study, the FMF algorithm based on maternal characteristics, MAP, UtA-PI, PlGF and PAPP-A predicted 77.4% of cases with pre-eclampsia with delivery < 34 weeks and 66.8% of cases with pre-eclampsia with delivery < 37 weeks of gestation at a SPR of 10%, suggesting that the performance of the algorithm in a Danish cohort matches that in other populations. © 2023 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.

Description and evaluation of a net energy intake model as a function of dietary chewing index.

Previously, a linear relationship has been found between net energy intake (NEI) and dietary chewing index (CI) of the diet for different types of cattle. Therefore, we propose to generalize and calibrate this relationship into a new model for direct prediction of NEI by dairy cows from CI values (CINE; min/MJ of NE). Furthermore, we studied the forage-to-concentrate substitution rate in this new NEI model. To calibrate the model on a diverse set of situations, we built a database of mean intake from 14 production experiments with a total of 986 primi- and multiparous lactating dairy cows of different breeds fed 136 different diets ad libitum. The NEI were estimated by the Nordic feed evaluation system. The CINE value of diets was estimated from the intake of concentrate, intake of forage neutral detergent fiber (NDFf), particle length of forage, indigestible NDFf/NDFf, body weight, NDFf/body weight, and the content of NE in DM. We show that the slope values in this regression are proportional to the squared intercepts, giving the nonlinear equation NEI=NEI0-k×NEI0a×CINE, where the parameter k represents the decline in NEI with the increasing CINE of the diet and a was estimated to have a value of 2, implying a constant maximum daily chewing time. The intercept NEI0 in the regression of NEI on CINE may be interpreted as metabolic net energy intake capacity of the cows fed without physical constraints on intake. Based on experimental data, the maximum chewing time was estimated as 1/(4 × k). The NEI0 values were parameterized as a linear function of metabolic body size, energy-corrected milk yield (kg/d), days in milk, and days in milk squared. Prediction accuracy was evaluated by mean square prediction error (MSPE) and its decomposition into central tendency, regression, and disturbance, across and within experiments on independent data from 19 experiments including 812 primi- and multiparous lactating dairy cows of different breeds fed 80 different diets ad libitum. The NEI model predicted NEI with an MSPE of 8% of observed, and across the 19 experiments the error central tendency, error regression, and error disturbance were 4.2, 40.6, and 84.9% of MSPE, respectively. The described intake model implies a variable forage-to-concentrate substitution rate as a nonlinear function of NEI0, CINE of forage, and supplementation of concentrate.

Variations in automatically recorded rumination time as explained by variations in intake of dietary fractions and milk production, and between-cow variation.

Individual recording of rumination time (RT) is now possible in commercial dairy herds, through development of a microphone-based sensor, which is able to record RT by the sound of rumination activity. The objectives of this study were to examine the relationship between daily RT and intakes of different dietary fractions, the relationship between RT in minutes per kilogram of dry matter intake (DMI) and milk production, and to examine the variation in RT within and between mid-lactating dairy cows. Data from 3 production trials were used in which a total of 27 different diets were fed. The data contained 761, 290, and 203 daily recordings of RT, milk yield, milk components, DMI, and intake of dietary fractions recorded on 29, 26, and 24 Holstein and Swedish Red cows from trials 1, 2, and 3, respectively. The dietary fractions included forage neutral detergent fiber (NDF), concentrate NDF, crude protein, sugar, starch, and the remaining fraction represented by organic matter--(forage NDF+concentrate NDF+crude protein+sugar+starch). The relationship between the dietary fractions and RT was analyzed in 2 steps. In step 1, the dietary fractions, which were significantly related to RT, were selected and simultaneously checked for multicollinearity between the dietary components; in step 2, a multivariate model, including the effect of repeated measurements, the main effect of the selected dietary fractions from step 1, random effects of cow(trial) and trial, and information on breed, days in milk, and parity was used to analyze the relationship between RT and the selected dietary fractions. Relationships between RT in minutes per kilogram of DMI and milk yield and milk components were analyzed, using the same multivariate model as in step 2. Approximately 32% of the variation in daily RT could be explained by variations in intakes of the dietary fractions, whereas 48% of the total variation in RT was accounted for by individual variations between cows. Intakes of forage NDF and starch were positively related to daily RT, whereas intakes of sugar and the remaining fraction were negatively related to daily RT. Rumination time in minutes per kilogram of DMI was negatively related to milk yield and protein percentage, but positively related to milk fat percentage.

Temporal effects of maternal and pregnancy characteristics on serum pregnancy-associated plasma protein-A and free ß-human chorionic gonadotropin at 7-14 weeks' gestation.

OBJECTIVE: The aim of this study was to investigate gestational age-dependent effects of racial origin, smoking status and mode of conception on maternal serum levels of free ß-human chorionic gonadotropin (ß-hCG) and pregnancy-associated plasma protein-A (PAPP-A) at 7-14 weeks' gestation. METHODS: This was an analysis of data from prospective first-trimester combined screening for aneuploidies in singleton pregnancies, with ß-hCG and PAPP-A measured at 7 + 1 to 14 + 3 weeks' gestation. We included 27,908 pregnancies from three centers in the U.K. and 125,461 pregnancies from 22 centers in Denmark, all with known normal fetal karyotype or resulting in the birth of a phenotypically normal neonate. Multiple regression modelling of log10 -transformed marker concentrations was used to produce log10 multiple of the median (MoM) values for free ß-hCG and PAPP-A and to examine pregnancy characteristics that have significant effects on marker concentrations. RESULTS: Serum free ß-hCG and PAPP-A concentrations were significantly affected by gestational age, maternal weight, racial origin, parity, smoking and mode of conception. There were significant gestational age-dependent effects attributed to Afro-Caribbean race, smoking and conception through in-vitro fertilization (IVF) on PAPP-A and free ß-hCG levels. In women of Afro-Caribbean race there was a weekly increase in PAPP-A of 5.3% and in free ß-hCG of 1.8%. In smokers there was a weekly decrease in PAPP-A of 2.4% and in free ß-hCG of 1.6%. In cases of IVF conceptions there was a weekly increase in PAPP-A of 4.5% and in free ß-hCG of 4.6%. CONCLUSIONS: Serum free ß-hCG and PAPP-A concentrations at 7-14 weeks' gestation are affected by several pregnancy characteristics. The effects of Afro-Caribbean race, smoking and IVF conception change with gestational age.

Particle size alterations of feedstuffs during in situ neutral detergent fiber incubation.

Particle size alterations during neutral detergent fiber (NDF) determination and in situ rumen incubation were analyzed by dry sieving and image analysis to evaluate the in situ procedure for estimation of NDF degradation parameters and indigestible NDF concentration in terms of particle size. Early-cut and late-cut grass silages, corn silage, alfalfa silage, rapeseed meal, and dried distillers grains were examined. Treatments were (1) drying and grinding of forage samples and grinding of concentrates; (2) neutral detergent-soluble (NDS) extraction; (3) machine washing and NDS extraction; (4) 24-h rumen incubation, machine washing, and NDS extraction; and (5) 288-h rumen incubation, machine washing, and NDS extraction. Degradation profiles for potentially degradable NDF were determined and image analysis was used to estimate particle size profiles and thereby the risk for particle loss. Particle dimensions changed during NDF determination and in situ rumen incubation and variations depended on feedstuff and treatment. Corn silage and late-cut grass silage varied most in particle area among feedstuffs, with an increase of 139% between 0 and 24h and a decrease of 77% between 24 and 288 h for corn silage and a decrease of 74% for late-cut grass silage between 24- and 288-h in situ rumen incubation. Especially for late-cut grass silage residues after 288 h in situ rumen incubation, a high mass proportion in the critical zone for escape was found. Particle area decreased linearly with increasing incubation time. Particle loss during in situ rumen incubation cannot be excluded and is likely to vary among feedstuffs.

Prospective study evaluating performance of first-trimester combined screening for trisomy 21 using repeat sampling of maternal serum markers PAPP-A and free ß-hCG.

OBJECTIVE: To prospectively evaluate the performance of first-trimester combined screening for trisomy 21 using the biochemical markers pregnancy-associated plasma protein-A (PAPP-A) and free beta-human chorionic gonadotropin (free ß-hCG) obtained before and at the time of the nuchal translucency (NT) scan. METHODS: Three fetal medicine departments in Denmark participated in the study. Screening for trisomy 21 was set up as a two-step approach with blood sampling performed before the NT scan (early sample) and again at the time of the NT scan (late sample). PAPP-A and free ß-hCG were measured on both the early and late samples. Age-standardized detection and false-positive rates for different screening protocols were calculated. RESULTS: We collected two blood samples in 27 pregnancies affected by trisomy 21 and in 3891 control pregnancies. The early samples were taken between gestational ages 8 + 0 and 13 + 6 weeks, and the late samples between 11 + 3 and 14 + 6 weeks. The median interval between the samples was 17 (range, 1-40) days. We found a significantly better estimated screening performance when using early sampling vs late sampling (P < 0.05). With a risk cut-off of 1 in 100, at the time of the risk assessment the estimated detection and false-positive rates when using the early sample were 91% (95% CI, 81-98%) and 1.6% (95% CI, 1.3-2.0%), respectively. For fixed false-positive rates the highest detection rates were achieved using both blood samples. When comparing early sampling vs double sampling there was no significant difference in screening performance. CONCLUSION: In combined first-trimester screening for trisomy 21, use of early sampling with measurement of PAPP-A and free ß-hCG before the time of the NT scan can optimize screening performance. Using maternal serum markers obtained both before and at the time of the NT scan has the potential to further improve performance, but larger studies are needed to confirm this potential.

Early lactation feed intake and milk yield responses of dairy cows offered grass silages harvested at early maturity stages.

The main objective was to evaluate the potential of grass silages of very high quality to support a high milk yield with a low or moderate, or even without concentrate supplementation. Production responses to increased levels of concentrate supplementation with 3 primary growth grass silages differing in digestibility were studied using 66 Norwegian Red dairy cows. Roundbale silage was produced from a timothy-dominated sward at very early (H1), early (H2), and normal (H3) stages of crop maturity. Crops were rapidly wilted (<24h) and a formic acid-based additive was applied. All silages were restrictedly fermented. Silage digestible organic matter in dry matter (DM) values were 747, 708, and 647 g/kg of DM for H1, H2, and H3, respectively. Dietary treatments were fed in a 3×3 factorial arrangement of the 3 silages supplemented with 3 concentrate levels (4, 8, and 12 kg/d) and, additionally, H1 was offered without concentrates and H3 with 16 kg/d, giving a total of 11 diets. Cows, blocked according to parity and calving date, were introduced to the experiment before calving and kept in the experiment until wk 16 of lactation. Silage was offered ad libitum in loose housing and concentrate was available in automatic feed stations. Intake of grass silage when fed as the sole feed was 16.9 kg of DM on average for lactation wk 1 to 16. When H1 was supplemented with 4 or 8 kg of concentrates, silage DM intake did not change, but total DM intake increased to 20.6 and 23.7 kg/d, respectively. Energy-corrected milk (ECM) yield increased from 23.4 kg when H1 was offered without concentrate supplement to 29.1 and 32.8 kg when supplemented with 4 or 8 kg concentrate, respectively. None of the other diets equaled the yield obtained by H1 plus 8 kg of concentrate. Feed intake and yield of cows offered H3 plus 4 kg of concentrates were strongly constrained by high dietary fiber concentration. They consumed 16.5 g of neutral detergent fiber/kg of body weight and spent more time eating silage than cows offered other diets. The highest concentrate level within each silage quality produced similar or lower ECM yield than that with 4 kg less concentrates. The obtained milk yield responses suggest that provision of 8.0, 8.4, and 11.5 kg of concentrates to H1, H2, and H3, respectively, would maximize ECM yield within each silage type. However, H1 may successfully be used with less concentrates, or even without, if future conditions should limit the amount of concentrates available for ruminant production.

Skin temperature measured by infrared thermography after specific ultrasound-guided blocking of the musculocutaneous, radial, ulnar, and median nerves in the upper extremity.

BACKGROUND: Sympathetic block causes vasodilatation and increases in skin temperature (T(s)). However, the T(s) response after specific nerve blocking is unknown. In this study, we hypothesized that T(s) would increase after specific blocking of the nerve innervating that area. METHODS: Forty-six patients undergoing hand surgery were included. We performed ultrasound-guided, specific nerve blocking of either the musculocutaneous, radial, ulnar, or median nerve in each patient and analysed T(s) in the forearm and hand at 2 min intervals in the following 22 min by the use of infrared thermography. Areas of interest corresponding to the cutaneous innervation area of each of the four nerves were defined and the mean T(s) in each area was analysed. RESULTS: Specific blocking of the ulnar and median nerves caused a substantial increase in mean (sd) T(s) in the areas innervated by these nerves [5.2 (3.2)°C and 5.1 (2.5)°C, respectively; both P<0.0001]. The increase was even larger at the fingertips. Median nerve blocking also increased T(s) in the area of the hand innervated by the radial nerve (P<0.0001). However, T(s) did not increase in any area after either musculocutaneous or radial nerve blocking. CONCLUSIONS: Specific blocking of the ulnar and median nerve causes substantial increases in T(s) in specific areas of the hand. In contrast, the specific blocking of the musculocutaneous or radial nerve does not increase T(s). Further studies are needed to clarify if these findings can be used to objectively evaluate brachial plexus block success.

Functional differences in yeast protein disulfide isomerases.

PDI1 is the essential gene encoding protein disulfide isomerase in yeast. The Saccharomyces cerevisiae genome, however, contains four other nonessential genes with homology to PDI1: MPD1, MPD2, EUG1, and EPS1. We have investigated the effects of simultaneous deletions of these genes. In several cases, we found that the ability of the PDI1 homologues to restore viability to a pdi1-deleted strain when overexpressed was dependent on the presence of low endogenous levels of one or more of the other homologues. This shows that the homologues are not functionally interchangeable. In fact, Mpd1p was the only homologue capable of carrying out all the essential functions of Pdi1p. Furthermore, the presence of endogenous homologues with a CXXC motif in the thioredoxin-like domain is required for suppression of a pdi1 deletion by EUG1 (which contains two CXXS active site motifs). This underlines the essentiality of protein disulfide isomerase-catalyzed oxidation. Most mutant combinations show defects in carboxypeptidase Y folding as well as in glycan modification. There are, however, no significant effects on ER-associated protein degradation in the various protein disulfide isomerase-deleted strains.

Short communication: salivary secretion during meals in lactating dairy cattle.

Four multiparous Holstein cows in midlactation were used in a 4 x 4 Latin square to evaluate whether source of forage influenced salivary secretion during eating in lactating dairy cows. The forages were allocated separately from the pelleted concentrates. Cows were offered 1 of 4 forages each period: barley silage, alfalfa silage, long-stemmed alfalfa hay, or chopped barley straw. Saliva secretion was measured during the morning meal by collecting masticates through the rumen cannula at the cardia of each cow. Rate of salivation (213 g/min) was not affected by forage source. However, the forage sources differed in eating rate (g of DM/min), which led to differences in ensalivation of forages (g of saliva/g of DM and g of saliva/g of NDF). On the basis of DM, ensalivation (g of saliva/g of DM) was greatest for straw (7.23) and similar for barley silage, alfalfa silage, and alfalfa hay (4.15, 3.40, and 4.34 g/g of DM, respectively). Higher ensalivation of straw could be accounted for by its higher neutral detergent fiber (NDF) content; ensalivation of NDF (g of saliva/g of NDF) was actually greatest for long-stemmed alfalfa hay (12.4) and similar for the other chopped forages (8.9). Cows consumed concentrate about 3 to 12 times faster than the various forages (DM basis), and ensalivation of concentrate was much lower (1.12 g of saliva/g of DM) than for forages. Feed characteristics such as particle size, DM, and NDF content affect salivary output during eating by affecting the eating rate. Slower eating rate and greater time spent eating may help prevent ruminal acidosis by increasing the total daily salivary secretion in dairy cows.

Effect of feed type and essential oil product on equine chewing activity.

The ingestive and post-digestion effect of a blend of special essential oil compounds (EO) on eating, chewing and faecal parameters were measured in horses. Ingestive effects appear after no adaptation. Post-digestion effects appear after adaptation. Six Icelandic horses were assigned to two groups in a Latin Square subplot design with EO treatments to four different roughage types and four different concentrates. The horses were fed four different roughage meals and two different concentrate meals on each of the four sampling days. Eating time and saliva were observed during meals. Jaw movements (JM) were recorded using a special chewing halter. Eating time was derived from JM and related to DM intake. The size characteristics of faecal particles were measured by using image analysis. All chewing characteristics measured were significantly affected by roughage (p < 0.001) and concentrate type (p < 0.01). EO had a significant ingestive effect on the frequency of observed saliva during concentrate meals. No significant (p < 0.05) post-digestive or ingestive effect of EO was found for any measured chewing characteristic, which was reflected in the absence of effect on faecal particle dimensions. In conclusion, effect of type of roughage and concentrate was more significant than potential effects of EO.

Presence of benzophenones commonly used as UV filters and absorbers in paired maternal and fetal samples.

BACKGROUND: Previous studies have demonstrated widespread exposure of humans to certain benzophenones commonly used as UV filters or UV absorbers; some of which have been demonstrated to have endocrine disrupting abilities. OBJECTIVES: To examine whether benzophenones present in pregnant women pass through the placental barrier to amniotic fluid and further to the fetal blood circulation. METHODS: A prospective study of 200 pregnant women with simultaneously collected paired samples of amniotic fluid and maternal serum and urine. In addition, unique samples of human fetal blood (n=4) obtained during cordocentesis: and cord blood (n=23) obtained at delivery, both with paired maternal samples of serum and urine collected simultaneously, were used. All biological samples were analyzed by TurboFlow-liquid chromatography - tandem mass spectrometry for seven different benzophenones. RESULTS: Benzophenone-1 (BP-1), benzophenone-3 (BP-3), 4-methyl-benzophenone (4-MBP), and 4-hydroxy-benzophenone (4-HBP) were all detectable in amniotic fluid and cord blood samples and except 4-HBP also in fetal blood; albeit at a low frequency. BP-1 and BP-3 were measured at ~10-times lower concentrations in fetal and cord blood compared to maternal serum and 1000-times lower concentration compared to maternal urine levels. Therefore BP-1 and BP-3 were only detectable in the fetal circulation in cases of high maternal exposure indicating some protection by the placental barrier. 4-MBP seems to pass into fetal and cord blood more freely with a median 1:3 ratio between cord blood and maternal serum levels. Only for BP-3, which the women seemed to be most exposed to, did the measured concentrations in maternal urine and serum correlate to concentrations measured in amniotic fluid. Thus, for BP-3, but not for the other tested benzophenones, maternal urinary levels seem to be a valid proxy for fetal exposure. CONCLUSIONS: Detectable levels of several of the investigated benzophenones in human amniotic fluid as well as in fetal and cord blood calls for further investigations of the toxicokinetic and potential endocrine disrupting properties of these compounds in order for better assessment of the risk to the developing fetus.

Maternal exposure to UV filters: associations with maternal thyroid hormones, IGF-I/IGFBP3 and birth outcomes.

BACKGROUND: Several chemical UV filters/absorbers ('UV filters' hereafter) have endocrine-disrupting properties in vitro and in vivo. Exposure to these chemicals, especially during prenatal development, is of concern. OBJECTIVES: To examine maternal exposure to UV filters, associations with maternal thyroid hormone, with growth factor concentrations as well as to birth outcomes. METHODS: Prospective study of 183 pregnant women with 2nd trimester serum and urine samples available. Maternal concentrations of the chemical UV filters benzophenone-1 (BP-1) and benzophenone-3 (BP-3) in urine and 4-hydroxy-benzophenone (4-HBP) in serum were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The relationships between 2nd trimester maternal concentrations of the three chemical UV filters and maternal serum concentrations of thyroid hormones and growth factors, as well as birth outcomes (weight, height, and head and abdominal circumferences) were examined. RESULTS: Positive associations between maternal serum concentrations of 4-HBP and triiodothyronine (T3), thyroxine (T4), insulin-like growth factor I (IGF-I) and its binding protein IGFBP3 were observed in mothers carrying male fetuses. Male infants of mothers in the middle 4-HBP exposure group had statistically significantly lower weight and shorter head and abdominal circumferences at birth compared to the low exposure group. CONCLUSIONS: Widespread exposure of pregnant women to chemical UV filters and the possible impact on maternal thyroid hormones and growth factors, and on fetal growth, calls for further studies on possible long-term consequences of the exposure to UV filters on fetal development and children's health.

Visceral leishmaniasis diagnosed in a patient with MALT lymphoma.

We report a case of visceral leishmaniasis in a 66-year-old female with a history of MALT lymphoma in the gastrointestinal tract. The patient presented with major hemorrhage per rectum and perforation of the small intestine. Due to unexplained decreasing platelets, lymphoma bone marrow involvement was suspected and bone marrow examination was performed. Surprisingly, Leishman-Donovan bodies were detected. The low platelet count, caused by the combination of MALT lymphoma and visceral leishmaniasis, appears to have aggravated the symptoms of the intestinal lymphoma. Leishmaniasis should be suspected even among asymptomatic patients with immune compromising illnesses and a travel history to areas where leishmaniasis is endemic.

Tetrahymena gene encodes a protein that is homologous with the liver-specific F-antigen and associated with membranes of the Golgi apparatus and transport vesicles.

The F-antigen is a prominent liver protein which has been extensively used in studies on natural and induced immunological tolerance. However, its intracellular localization and biological function have remained elusive. It has generally been assumed that the F-antigen is confined phylogenetically to vertebrates. Now we have cloned and characterized a gene from the ciliated protozoan Tetrahymena thermophila encoding a protein which clearly is homologous with the rat F-antigen. The coding region of the Tetrahymena F-antigen (TF-ag) gene specifies a 46,051 M(r) protein and is interrupted by three introns. In accordance with the predicted molecular mass of the TF-ag protein, antibodies raised against a cro-lacZ'-TF-ag fusion protein specifically recognized a 45,000 M(r) protein in Western blots of total T. thermophila protein. Immunoelectron microscopy demonstrated that the TF-ag is associated with membranes of the Golgi apparatus and transport vesicles pointing to a role of TF-ag in membrane trafficking. Transcription of the TF-ag gene, as determined by run-on analyses, was only detectable in growing cells, and following transfer to starvation condition pre-existing TF-ag mRNA was rapidly degraded. The abundance of the TF-ag protein, however, declined only moderately during prolonged periods of starvation demonstrating that extensive release of the TF-ag did not take place. In combination these results suggest that the TF-ag protein is a recycled constituent of the intracellular membrane network in T. thermophila.

Treatment with farnesyl-protein transferase inhibitor induces regression of mammary tumors in transforming growth factor (TGF) alpha and TGF alpha/neu transgenic mice by inhibition of mitogenic activity and induction of apoptosis.

Mouse mammary tumor virus-transforming growth factor alpha (MMTV-TGF alpha) and MMTV-TGF alpha/neu transgenic mice develop mammary tumors after a long latency and therefore provide useful model systems for breast cancer with its recognized activation of receptor tyrosine kinase signaling. We used these mice to study the antitumor effect of L-744,832 (FTI), a potent and selective inhibitor of farnesyl-protein transferase, and hence of Ras function. A total of 55 mice were assigned randomly to treatment with FTI or vehicle, and one-half of the mice were crossed over after initial treatment to the opposite group. L-744,832 induced reversible regression of mammary tumors that was paralleled by a decrease in serum levels of TGF alpha secreted by the tumor cells. There was no difference in response to treatment with FTI between MMTV-TGF alpha mice, in which tumorigenesis was accelerated by multiparity or the chemical carcinogen 7,12-dimethylbenzanthracene, and MMTV-TGF alpha/neu mice. The tumor histological type had no impact on FTI sensitivity. For mechanistic analyses, tumor excision biopsies were obtained from 12 mice before and after treatment with L-744,832. In these samples, tumor regression was paralleled biochemically by inhibition of mitogen-activated protein kinase activity and biologically by an increase in G1-phase and decrease in S-phase fractions, as well as induction of apoptosis. These results suggest that the potential clinical use of FTI could be expanded to include cancers harboring activated receptor tyrosine kinases as well as those containing activated Ras.

Evaluation of physical structure value in spring-harvested grass/clover silage and hay fed to heifers.

The physical structure value of conserved grass/clover forages of spring harvest was evaluated by assessing effects of harvest time, conservation method, iNDF/NDF ratio and NDF intake (NDFI) per kg BW on chewing activity and fecal particle size in dairy heifers. A mixed sward consisting of ryegrass (Lolium perenne), red clover (Trifolium pratense) and white clover (Trifolium repens) was harvested in 2009 on May 9 (early) and 25 (late), and both cuts were conserved as silage and hay. The early silage, early hay, late silage and late hay contained dry matter (DM) of 454, 842, 250 and 828 g/kg, and NDF of 315, 436, 414 and 503 g/kg DM, respectively. Forages were fed as sole feed to four Jersey heifers of 435±30 kg BW in a 4×4 Latin square experiment. Feeding level was 90% of individual ad libitum intake, divided equally across two daily meals offered at 0800 and 1530 h. Chewing activity was estimated from recorded jaw movements (JM) oscillations continuously logged for 96 h and summarized per 24 h as mean effective rumination time and eating time. Eating behavior was further observed during four 20-min test meals. Weight proportion of large feces particles (>1.0 mm) and geometric mean fecal particle size (GPS) were calculated. Potentially indigestible NDF (iNDF) was estimated by incubation for 288 h in situ. The daily DM intake (DMI) decreased with progressing maturity at harvest (P<0.001) while daily NDFI was unaffected by harvest time (P>0.05). Earlier harvest led to less rumination per kg NDFI (P<0.01), similar eating time per kg NDFI (P>0.05) and similar proportion of large particles (P>0.01) compared with later harvest. Rumination time per kg NDFI decreased with higher NDFI per kg BW (P<0.001) and with lower iNDF/NDF ratio (P<0.01). Content and potential digestibility of NDF was greater in hay than in silage from the same harvest probably due to field loss and therefore confounded effects of conservation method. This study of high digestibility grass/clover silage and hay showed that NDF content and NDFI per kg BW affect fecal particle size and rumination time per kg NDF, and suggests implementation of NDFI per kg BW in systems evaluating physical structure in diets.

High value of the radiobiological parameter Dq correlates to expression of the transforming growth factor beta type II receptor in a panel of small cell lung cancer cell lines.

Our panel of SCLC cell lines have previously been examined for their radiobiological characteristics and sensitivity to treatment with TGF beta 1. In this study we examined the possible correlations between radiobiological parameters and the expression of the TGF beta type II receptor (TGF beta-rII). We have, in other studies, shown that the presence of TGF beta-rII was mandatory for transmitting the growth inhibitory effect of TGF beta. The results showed a statistically significant difference in Dq, i.e. the shoulder width of the survival curve, between cell lines expressing TGF beta-rII and cell lines which did not express the receptor (P = 0.01). Cell lines expressing TGF beta-rII had a high Dq-value. TGF beta-rII expression did not correlate with any other radiobiological parameters. We suggest that an intact growth inhibitory pathway mediated by the TGF beta-rII may have a significant role for the repair of radiation induced DNA damage in SCLC.

Acquired TGF beta 1 sensitivity and TGF beta 1 expression in cell lines established from a single small cell lung cancer patient during clinical progression.

Three small cell lung cancer cell lines established from a single patient during longitudinal follow-up were examined for in vitro expression of TGF beta and TGF beta receptors, i.e. the components of an autocrine loop. GLC 14 was established prior to treatment, GLC 16 on relapse after chemotherapy and GLC 19 on recurrence after radiotherapy. TGF beta was detected by ELISA and TGF beta receptors by chemical crosslinking to radiolabelled TGF beta 1. Furthermore, TGF beta and TGF beta receptor mRNAs were detected by northern blot analysis. Expression of type II TGF beta receptor mRNA and protein was found in GLC 16 and GLC 19. These cell lines were also growth inhibited by exogenously administrated TGF beta 1. TGF beta 1 mRNA and protein in its latent form was only expressed in the radiotherapy-resistant cell line, GLC 19. The results indicate that disease progression in this patient was paralleled by a gain in sensitivity to the growth inhibition by TGF beta 1 due to type II TGF beta receptor, and a gain of latent TGF beta 1 protein. Lack of type II receptor expression in GLC 14, which was also resistant to growth inhibition by exogenous TGF beta 1, was not due to gross structural changes in the type II receptor gene, as examined by Southern blotting. Also, the type I receptor could not be detected by ligand binding assay in this cell line, despite expression of mRNA for this receptor. This agrees with previous findings that type I receptor cannot bind TGF beta 1 without co-expression of the type II receptor.