Differences during the first lactation between cows cloned by somatic cell nuclear transfer and noncloned cows.

Lactation performance is dependent on both the genetic characteristics and the environmental conditions surrounding lactating cows. However, individual variations can still be observed within a given breed under similar environmental conditions. The role of the environment between birth and lactation could be better appreciated in cloned cows, which are presumed to be genetically identical, but differences in lactation performance between cloned and noncloned cows first need to be clearly evaluated. Conflicting results have been described in the literature, so our aim was to clarify this situation. Nine cloned Prim' Holstein cows were produced by the transfer of nuclei from a single fibroblast cell line after cell fusion with enucleated oocytes. The cloned cows and 9 noncloned counterparts were raised under similar conditions. Milk production and composition were recorded monthly from calving until 200d in milk. At 67d in milk, biopsies were sampled from the rear quarter of the udder, their mammary epithelial cell content was evaluated, and mammary cell renewal, RNA, and DNA were then analyzed in relevant samples. The results showed that milk production did not differ significantly between cloned and noncloned cows, but milk protein and fat contents were less variable in cloned cows. Furthermore, milk fat yield and contents were lower in cloned cows during early lactation. At around 67 DIM, milk fat and protein yields, as well as milk fat, protein, and lactose contents, were also lower in cloned cows. These lower yields could be linked to the higher apoptotic rate observed in cloned cows. Apoptosis is triggered by insulin-like factor growth binding protein 5 (IGFBP5) and plasminogen activator inhibitor (PAI), which both interact with CSN1S2. During our experiments, CSN1S2 transcript levels were lower in the mammary gland of cloned cows. The mammary cell apoptotic rate observed in cloned cows may have been related to the higher levels of DNA (cytosine-5-)-methyltransferase 1 (DNMT1) transcripts, coding for products that maintain the epigenetic status of cells. We conclude, therefore, that milk production in cloned cows differs slightly from that of noncloned cows. These differences may be due, in part, to a higher incidence of subclinical mastitis. They were associated with differences in cell apoptosis and linked to variations in DNMT1 mRNA. However, milk protein and fat contents were more similar among cloned cows than among noncloned cows.

Milk from dams fed an obesogenic diet combined with a high-fat/high-sugar diet induces long-term abnormal mammary gland development in the rabbit.

Alterations to the metabolic endocrine environment during early life are crucial to mammary gland development. Among these environmental parameters, the initial nutritional event after birth is the consumption of milk, which represents the first maternal support provided to mammalian newborns. Milk is a complex fluid that exerts effects far beyond its immediate nutritional value. The present study, therefore, aimed to determine the effect of the nutritional changes during the neonatal and prepubertal periods on the adult mammary phenotype. Newborn rabbits were suckled by dams fed a high-fat/high-sugar obesogenic (OD) or a control (CON) diet and then subsequently fed either the OD or CON diets from the onset of puberty and throughout early pregnancy. Mammary glands were collected during early pregnancy (Day 8 of pregnancy). Rabbits fed with OD milk and then subjected to an OD diet displayed an abnormal development of the mammary gland: the mammary ducts were markedly enlarged (P < 0.05) and filled with abundant secretory products. Moreover, the alveolar secretory structures were disorganized, with an abnormal aspect characterized by large lumina. Mammary epithelial cells contained numerous large lipid droplets and exhibited fingering of the apical membrane and abnormally enlarged intercellular spaces filled with casein micelles. Leptin has been shown to be involved in modulating several developmental processes. We therefore analyzed its expression in the mammary gland. Mammary leptin mRNA was strongly expressed in rabbits fed with OD milk and subjected to an OD diet by comparison with the CON rabbits. Leptin transcripts and protein were localized in the epithelial cells, indicating that the increase in leptin synthesis occurs in this compartment. Taken together, these findings suggest that early-life nutritional history, in particular through the milking period, can determine subsequent mammary gland development. Moreover, they highlight the potentially important regulatory role that leptin may play during critical early-life nutritional windows with respect to long-term growth and mammary function.

Nuclear and cytoplasmic maturation of canine oocytes related to in vitro denudation.

In vitro maturation (IVM) of bitch oocytes is, to date, a very inefficient process, with common metaphase rates approximately 0-20% and mean degeneration rates approximately 20-30%. In other mammals, meiotic resumption is controlled in the cumulus-oocyte complex by the disappearance of the coupling between granulosa cells and the oocyte. The first aim of this study was to evaluate the influence on meiotic resumption of a mechanical denudation of the oocytes before maturation. The nuclear stage was determined by DNA staining with ethidium-homodimer-2 under confocal microscopy. Denuded (n = 318) and control (n = 378; no mechanical denudation) oocytes had similar degeneration rates (respectively 32.1 vs 28.6%). However, meiosis resumption rates were significantly higher for denuded oocytes (DO; 26.9 vs 17.8%). Secondly, we aimed to evaluate oocytes experiencing spontaneous denudation during the 72 h IVM period. Denuded oocytes, having lost cumulus cells on at least 75% of their perimeter (n = 440), were compared with surrounded oocytes (SO), with 100% of their perimeter surrounded by granulosa cells (n = 860). As above, the nuclear stage was determined by confocal microscopy, but cytoplasmic maturation was also evaluated through transmission electron microscopy. Degeneration rates but also meiosis resumption and metaphase rates were significantly higher for denuded than for SO (9.6 vs 2.8% for metaphase rate). Nevertheless, ultrastructurally, metaphase DO have scarcer organelles unevenly distributed, with smooth endoplasmic reticulum concentrated in aggregates in the cortical zone. Denudation, whether mechanical or spontaneous, is thus an inefficient mean to obtain metaphase II oocytes suitable for in vitro fertilization.

Inter-relationships between DNA damage, ascorbic acid and glycaemic control in Type 2 diabetes mellitus.

AIMS: The onset of complications in Type 2 diabetes mellitus (DM) patients cannot be predicted in individuals. Evidence suggests a link between complications and hyperglycaemia, oxidative stress and antioxidants, but causality is unclear. This study investigated baseline (entry) fasting plasma ascorbic acid, lymphocytic DNA damage and glycaemic control in Type 2 DM as part of a long-term study, the aim of which is to explore a biomarker profiling approach to identify and improve outcome in high-risk subjects. METHODS: A cross-sectional study, in which DNA damage, glycated haemoglobin (HbA(1c)), fasting plasma glucose (FPG) and ascorbic acid (AA) were measured on fasting blood samples collected from 427 Type 2 DM subjects. RESULTS: DNA damage was significantly (P < 0.0001) and directly correlated to both FPG (r = 0.540) and HbA(1c) (r = 0.282), and was significantly (P < 0.0001), independently and inversely correlated to plasma AA (r = -0.449). In those subjects with both poor glycaemic control and low AA (< 48 microm, the overall mean value for the study group), DNA damage was significantly (P < 0.005) higher compared with those subjects with a similar degree of hyperglycaemia but with AA above the mean. CONCLUSIONS: The novel finding of a significant inverse relationship between plasma AA and DNA damage in Type 2 DM indicates that poorly controlled diabetic subjects might benefit from increased dietary vitamin C. The data also have important implications for biomarker profiling to identify those subjects who might benefit most from intensive therapy. Longer-term follow-up is underway.

Clinical evaluation of the Shin-Nippon SRW-5000 autorefractor in children.

The Canon Autoref R-1 is an 'open-field' autorefractor which has been widely used for research purposes for the past 20 years, but is no longer manufactured. A new autorefractor, the Shin-Nippon SRW-5000, is now available, and if measures using this instrument are shown to be equally accurate and reliable, is likely to replace the R-1. Here we report on the accuracy and reliability (repeatability and reproducibility) of refraction measures in a paediatric population (from 4 to 8 years of age). Subject numbers were 44 for cycloplegic measures and 53 for non-cycloplegic measures. As would be expected, agreement with cycloplegic refraction and reliability were better when SRW-5000 measures were taken using cycloplegia. Repeatability results from the SRW-5000 autorefractor, both with and without cycloplegia were similar to those reported for the Canon R-1.