Changes of beta-casomorphin content in human milk during lactation

Milk is the best, complete food important for the development and nourishment of a neonate. Except for nutrients, milk contains biologically active opioid peptides derived from beta-casein, named beta-casomorphins (BCMs), which can exert effects in the gastrointestinal tract as well as in the whole body of neonates. The content of beta-casomorphins in human milk during maturation phases has not been studied so far. The aim of this study was to determine the content of beta-casomorphin-5 and -7 in human milk in different phases of lactation. A significantly higher concentration of both beta-casomorphins was found in colostrum than in mature milk. The concentration of beta-casomorphin in milk collected in the second month of lactation was similar to the level obtained in the fourth month of lactation. The content of beta-casomorphins in human milk was observed with the period of lactation. The level of opioid peptides may depend on the function of these peptides in neonate's body and may be associated with the maturation process.     

Transgenic Mice Expressing Recombinant Human Protein C Exhibit Defects in Lactation and Impaired Mammary Gland Development

To determine if the production of recombinant human protein C (rHPC) could be increased in milk, we created two lines of mice homozygous for the mouse whey acidic protein (WAP)/human protein C (HPC) transgene. Females of both lines had normal growth, activity and fertility, but failed to lactate normally and were unable to raise litters. Histological analyses of mammary glands from lactating homozygous females showed barely distended alveoli filled with dense-staining milk. Epithelial cells within these alveoli had distinct, centrally located nuclei and contained intracellular lipid droplets. Hemizygous animals derived from these lines were able to lactate and raised normal sized litters. Northern blot analysis showed that the 6.4 homozygous (6.4H) line expressed the transgene at higher levels then corresponding hemizygous (6.4) animals, but the 4.2 homozygous (4.2H) line expressed the transgene at lower levels than the 4.2 hemizygous line. The 6.4H line also had increased rHPC levels in the milk as revealed by western blot analysis. The 4.2H, 6.4, and 6.4H lines showed decreased and/or delayed expression of WAP, -casein, and -lactalbumin mRNA's compared to wild type animals during lactogenesis. The 4.2 line showed decreased mRNA expression for -casein and -lactalbumin, but normal or higher expression of WAP during lactogenesis. Elevated levels of some proteins were detected in the milk of transgenic mice. From these results, it is concluded that expression of rHPC induced a lactational phenotype that involves abnormal morphological, biochemical, and functional differentiation of mammary epithelial cells. However, the induction of this phenotype does not appear to be directly related to the level of rHPC mRNA expression, thus suggesting that the basis of this phenotype may involve secondary, rather than primary, effects of rHPC on mammary gland development.Deceased.     

Frequent coinfection of cells explains functional in vivo complementation between cytomegalovirus variants in the multiply infected host

In contrast to many other virus infections, primary cytomegalovirus (CMV) infection does not fully protect against reinfection. Accordingly, clinical data have revealed a coexistence of multiple human CMV variants/strains in individual patients. Notably, the phenomenon of multiple infection was found to correlate with increased virus load and severity of CMV disease. Although of obvious medical relevance, the mechanism underlying this correlation is unknown. A weak immune response in an individual could be responsible for a more severe disease and for multiple infections. Alternatively, synergistic contributions of variants that differ in their biological properties can lead to qualitative changes in viral fitness by direct interactions such as genetic recombination or functional complementation within coinfected host cells. We have addressed this important question paradigmatically with the murine model by differently designed combinations of two viruses employed for experimental coinfection of mice. Specifically, a murine cytomegalovirus (MCMV) mutant expressing Cre recombinase was combined for coinfection with a mutant carrying Cre-inducible green fluorescent protein gene, and attenuated mutants were combined for coinfection with wild-type virus followed by two-color in situ hybridization studies visualizing the replication of the two viruses in infected host organs. These different approaches concurred in the conclusion that coinfection of host cells is more frequent than statistically predicted and that this coinfection alters virus fitness by functional trans-complementation rather than by genetic recombination. The reported findings make a major contribution to our molecular understanding of enhanced CMV pathogenicity in the multiply infected host.     

The prevalence of overweight and obesity among Croatian hospitalized coronary heart disease patients

The aim of this article was to investigate the prevalence of overweight and obesity using selected anthropometric variables in a sample of hospitalized coronary heart disease (CHD) patients in Croatia (N = 1,298). Prevalence of overweight and obesity in surveyed patient population was high: 48.2% of participants were overweight and 28.6% were obese according to their body mass index; measured through waist-to-hip ratio 54.5% of participants were centrally obese. These data on prevalences of overweight, obesity and central obesity show that although there are some reassuring trends, there is still considerable amount of work to be done if the prevalence of this cardiovascular risk factor is to be reduced further among Croatian CHD patients. While the prevalence of obesity seems to be on the decline, the prevalence of overweight is rising, which may be just an early warning sign of an incoming wave of obesity epidemic in future years.     

Expression of stress gene networks in tomato lines susceptible and resistant to Tomato yellow leaf curl virus in response to abiotic stresses.

The defense response to several abiotic stresses has been compared in two tomato inbred lines issued from the same breeding program, one susceptible and the other resistant to Tomato yellow leaf curl virus (TYLCV) infection. The level of oxidative burst and the amounts of key regulatory stress proteins: pathogenesis-related proteins (PRs), heat shock proteins (HSPs) and mitogen-activated protein kinases (MAPKs) were appraised following treatments with NaCl, H(2)O(2), and ethanol. Significant differences in the response of the two tomato genotypes to these stresses have been found for HSPs and MAPKs patterns at the level of down-regulation but not activation. The higher abundance of HSPs and MAPKs in tomatoes resistant to TYLCV could result in enhanced defense capacity against abiotic stresses.     

Fusion of oil bodies in endosperm of oat grains

Few microscopical studies have been made on lipid storage in oat grains, with variable results as to the extent of lipid accumulation in the starchy endosperm. Grains of medium- and high-lipid oat (Avena sativa L.) were studied at two developmental stages and at maturity, by light microscopy using different staining methods, and by scanning and transmission electron microscopy. Discrete oil bodies occurred in the aleurone layer, scutellum and embryo. In contrast, oil bodies in the starchy endosperm often had diffuse boundaries and fused with each other and with protein vacuoles during grain development, forming a continuous oil matrix between the protein and starch components. The different microscopical methods were confirmative to each other regarding the coalescence of oil bodies, a phenomenon probably correlated with the reduced amount of oil-body associated proteins in the endosperm. This was supported experimentally by SDS-PAGE separation of oil-body proteins and immunoblotting and immunolocalization with antibodies against a 16 kD oil-body protein. Much more oil-body proteins per amount of oil occurred in the embryo and scutellum than in the endosperm. Immunolocalization of 14 and 16 kD oil-body associated proteins on sectioned grains resulted in more heavy labeling of the embryo, scutellum and aleurone layer than the rest of the endosperm. Observations on the appearance of oil bodies at an early stage of development pertain to the prevailing hypotheses of oil-body biogenesis.     

Histone demethylase LSD1 is a folate-binding protein

Methylation of lysine residues in histones has been known to serve a regulatory role in gene expression. Although enzymatic removal of the methyl groups was discovered as early as 1973, the enzymes responsible for their removal were isolated and their mechanism of action was described only recently. The first enzyme to show such activity was LSD1, a flavin-containing enzyme that removes the methyl groups from lysines 4 and 9 of histone 3 with the generation of formaldehyde from the methyl group. This reaction is similar to the previously described demethylation reactions conducted by the enzymes dimethylglycine dehydrogenase and sarcosine dehydrogenase, in which protein-bound tetrahydrofolate serves as an accepter of the formaldehyde that is generated. We now show that nuclear extracts of HeLa cells contain LSD1 that is associated with folate. Using the method of back-scattering interferometry, we have measured the binding of various forms of folate to both full-length LSD1 and a truncated form of LSD1 in free solution. The 6R,S form of the natural pentaglutamate form of tetrahydrofolate bound with the highest affinity (K(d) = 2.8 μM) to full-length LSD1. The fact that folate participates in the enzymatic demethylation of histones provides an opportunity for this micronutrient to play a role in the epigenetic control of gene expression.