Effects of branched-chain amino acids on glucose uptake and lactose synthesis rates in bovine mammary epithelial cells and lactating mammary tissue slices.

Even though supplementations of essential AA (EAA) are often related to increased lactose yields in dairy cows, underlying mechanisms connecting EAA availability to the mammary glands and lactose synthesis are poorly understood. The objective of this study was to examine the effects of branched-chain AA (BCAA) including Leu, Ile, and Val on (1) glucose transporter (GLUT1) abundance and glucose uptake, (2) the abundance of proteins regulating lactose synthesis pathway, and (3) fractional synthesis rates of lactose (FSR) using bovine mammary epithelial cells (BMEC) and mammary tissues slices (MTS). The BMEC (n = 4) were allocated randomly to regular Dulbecco's Modified Eagle Medium with Ham's F12 (DMEM/F12) medium (+EAA) or +EAA deficient (by 90%) in all EAA (-EAA), all BCAA (-BCAA), only Leu (-Leu), only Ile (-Ile) or only Val (-Val). Western immunoblotting analyses, depletion of glucose in media, and a proteomic analysis were performed to determine the abundance of GLUT1 in the cell membrane, net glucose uptake, and the abundance of enzymes involved in lactose synthesis pathway in BMEC, respectively. The MTS (n = 6) were allocated randomly to DMEM/F12 medium having all EAA and 13C-glucose at concentrations similar to plasma concentrations of cows (+EAAp), and +EAAp deprived of all BCAA (-BCAAp) or only Leu (-Leup) for 3 h. The 13C enrichments of free glucose pool in MTS (EGlu-free) and the enrichments of glucose incorporated into lactose in MTS and media [ELactose-bound (T&M)] were determined and used in calculating FSR. In BMEC, -BCAA increased the fraction of total GLUT1 translocated to the cell membrane and the fraction that was potentially glycosylated compared with +EAA. Among individual BCAA, only -Leu was associated with a 63% increase in GLUT1 translocated to the cell membrane and a 40% increase in glucose uptake of BMEC. The -BCAA tended to be related to a 75% increase in the abundance of hexokinase in BMEC. Deprivation of Leu tended to increase glucose uptake of MTS but did not affect EGlu-free, ELactose-bound (T&M), or FSR relative to +EAAp. On the other hand, -BCAAp did not affect glucose uptake of MTS but was related to lower ELactose-bound (T&M), or FSR relative to +EAAp. Considering together, decreasing Leu supply to mammary tissues enhances GLUT1 and thus glucose uptake, which, however, does not affect lactose synthesis rates. Moreover, the deficiency of other BCAA, Ile, and Val alone or together with the deficiency of Leu seemed to decrease lactose synthesis rates without affecting glucose uptake. The data also emphasize the importance of addressing the effect of the supply of other nutrients to the mammary glands than the precursor supply in describing the synthesis of a milk component.

In vitro photodynamic lipid peroxidation of total lipophilic extracts from leaves of bean plants.

In vitro lipid peroxidation, induced by reactive oxygen species photochemically generated, was carried out on purified alpha-linolenic acid (18:3(n - 3)) and on bean leaf total lipophilic extracts. The photosensitizer used was meso-tetraphenylporphine. The time-course of the reaction was evaluated by ultraviolet (UV) spectra analysis. The 18:3(n - 3) photoperoxidation kinetics comprised three steps: monohydroperoxidation, characterized by the appearance of conjugated diene patterns; dihydroperoxidation characterized by the appearance of conjugated triene patterns, oxidative cleavage of the mono- and dihydroperoxides, characterized by the disappearance of conjugated patterns. In contrast, for hydrated plant total lipophilic extract photoperoxidation, conjugated pattern appearance was slow and the maximum plateau was not obtained. In order to explain plant extract behaviour, we tested the effects of beta-carotene and quercetin (important components of the chloroplast membrane) on the time-course of 18:3(n - 3) photoperoxidation. The first step was inhibited by beta-carotene implying that type II photoperoxidation involving singlet oxygen (1O2) was predominant. Whilst the two last steps were inhibited by quercetin implying that type I photoperoxidation involving free radicals, prevailed. Since 18:3(n - 3) foliar content decreased under water deficit, we tested the behaviour of total lipophilic extracts from droughted plant leaves in presence of reactive oxygen species. In the case of droughted plants, the maximum of conjugated diene patterns was attained later than for hydrated plants, suggesting that drought affects the resistance capability of total lipophilic extracts from bean leaves to lipid photoperoxidation.

Evaluation of the purified fraction of Wilbrandia (c.f.) verticillata for antitumour activity.

Cucurbitacins are known to produce cytotoxic and anticancer activities. Two novel norcucurbitacin glucosides (Wv1 and Wv2) have recently been isolated from a purified fraction obtained from the rhizome of Wilbrandia verticillata. The present study evaluates the cytotoxic and antitumour activities of these norcucurbitacins. We have found a regular cytotoxicity in KB cells (Cy50 = 12 micrograms/ml) as well as a significant inhibition in the Walker 256 carcinosarcoma growth (approximately 75%).

Evaluation of the purified fraction of Wilbrandia (c. f. ) verticillata for antitumour activity

Cucurbatacins are known to produce cytotoxic and anticancer activities. Two novel norcucurbitacin glucosides (Wvl and Wv2) have recently been isolated from a purified fraction obtained from the rhizome of Wilbrandia verticillata. The present study evaluates the cytotoxic and anti-tumour activities of the norcucurbitacins. We have found a regular cytotoxicity in KB cells (Cy50 = 12µg/ml) as well as a significant inhibition in the Walker 256 carcinosarcoma growth (approximately 75%).