Chronic liver disease due to alcohol use disorder contributes markedly to the global burden of disease and mortality 1-3. Alcoholic hepatitis is a severe and life-threatening form of alcohol-Duan et al.

We found that Ku70, a known DNA repair factor, has a novel function to bind and inhibit Bax (Bcl-2-associated X protein), a key mediator of apoptosis. Pentapeptides derived from the Bax-binding domain of Ku70 were cell-permeable and protected cells from Bax-mediated apoptosis. These pentapeptides were called BIPs (Bax-inhibiting peptides). BIPs may become a useful therapeutic tool to reduce cellular damage. We also generated BIP mutant pentapeptides that do not inhibit Bax, but retain their cell-penetrating activity. Since both BIPs and BIP mutants are cell-permeable, these peptides were designated CPP5s (cell-penetrating pentapeptides). Among the CPP5s discovered, VPTLK (BIP) and KLPVM (BIP mutant) were confirmed to possess protein transduction activity by examination of the delivery of GFP (green fluorescent protein) into cells by these peptides. The mechanism of cell penetration by CPP5s is not known. CPP5s enter the cell at 0 and 4 degrees C. In preliminary studies, various inhibitors of endocytosis and pinocytosis did not show any significant suppression of CPP5 cell entry. CPP5s have very low toxicity in vitro and in vivo and so may be useful tools in order to develop non-toxic drug-delivery technologies.

The expression level of the telomerase catalytic subunit (telomerase reverse transcriptase, TERT) positively correlates with cell survival after exposure to several lethal stresses. However, whether the protective role of TERT is independent of telomerase activity has not yet been clearly explored. Here, we genetically evaluated the protective roles of both TERT and telomerase activity against cell death induced by staurosporine (STS) and N-methyl-Daspartic acid (NMDA). First generation (G1) TERTdeficient mouse embryonic fibroblasts (MEFs) displayed an increased sensitivity to STS, while TERT transgenic MEFs were more resistant to STS-induced apoptosis than wildtype. Deletion of the telomerase RNA component (TERC) failed to alter the sensitivity of TERT transgenic MEFs to STS treatment. Similarly, NMDA-induced excitotoxic cell death of primary neurons was suppressed by TERT, but not by TERC both in vitro and in vivo. Specifically, NMDA accelerated death of TERT-deficient mice, while TERT transgenic mice showed enhanced survival when compared with wild-type littermates after administration of NMDA. In addition, the transgenic expression of TERT protected motor neurons from apoptosis induced by sciatic nerve axotomy. These results indicate that telomerase activity is not essential for the protective function of TERT. This telomerase activity-independent TERT function may contribute to cancer development and aging independently of telomere lengthening.

Reference genes are used as internal controls in gene expression studies, but their expression levels vary according to tissue types and experimental treatments. Quantitative real-time PCR (qPCR) is the most sensitive technique for transcript quantification provided that gene transcription patterns are normalized to an evaluated reference gene. In this study, the suitability of eight commonly used genes (β?-actin, 5.8SrRNA, α?-TUB, GAPDH, RPL13a, RPS18, TBP, SDHA) were cloned and investigated to find the most stable candidates for normalizing real-time PCR data generated from the four different strains (abamectin-resistant, fenpropathrin-resistant, omethoate-resistant, and susceptible strains) and different developmental stages (eggs, protonymphs, nymphs, and adults) of carmine spider mite, Tetranychus cinnabarinus (Boisduval) (Acarina: Tetranychidae). The stability of gene expression was assessed using two different analysis programs, geNorm and NormFinder. Using these analyses, RPS18 and 5.8SrRNA had the most stable expression regardless of the four different strains, whereas RPS18 and α?-TUB were expressed most stably in different developmental stages.

Vitamin A serves essential functions in mammalian biology as a signaling molecule and chromophore. This lipid can be synthesized from more than 50 putative dietary provitamin A precursor molecules which contain at least one unsubstituted β-ionone ring. We here scrutinized the enzymatic properties and substrate specificities of the two structurally related carotenoid cleavage dioxygenases (CCDs) which catalyze this synthesis. Recombinant BCO1 split substrates across the C15,C15' double bond adjacent to a canonical β-ionone ring site to vitamin A aldehyde. Substitution of the ring with a hydroxyl group prevented this conversion. The removal of methyl groups from the polyene carbon backbone of the substrate did not impede enzyme activity. Homology modeling and site-directed mutagenesis identified amino acid residues at the entrance of the substrate tunnel, which determined BCO1's specificity for the canonical β-ionone ring site. In contrast, BCO2 split substrates across the C9,C10 double bond adjacent to assorted ionone ring sites. Kinetic analysis revealed a higher catalytic efficiency of BCO2 with substrates bearing 3-hydroxy-β-ionone rings. In the mouse intestine, the asymmetric carotenoid β-cryptoxanthin with one canonical and one 3-hydroxy-β-ionone ring site was meticulously converted to vitamin A. The tailoring of this asymmetric substrate occurred by a stepwise processing of the carotenoid substrate by both CCDs and involved a β-apo-10'-carotenal intermediate. Thus, opposite selectivity for ionone ring sites of the two mammalian CCDs complement each other in the metabolic challenge of vitamin A production from a chemically diverse set of precursor molecules.

This study was conducted to compare the effects of adding sodium butyrate (SB), medium-chain fatty acids (MCFAs), or n-3 polyunsaturated fatty acids (n-3 PUFAs) to the diet of sows during late gestation and lactation on the reproductive performance of sows and the growth performance and intestinal health of suckling piglets. Twenty-four sows (Landrace × Large-White hybrid; third parity; 200 ± 15 kg) were randomly assigned to receive 1 of 4 diets: basal diet (control group), basal diet + 1 g SB/kg (SB group), basal diet + 7.75 g MCFA/kg (MCFA group), or basal diet + 68.2 g n-3 PUFA/kg (n-3 PUFA group). The experiment began on day 85 of gestation and ended day 22 of lactation. Colostrum samples were collected from each sow. After the experiment, blood and tissue samples were collected from 1 randomly selected piglet. The results showed that the weaning-to-estrus interval of sows in the SB, MCFA, and n-3 PUFA groups was shorter than that of sows in the control group (P < 0.05). The incidence of diarrhea in suckling piglets in the SB, MCFA, and n-3 PUFA groups was lower than that of piglets in the control group (P < 0.05). The fat, protein, IgA, IgG, and IgM concentration in colostrum from sows increased following dietary supplementation with SB, MCFA, or n-3 PUFA (P < 0.05). Comparison with the control group, the mRNA expression of claudin-1, zona occludens 1, and interleukin-10 increased in the jejunum mucosa of suckling piglets in the SB, MCFA, and n-3 PUFA groups, while that of TLR4 decreased (P < 0.05). Compared with the control group, the Chao1 and ACE indexes of microbial flora in the colon contents of piglets in the SB, MCFA, and MCFA groups increased (P < 0.05), while the relative abundance of Firmicutes, Actinobacteria, and Synergistetes decreased at the phylum level (P < 0.05). In conclusion, during late pregnancy and lactation, dietary SB supplementation had a greater effect on intestinal health and caused a greater decrease in preweaning mortality of suckling piglets than did dietary MCFA or n-3 PUFA supplementation; dietary MCFA supplementation shortened the weaning-to-estrus interval of sows to a greater extent than did dietary SB or n-3 PUFA supplementation; and dietary n-3 PUFA supplementation increased the fat and protein content in the colostrum to the greatest extent.

ABSTRACT. Immune-related miRNAs in breast milk are extracellular miRNAs that are related to immune organ development and regulation of the immune function in infants and young animals. The goal of this study was to compare the expression levels of five immune-related miRNAs in breast milk in black goats, humans, and dairy cattle. The miRNAs from milk were extracted and the expression levels were assessed using quantitive RT-PCR methods. MiR-146, miR-155, miR-181a, miR-223, and miR-150 were all detected in Dazu black goat milk, and these miRNAs were significantly more highly expressed in colostrum than in mature milk of goats (P < 0.01), except for miR-150. Further, all five miRNAs were expressed in human colostrum, but patterns differed from those in goats: miR-146 and miR-155 were highly expressed (P < 0.01) in human colostrum, whereas miR-223 was abundant in goat colostrum (P < 0.01). In addition, five miRNAs were significantly higher in bovine mature milk than in goat milk (P < 0.01). Taken together, R.S. Na et al. 11372©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 14 (3): 11371-11376 (2015) these results confirm that immune-related miRNAs are rich in breast milk with different expression levels depending on the lactation phase and species.

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