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Hata T.,Mie University | Murakami K.,Nagoya University | Nakatani H.,Nagoya University | Yamamoto Y.,Mie Prefecture Livestock Research Institute | And 2 more authors.
Biochemical and Biophysical Research Communications | Year: 2010

By a series of centrifugation and ultracentrifugation, we could isolate microvesicles with approximately 100 nm in diameter from bovine milk. We also found that approximately 1700 and 1000 ng of total RNA, in which small RNAs were major components, was contained inside the microvesicles isolated from 6 ml of colostrum and mature milk, respectively, despite high RNase activity in the milk. Polyadenylated gene transcripts for major milk proteins and translation elongation factor-1α (EF-1α) were present in the microvesicles, and integrity of some transcripts was confirmed by real-time PCR targeting 5′- and 3′-ends of mRNA and by in vitro translation analysis. Moreover, a considerable amount of mammary gland and immune-related microRNAs were present in the milk-derived microvesicles. Acidification of milk to mimic gastrointestinal tract did not mostly affected RNA yield and quality. The milk related gene transcripts were detected in cultured cells when incubated with milk-derived microvesicles, suggesting cellular uptake of the microvesicle contents including RNA. Our findings suggest that bovine breast milk contains RNAs capable for being transferred to living cells and involved in the development of calf's gastrointestinal and immune systems. © 2010 Elsevier Inc. All rights reserved. Source

Kobayashi K.-I.,Toyohashi University of Technology | Matsui Y.,Mie Prefecture Livestock Research Institute | Maebuchi Y.,Toyohashi University of Technology | Nishino K.,Toyohashi University of Technology | And 2 more authors.
5th European Conference on Colour in Graphics, Imaging, and Vision and 12th International Symposium on Multispectral Colour Science 2010, CGIV 2010/MCS'10 | Year: 2010

The beef quality grade is greatly affected by visible fat content. Especially, in Japanese black (Wagyu) cattle, high fat content is typically valued highly In this paper, we describe the feasibility of beef evaluation by visualizing fat characteristics using near-infrared (NIR) multispectral imaging. An intact raw beef cut from Wagyu cattle was used as an evaluation target. The content of fat and fatty acid, such as the total saturated fatty acid (SFA) content, the total unsaturated fatty acid (UFA) content, myristic acid (C14:0), palmitic acid (C16:0), stearic acid (C18:0), myristoleic acid (C14:1), palmitoleic acid (C16:1), oleic acid (C18:1), and linoleic acid (C18:2) were estimated and visualized. The total SFA content was calculated as the sum of myristic acid, palmitic acid, and stearic acid. Also, the total UFA content was calculated as the sum of myristoleic acid, palmitoleic acid, oleic acid, and linoleic acid. Reference values for the fat content and fatty acid composition were determined by conventional physical and chemical methods. The fatty acid composition was determined from the extracted lipids by Folch's method, by gas chromatography (GC) using its methyl ester. The fat content was determined by using the Gerhardt SOXTHERM. The NIR multispectral images of the sample were acquired by using the SPECIM Spectral Camera SWIR. It works in the wavelength range of 970-2500 nm with 6.3 nm of bandwidth at 320 pixels resolution in spatial domain. The absorbance spectra of each pixel calculated from pixel intensity of subject and reference white standard was used for constructing the prediction model. In total, 33 samples from various parts of the 2 head of Wagyu cattle were measured. Calibrations were performed by a partial least squares (PLS) regression using mean extracted spectra from each individual sample, limited wavelength range from 1000 to 2300 nm. The coefficients of determination (R2) were between 0.68 and 0.87. The ranks by evaluation index (EI) were "B (high accuracy)" and "C (slightly high)". The ratios of the standard error of prediction to the standard deviation (RPD) were between 1.74 and 2.74. These results indicate a sufficient feasibility of the prediction except for myristoleic acid content. The visualizations, which show the spatial distribution of fatty acid content, were performed by applying the model to predict the content of each pixel. Source

Fujita S.,Mie University | Okamoto R.,Mie University | Taniguchi M.,Mie University | Ban-Tokuda T.,Mie University | And 9 more authors.
Journal of Biochemistry | Year: 2013

Hibernation-specific protein (HP) is a plasma protein that regulates hibernation in chipmunks. The HP complex (HP20c) consists of three homologous proteins, HP20, HP25 and HP27, all produced by liver and belonging to the C1q family. To date, HP20c has not been identified in any mammalian species except chipmunk and ground squirrel hibernators. Here, we report a bovine HP20 gene isolated from liver tissue and aortic endothelial cells. Total homology between bovine and chipmunk variants was 63% at the amino acid level. Gene expression was highest in the liver. Western blot revealed HP20 protein in foetal, newborn, calf and adult serum, with highest concentrations in the adult. Similar proteins were detected in sera of other ruminants but not in humans, bears, mice or rats. Bovine HP20 protein was found mainly in ovaries, stomach, heart, kidneys, lungs, testes and prostate, but not in the skeletal muscle. Native HP20 was purified from bovine adult serum as a complex containing 25 and 27 kDa proteins. Mass spectrometry revealed that these proteins are orthologues of chipmunk HP25 and HP27, respectively. Interestingly, bovine HP20 was highly expressed in cattle serum after fasting. Native bovine HP20c may be a useful tool for investigating HP function. © 2013 The Authors 2013. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved. Source

Nishino N.,Okayama University | Ogata Y.,Okayama University | Han H.,Okayama University | Yamamoto Y.,Mie Prefecture Livestock Research Institute
Animal Science Journal | Year: 2015

As a forage source for total mixed ration (TMR) silage production, locally produced crop silage is now used in addition to imported hay. This type of TMR ensiling is regarded as a two-step fermentation process; hence, a survey was carried out to determine whether the bacteria in crop silage affect the subsequent TMR ensiling. Fermentation product contents and bacterial community were determined for TMR silage and its ingredient silages collected in August, October and November. August product contained corn, sorghum and Italian ryegrass silages, October product had wheat silage exclusively and November product did not include any crop silages. Acetic acid, lactic acid, 2,3-butanediol and ethanol were predominant fermentation products in corn, sorghum, Italian ryegrass and wheat silages, respectively. Robust lactic acid fermentation was seen in TMR silage, even if acetate-type and alcohol-type silages were mixed as ingredients. The finding that bacterial community of the TMR silage appeared unrelated to those of ingredient silage supported this. Silages of various fermentation types can therefore be formulated without interfering with lactate-type fermentation in TMR silage. © 2014 Japanese Society of Animal Science. Source

Han H.,Okayama University | Ogata Y.,Okayama University | Yamamoto Y.,Mie Prefecture Livestock Research Institute | Nagao S.,Okayama Prefecture Livestock Research Institute | Nishino N.,Okayama University
Journal of Dairy Science | Year: 2014

The survival of silage lactic acid bacteria (LAB) in the gut of dairy cows was evaluated by examining the LAB communities of silage and gut contents. Samples were collected at 2 different research institutes (Mie and Okayama) that offered total mixed ration (TMR) silage throughout the year. Silage and feces were sampled in August, October, and November at the Mie institute, whereas silage, rumen fluid, and feces were sampled in June and August at the Okayama institute. Denaturing gradient gel electrophoresis using Lactobacillus-specific primers was performed to detect LAB species in the samples. The selected bands were purified for species identification and the band patterns were used for principal component analysis. Lactic acid was the predominant fermentation product in all the TMR silages analyzed, and the lactic acid level tended to be constant regardless of the sampling time and region. A total of 14 LAB species were detected in the TMR silage samples, of which 5 (Lactobacillus acetotolerans, Lactobacillus pontis, Lactobacillus casei, Lactobacillus suebicus, and Lactobacillus plantarum) were detected in the dairy cow feces. Most of the denaturing gradient gel electrophoresis bands for the feces samples were also detected in the rumen fluid, suggesting that any elimination of silage LAB occurred in the rumen and not in the postruminal gut segments. The principal component analysis indicated that the LAB communities in the silage, rumen fluid, and feces were separately grouped; hence, the survival of silage LAB in the cow rumen and lower gut was deemed difficult. It was concluded that, although the gut LAB community is robust and not easily affected by the silage conditions, several LAB species can inhabit both silage and feces, which suggests the potential of using silage as a vehicle for conveying probiotics. © 2014 American Dairy Science Association. Source

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