Utilization of digital differential display to identify differentially expressed genes related to rumen development

This study aimed to identify the genes associated with the development of the rumen epithelium by screening for candidate genes by digital differential display (DDD) in silico. Using DDD in NCBI's UniGene database, expressed sequence tag (EST)‐based gene expression profiles were analyzed in rumen, reticulum, omasum, abomasum and other tissues in cattle. One hundred and ten candidate genes with high expression in the rumen were derived from a library of all tissues. The expression levels of 11 genes in all candidate genes were analyzed in the rumen, reticulum, omasum and abomasum of nine Japanese Black male calves (5‐week‐old pre‐weaning: n = 3; 15‐week‐old weaned calves: n = 6). Among the 11 genes, only 3‐hydroxy‐3‐methylglutaryl‐CoA synthase 2 (HMGCS2), aldo‐keto reductase family 1, member C1‐like (AKR1C1), and fatty acid binding protein 3 (FABP3) showed significant changes in the levels of gene expression in the rumen between the pre‐ and post‐weaning of calves. These results indicate that DDD analysis in silico can be useful for screening candidate genes related to rumen development, and that the changes in expression levels of three genes in the rumen may have been caused by weaning, aging or both. © 2015 Japanese Society of Animal Science     

Characterization of ethylene biosynthesis and its regulation during fruit ripening in kiwifruit,Actinidia chinensis ‘Sanuki Gold’

Ethylene biosynthesis in kiwifruit, Actinidia chinensis ‘Sanuki Gold’ was characterized using propylene, an ethylene analog, and 1-methylcyclopropene (1-MCP), an inhibitor of ethylene perception. In fruit harvested between a young stage (66 days after pollination) (DAP) and an early commercial harvesting stage (143 DAP), 2 days of exposure to propylene were sufficient to initiate ethylene biosynthesis while in fruit harvested at commercial harvesting stage (154 DAP), 4 days of propylene treatment were required. This observation suggests that response of ethylene biosynthesis to propylene treatment in kiwifruit declined with fruit maturity. Propylene treatment resulted in up-regulated expression of AC-ACO1, AC-ACO2, AC-SAM1 and AC-SAM2, prior to the induction of AC-ACS1 and ethylene production, confirming that AC-ACS1 is the rate limiting step in ethylene biosynthesis in kiwifruit. Treatment of fruit with more than 5 μL L^?1 of 1-MCP after the induction of ethylene production subsequently suppressed ethylene production and expression of ethylene biosynthesis genes. Treatment of fruit with 1-MCP at harvest followed with propylene treatment delayed the induction of ethylene production and AC-ACS1 expression for 5 days. These observations suggest that in ripening kiwifruit, ethylene biosynthesis is regulated by positive feedback mechanism and that 1-MCP treatment at harvest effectively delays ethylene production by 5 days.     

Trophinin is expressed in the porcine endometrium during the estrous cycle

We investigated endometrial expression of trophinin mRNA and protein, homophilic cell adhesion molecules, during the estrous cycle of gilts. An immunopositive reaction for trophinin was observed in the luminal and glandular epithelia of the endometrium at all stages of the estrous cycle, but not in endometrial stromal cells or the myometrium. A partial coding sequence of porcine trophinin was similar to sequences in humans and mice, with homologies of 75% and 70%, respectively. As in humans and mice, the trophinin gene is expressed in the endometrium. Trophinin, however, is expressed in the endometrium of the pig throughout the estrous cycle, higher expression levels were observed at some points of the luteal phase, as in humans. These findings suggest that regulation of trophinin gene expression in the pig is different from that in mice, but similar to that in humans. Furthermore, the present results suggest that the pig might be a suitable model for studying the physiological importance of trophinin in early pregnancy in humans.     

Establishment of a new canine cell line (CCT) originated from a cutaneous malignant histiocytosis

A new canine cell line, named CCT, was established from the cutaneous malignant histiocytosis in a 4-year-old male Borzoi. CCT proliferated with loose adherence and doubling time was approximately 30 hr. When co-cultured with latex beads, CCT phagocytized beads vigorously. Lysozyme and vimentin were positive by immunostaining, and non-specific esterase and acid phosphatase were positive by cytochemical staining. These features indicated the cells had a histiocytic nature. Furthermore, by subcutaneous injection to nude mice CCT could successfully form tumors with the morphological and immunohistochemical features similar to the original tumor.     

Effects of a high-protein diet versus dietary supplementation with ammonium chloride on struvite crystal formation in urine of clinically normal cats

OBJECTIVE: To evaluate the effects of a high-protein diet versus dietary supplementation with ammonium chloride (NH4Cl) on struvite crystal formation in the urine of clinically normal cats by measuring the urine concentration of hydrochloric acid (HCl)-insoluble sediment, urine pH, struvite activity product (SAP), number of struvite crystals in urine, and urine volume. ANIMALS: 23 healthy adult cats. PROCEDURE: Urine was fractionated by centrifugation with subsequent extraction of the sediment with 1 N HCl (study 1). Diets containing either 29% crude protein or 55% crude protein were fed to cats in a crossover trial of 3 weeks/period (study 2). Diets supplemented with either sodium chloride (NaCl) or NH4Cl were fed, by use of a 3 x 3 Latin-square design with 3 wk/period (study 3). In studies 2 and 3, urine samples were collected for the last 7 days of each period. RESULTS: The HCl-insoluble sediment contained Tamm-Horsfall glycoprotein (THP; study 1). The high-protein diet (study 2) and dietary supplementation with NH4Cl (study 3) resulted in a decrease in urine pH, SAP, and the number of struvite crystals in urine. However, the high-protein diet decreased urine concentrations of HCl-insoluble sediment containing THP (study 2), in contrast to the NH4Cl supplementation that increased urine volume without a significant effect on the urine concentration of the HCl-insoluble sediment (study 3). CONCLUSIONS AND CLINICAL RELEVANCE: Our results indicate that compared with dietary supplementation with NH4Cl, the high-protein diet is preferable as a urine acidifier for the prevention of struvite crystal formation in clinically normal cats.     

Daily incremental lines in sika deer (Cervus nippon) dentine

This work was designed to observe the dentine incremental lines of the sika deer (Cervus nippon) fawns and to investigate their periodicity using the chronological labeling method with fluorochromes. The incremental lines were observed in decalcified specimens stained by Bodian's silver technique, and the fluorescence-labeled lines were observed in undecalcified and ground specimens. In the silver stained specimens, there were two types of lines, deeply stained thick lines and faintly stained minute regular incremental lines. The intervals and staining intensities of the deeply stained thick lines were very similar to those of the fluorescence-labeled lines in the ground specimens obtained from the same tooth, and hence, it appeared that the both lines were identical. The number of minute incremental lines between the deeply stained thick lines was the same as that of days between the time when each fluorescent labeling injection was made. Therefore, it seemed that each minute incremental line was formed each day. The possibility of age estimation in days using diurnal dentine increments was discussed.     

Production of inter-section hybrids between Primula filchnerae and P. sinensis through ovule culture

Inter-section hybrids were obtained in the reciprocal crosses between Primula filchnerae (2n = 2x = 24) of Sect. Pinnatae and P. sinensis ‘Fanfare’ (2n = 2x = 24) of Sect. Auganthus by rescuing ovules on half-strength (1/2) Murashige and Skoog's (MS) medium supplemented with 50 g l⁻¹ sucrose, 2.5 g l⁻¹ gellan gum, 0.1 mg l⁻¹ α-naphthaleneacetic acid (NAA), 0.1 mg l⁻¹ 6-benzyladenine (BA) and 50 mg l⁻¹ gibberellic acid (GA₃). In ovule culture, germination occurred with radicle elongation but no plumule was observed. The radicle kept on the initial medium showed root proliferation with callus formation. When the calluses were transferred to (1/2)MS media containing 30 g l⁻¹ sucrose and 3 g l⁻¹ gellan gum, without plant growth regulators (PGRs) or with 1 mg l⁻¹ zeatin and 0.1 mg l⁻¹ NAA, plantlets were regenerated. The plants thus obtained were confirmed to be hybrids through flow cytometry (FCM) and random amplified polymorphic DNA (RAPD) analyses. The hybrid obtained when P. filchnerae was used as the maternal parent was diploid, whereas hexaploid hybrid was obtained when using P. sinensis as the maternal parent. The hexaploid hybrid might be produced through chromosome doubling of a triploid originated from the fertilization of P. sinensis with unreduced pollen of P. filchnerae.