title: Expression of innate resistance factors in mammary secretion from periparturient dairy heifers and their association with subsequent infection status

authors: Compton, Chris W.R.

subjects: bovine

abstract: Mastitis in dairy heifers in the peripartum period is a common and costly problem for producers, and mammary innate resistance is of key importance in defense of the gland from bacterial invasion. A prospective observational study was undertaken in 97 dairy heifers to measure associations between expression of eight innate resistance factors in mammary secretions collected from the same animals within 14 days prior to calving and at calving, and intramammary infection (IMI) status at calving, and to describe changes in expression of these factors over time. Relative expression (RE) of eight candidate resistance mediator genes from cells from intramammary secretion was measured by quantitative real time polymerase chain reaction. Glands which were IMI-free pre-calving and did not develop a new IMI had significantly higher RE of molecule possessing ankyrin-repeat (MAIL) and beta defensin (Bdef) genes compared to glands which subsequently did develop a new IMI. Also, Bdef RE increased up to the day of calving in glands that did not develop a new IMI, but was unresponsive in glands that did develop a new IMI. Relative expression of complement 5 alpha receptor, interleukin 1beta and interleukin 8 increased in glands that did develop a new IMI. Serum amyloid A3 and toll-like receptor 2 RE increased in all glands up to the day of calving. Transforming growth factor beta RE was not associated with new infection status or time relative to calving. These findings support further investigation of function and gene polymorphisms of MAIL and Bdef as potential markers of mastitis resistance in dairy heifers.

text: Expression of innate resistance factors in mammary secretion from periparturient dairy heifers and their association with subsequent infection status Mastitis in dairy heifers in the peripartum period is a common and costly problem for producers, and mammary innate resistance is of key importance in defense of the gland from bacterial invasion. A prospective observational study was undertaken in 97 dairy heifers to measure associations between expression of eight innate resistance factors in mammary secretions collected from the same animals within 14 days prior to calving and at calving, and intramammary infection (IMI) status at calving, and to describe changes in expression of these factors over time. Relative expression (RE) of eight candidate resistance mediator genes from cells from intramammary secretion was measured by quantitative real time polymerase chain reaction. Glands which were IMI-free pre-calving and did not develop a new IMI had significantly higher RE of molecule possessing ankyrin-repeat (MAIL) and beta defensin (Bdef) genes compared to glands which subsequently did develop a new IMI. Also, Bdef RE increased up to the day of calving in glands that did not develop a new IMI, but was unresponsive in glands that did develop a new IMI. Relative expression of complement 5 alpha receptor, interleukin 1beta and interleukin 8 increased in glands that did develop a new IMI. Serum amyloid A3 and toll-like receptor 2 RE increased in all glands up to the day of calving. Transforming growth factor beta RE was not associated with new infection status or time relative to calving. These findings support further investigation of function and gene polymorphisms of MAIL and Bdef as potential markers of mastitis resistance in dairy heifers.

year: 2008

title: Epigenetic reprogramming of somatic cells by zygotic factors

authors: Sowry, Blair Gavin

subjects: Reprogramming

abstract: Cloning cattle using somatic cell nuclear transfer (SCNT) is an inefficient process, with approximately only 5% of transferred embryos developing to live offspring. SCNT produced cattle have a high mortality rate due to a number of developmental abnormalities caused mainly by the incorrect epigenetic reprogramming of the donor cell to a pluripotent state. Conventional bovine SCNT involves fusion of a somatic donor cell into an enucleated metaphase II (MII) arrested oocyte. The resulting embryo is cultured to the blastocyst stage before being transferred to a surrogate cow to produce live offspring. Zygotes were initially thought to be unsuitable as SCNT recipients in bovine, until it was revealed that using telophase II (TII) zygotes as opposed to MII oocytes results in improved in vivo development. Metaphase zygotes have also been used successfully as SCNT recipients in mouse to produce cloned blastocysts and it has been proposed that reprogramming factors sequester in the pronuclei of interphase zygotes. Little is known about the nature the nuclear reprogramming, however a few candidate reprogramming factors have emerged recently. TCTP is known to activate key pluripotency genes (POU5F1 and NANOG) in somatic cell nuclei. It has also been identified as present in bovine oocytes with a high potential to reprogram somatic cells. Reprogramming of cell nuclei by Xenopus egg extract has found to require BRG1. Immunodepletion of BRG1 was shown to decrease the reprogramming ability of the egg extract, whist its over-expression increased reprogramming potential. HDAC1 has been found to initiate a transcriptionally repressive state in preimplantation mouse embryos possibly inhibiting transcription of reprogramming factors. Knockdown of HDAC1 using TSA is known to increase development of mouse NT embryos. 
The aim of this study was to produce blastocysts using metaphase zygotes as recipients for SCNT in bovine. In addition, localisation and abundance of candidate reprogramming factors TCTP (TPT1 gene), BRG1 and HDAC1 were examined in MII oocytes as well as TII, interphase and metaphase zygotes.

This study found metaphase zygotes are unsuitable as recipients for bovine SCNT using current methodologies, possibly due to the premature cleavage of the embryos. Control embryos produced using MII oocytes as recipients developed to blastocyst with an efficiency of ~ 11%. mRNA analysis of zygotes and oocytes did not reveal any significant differences in the relative concentrations of TPT1 or BRG1 between the samples. The TCTP and HDAC1 proteins showed a similar pattern of localisation in the MII oocytes and all stages of zygotes. Both proteins clearly localise to the maternal chromatin in the second polar body of TII zygotes. This finding has never previously been described and may in part explain why there is increased cloning efficiency observed when using TII zygotes as SCNT recipients.

This increased understanding of these reprogramming factors may increase our knowledge of the processes which occur during NT and lead to greater efficiency of bovine cloning for commercial and therapeutic purposes.

text: Epigenetic reprogramming of somatic cells by zygotic factors Cloning cattle using somatic cell nuclear transfer (SCNT) is an inefficient process, with approximately only 5% of transferred embryos developing to live offspring. SCNT produced cattle have a high mortality rate due to a number of developmental abnormalities caused mainly by the incorrect epigenetic reprogramming of the donor cell to a pluripotent state. Conventional bovine SCNT involves fusion of a somatic donor cell into an enucleated metaphase II (MII) arrested oocyte. The resulting embryo is cultured to the blastocyst stage before being transferred to a surrogate cow to produce live offspring. Zygotes were initially thought to be unsuitable as SCNT recipients in bovine, until it was revealed that using telophase II (TII) zygotes as opposed to MII oocytes results in improved in vivo development. Metaphase zygotes have also been used successfully as SCNT recipients in mouse to produce cloned blastocysts and it has been proposed that reprogramming factors sequester in the pronuclei of interphase zygotes. Little is known about the nature the nuclear reprogramming, however a few candidate reprogramming factors have emerged recently. TCTP is known to activate key pluripotency genes (POU5F1 and NANOG) in somatic cell nuclei. It has also been identified as present in bovine oocytes with a high potential to reprogram somatic cells. Reprogramming of cell nuclei by Xenopus egg extract has found to require BRG1. Immunodepletion of BRG1 was shown to decrease the reprogramming ability of the egg extract, whist its over-expression increased reprogramming potential. HDAC1 has been found to initiate a transcriptionally repressive state in preimplantation mouse embryos possibly inhibiting transcription of reprogramming factors. Knockdown of HDAC1 using TSA is known to increase development of mouse NT embryos. 
The aim of this study was to produce blastocysts using metaphase zygotes as recipients for SCNT in bovine. In addition, localisation and abundance of candidate reprogramming factors TCTP (TPT1 gene), BRG1 and HDAC1 were examined in MII oocytes as well as TII, interphase and metaphase zygotes.

This study found metaphase zygotes are unsuitable as recipients for bovine SCNT using current methodologies, possibly due to the premature cleavage of the embryos. Control embryos produced using MII oocytes as recipients developed to blastocyst with an efficiency of ~ 11%. mRNA analysis of zygotes and oocytes did not reveal any significant differences in the relative concentrations of TPT1 or BRG1 between the samples. The TCTP and HDAC1 proteins showed a similar pattern of localisation in the MII oocytes and all stages of zygotes. Both proteins clearly localise to the maternal chromatin in the second polar body of TII zygotes. This finding has never previously been described and may in part explain why there is increased cloning efficiency observed when using TII zygotes as SCNT recipients.

This increased understanding of these reprogramming factors may increase our knowledge of the processes which occur during NT and lead to greater efficiency of bovine cloning for commercial and therapeutic purposes.

year: 2009

title: Hau

authors: Lodge, Martin

subjects: music

abstract: The article presents reflections on some issues encountered when combining traditional Maori and Western concert music. These issues include tradition and repertoire versus innovation, control versus freedom, collaboration versus appropriation, and the overlapping roles of creator and performer. Focal to the article is "Hau," a Maori word meaning breath or wind. The author also presents a comparison of Western music with traditional Maori instrumental music.

text: Hau The article presents reflections on some issues encountered when combining traditional Maori and Western concert music. These issues include tradition and repertoire versus innovation, control versus freedom, collaboration versus appropriation, and the overlapping roles of creator and performer. Focal to the article is "Hau," a Maori word meaning breath or wind. The author also presents a comparison of Western music with traditional Maori instrumental music.

year: 2007