{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,18]],"date-time":"2026-09-18T09:44:59Z","timestamp":1789724699214,"version":"4.0.1"},"reference-count":32,"publisher":"Springer Science and Business Media LLC","issue":"1","content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Reprod Biol Endocrinol"],"published-print":{"date-parts":[[2012,12]]},"abstract":"<jats:title>Abstract<\/jats:title>\n          <jats:sec>\n            <jats:title>Background<\/jats:title>\n            <jats:p>Bovine <jats:italic>Herpesvirus<\/jats:italic> type-5 (BoHV-5) is a neurovirulent \u03b1-<jats:italic>Herpesvirus<\/jats:italic> which is potentially pathogenic for cows and suspected to be associated with reproductive disorders. Interestingly, natural transmission of BoHV-5 by contaminated semen was recently described in Australia. Additionally, BoHV-5 was also isolated from the semen of a healthy bull in the same country and incriminated in a natural outbreak of reproductive disease after artificial insemination. In contrast with BoHV-1, experimental exposure of in vitro produced bovine embryos to BoHV-5 does not affect embryo viability and seems to inhibit some pathways of apoptosis. However, the mechanisms responsible for these phenomena are poorly understood. In this study, we examined mitochondrial activity, antioxidant protection, stress response and developmental rates of in vitro produced bovine embryos that were exposed and unexposed to BoHV-5.<\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Methods<\/jats:title>\n            <jats:p>For this purpose, bovine embryos produced in vitro were assayed for cell markers after experimental infection of oocytes (n\u2009=\u200930; five repetitions), in vitro fertilization and development. The indirect immunofluorescence was employed to measure the expression of superoxide dismutase 1 (SOD1), anti-oxidant like protein 1 (AOP-1), heat shock protein 70.1 (Hsp 70.1) and also viral antigens in embryos derived from BoHV-5 exposed and unexposed oocytes. The determination of gene transcripts of mitochondrial activity (<jats:italic>SOD1<\/jats:italic>), antioxidant protection (<jats:italic>AOP-1<\/jats:italic>) and stress response (<jats:italic>Hsp70.1<\/jats:italic>) were evaluated using the reverse transcriptase polymerase chain reaction (RT-PCR). MitoTracker Green FM, JC-1 and Hoechst 33342-staining were used to evaluate mitochondrial distribution, segregation patterns and embryos morphology. The intensity of labeling was graded semi-quantitatively and embryos considered intensively marked were used for statistical analysis.<\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Results<\/jats:title>\n            <jats:p>The quality of the produced embryos was not affected by exposure to BoHV-5. Of the 357 collected oocytes, 313 (+\/\u2212 6.5; 87,7%) were cleaved and 195 (+\/\u2212 3.2; 54,6%) blastocysts were produced without virus exposure. After exposure, 388 oocytes were cleaved into 328 (+\/\u2212 8.9, 84,5%), and these embryos produced 193 (+\/\u2212 3.2, 49,7%) blastocysts. Viral DNA corresponding to the US9 gene was only detected in embryos at day 7 after in vitro culture, and confirmed by indirect immunofluorescence assay (IFA). These results revealed significant differences (<jats:italic>p<\/jats:italic>\u2009&lt;\u20090.05) between exposed and unexposed oocytes fertilized, as MitoTracker Green FM staining Fluorescence intensity of Jc-1 staining was significantly higher (<jats:italic>p<\/jats:italic>\u2009&lt;\u20090.005) among exposed embryos (143 +\/\u2212 8.2). There was no significant difference between the ratios of Hoechst 33342-stained nuclei and total cells in good-quality blastocysts (in both the exposed and unexposed groups). Using IFA and reverse transcriptase polymerase chain reaction (RT-PCR) for the set of target transcripts (<jats:italic>SOD1, AOP-1 and Hsp 70.1<\/jats:italic>), there were differences in the mRNA and respective proteins between the control and exposed embryos. Only the exposed embryos produced anti-oxidant protein-like 1 (<jats:italic>AOP-1<\/jats:italic>). However, neither the control nor the exposed embryos produced the heat shock protein <jats:italic>Hsp 70.1<\/jats:italic>. Interestingly, both the control and the exposed embryos produced superoxide dismutase (<jats:italic>SOD1<\/jats:italic>), revealing intense mitochondrial activity.<\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Conclusion<\/jats:title>\n            <jats:p>This is the first demonstration of <jats:italic>SOD1<\/jats:italic> and <jats:italic>AOP-1<\/jats:italic> production in bovine embryos exposed to BoHV-5. Intense mitochondrial activity was also observed during infection, and this occurred without interfering with the quality or number of produced embryos. These findings further our understanding on the ability of \u03b1-<jats:italic>Herpesviruses<\/jats:italic> to prevent apoptosis by modulating mitochondrial pathways.<\/jats:p>\n          <\/jats:sec>","DOI":"10.1186\/1477-7827-10-53","type":"journal-article","created":{"date-parts":[[2012,7,23]],"date-time":"2012-07-23T12:14:59Z","timestamp":1343045699000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":11,"title":["Evaluation of developmental changes in bovine in vitro produced embryos following exposure to bovine Herpesvirus type 5"],"prefix":"10.1186","volume":"10","author":[{"given":"Mariana PC","family":"Brenner","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Camila","family":"Silva-Frade","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marina C","family":"Ferrarezi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrea F","family":"Garcia","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Eduardo F","family":"Flores","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tereza C","family":"Cardoso","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2012,7,23]]},"reference":[{"key":"991_CR1","first-page":"103","volume-title":"Manual of the International Embryo Transfer Society Volume 3","author":"A Robertson","year":"1998","unstructured":"Robertson A, Nelson RE: Certification and identification of the embryo. Manual of the International Embryo Transfer Society Volume 3. Edited by: Stringfellow DA, Seidel SM, Savoy IL. 1998, 116, 116, 103-116."},{"key":"991_CR2","doi-asserted-by":"publisher","first-page":"584","DOI":"10.1016\/j.theriogenology.2009.04.015","volume":"72","author":"WJ Jeong","year":"2009","unstructured":"Jeong WJ, Cho SJ, Lee HS, Deb GK, Lee YS, Kwon TH, Kong IK: Effect of cytoplasmic lipid content on in vitro developmental efficiency of bovine IVP embryos. Theriogenology. 2009, 72: 584-589. 10.1016\/j.theriogenology.2009.04.015.","journal-title":"Theriogenology"},{"key":"991_CR3","doi-asserted-by":"publisher","first-page":"242","DOI":"10.1210\/en.136.1.242","volume":"136","author":"JL Tilly","year":"1995","unstructured":"Tilly JL, Tilly KI: Inhibitors of oxidative stress mimic the ability of follicle stimulating bovine to suppress apoptosis in cultured rat ovarian follicles. Endocrinology. 1995, 136: 242-252. 10.1210\/en.136.1.242.","journal-title":"Endocrinology"},{"key":"991_CR4","doi-asserted-by":"publisher","first-page":"132","DOI":"10.1016\/j.anireprosci.2007.02.002","volume":"104","author":"AG Corr\u00eaa","year":"2008","unstructured":"Corr\u00eaa AG, Rumpf R, Mundim TCD, Franco MM, Dode MAN: Oxygen tension during in vitro culture of bovine embryos: effect in production and expression of genes related to oxidative stress. Anim Reprod Sci. 2008, 104: 132-142. 10.1016\/j.anireprosci.2007.02.002.","journal-title":"Anim Reprod Sci"},{"key":"991_CR5","doi-asserted-by":"publisher","first-page":"265","DOI":"10.1016\/j.theriogenology.2007.05.044","volume":"68","author":"S Balasubramanian","year":"2007","unstructured":"Balasubramanian S, Son WJ, Kumar BM, Ock SA, Yoo JG, Im GS, Choe SY, Rho GJ: Expression pattern of oxygen and stress-responsive gene transcripts at various developmental stages of in vitro and in vivo preimplantation bovine embryos. Theriogenology. 2007, 68: 265-275. 10.1016\/j.theriogenology.2007.05.044.","journal-title":"Theriogenology"},{"key":"991_CR6","doi-asserted-by":"publisher","first-page":"173","DOI":"10.1016\/j.anireprosci.2008.02.014","volume":"111","author":"TM Rodina","year":"2009","unstructured":"Rodina TM, Cooke FNT, Hansen PJ, Ealy AD: Oxygen tension and medium type actions on blastocyst development and interferon-tau secretion in cattle. Anim Reprod Sci. 2009, 111: 173-188. 10.1016\/j.anireprosci.2008.02.014.","journal-title":"Anim Reprod Sci"},{"key":"991_CR7","doi-asserted-by":"publisher","first-page":"316","DOI":"10.1016\/j.mito.2010.02.005","volume":"10","author":"I Scott","year":"2010","unstructured":"Scott I: The role of mitochondria in the mammalian antiviral defense system. Mitochondrion. 2010, 10: 316-320. 10.1016\/j.mito.2010.02.005.","journal-title":"Mitochondrion"},{"key":"991_CR8","doi-asserted-by":"publisher","first-page":"297","DOI":"10.1017\/S0967199411000050","volume":"19","author":"M Crocco","year":"2011","unstructured":"Crocco M, Alberio RH, Lauria L, Mariano MI: Effect of serum on the mitochondrial active area on developmental days 1 to 4 in in vitro produced bovine embryos. Zygote. 2011, 19: 297-306. 10.1017\/S0967199411000050.","journal-title":"Zygote"},{"key":"991_CR9","doi-asserted-by":"publisher","first-page":"5","DOI":"10.1111\/j.1439-0531.2006.00615.x","volume":"41","author":"AM Tarazona","year":"2006","unstructured":"Tarazona AM, Rodriguez JI, Restrepo LF, Olivera-Angel M: Mitochondrial activity, distribution and segregation in bovine oocytes and in embryos produced in vitro. Reprod Dom Anim. 2006, 41: 5-11. 10.1111\/j.1439-0531.2006.00615.x.","journal-title":"Reprod Dom Anim"},{"key":"991_CR10","first-page":"201","volume":"107","author":"M Paroli","year":"2000","unstructured":"Paroli M, Schiafella E, Di Rosa F, Barnaba V: Persisting viruses and autoimmunity. J Neuroimmunol. 2000, 107: 201-204.","journal-title":"Persisting viruses and autoimmunity. J Neuroimmunol"},{"key":"991_CR11","doi-asserted-by":"publisher","first-page":"324","DOI":"10.1016\/j.theriogenology.2009.09.016","volume":"73","author":"C Silva-Frade","year":"2010","unstructured":"Silva-Frade C, Martins J\u00fanior A, Borsanelli AC, Cardoso TC: Effects of bovine Herpesvirus type 5 on development of in vitro produced bovine embryos. Theriogenology. 2010, 73: 324-331. 10.1016\/j.theriogenology.2009.09.016.","journal-title":"Theriogenology"},{"key":"991_CR12","doi-asserted-by":"publisher","first-page":"1296","DOI":"10.1016\/j.theriogenology.2010.06.001","volume":"74","author":"C Silva-Frade","year":"2010","unstructured":"Silva-Frade C, Gameiro R, Martins A, Cardoso TC: Apoptotic and developmental effects of bovine Herpesvirus type-5 infection on in vitro-produced bovine embryos. Theriogenology. 2010, 74: 1296-1303. 10.1016\/j.theriogenology.2010.06.001.","journal-title":"Theriogenology"},{"key":"991_CR13","doi-asserted-by":"publisher","first-page":"254","DOI":"10.1016\/j.vetmic.2009.10.016","volume":"142","author":"APM Varela","year":"2010","unstructured":"Varela APM, Holz CL, Cibulski SP, Teixeira TF, Antunes AC, Franco AC, Roehe LR, Oliveira MT, Campos FS, Dezen D, Cenci A, Brito WD, Roehe PM: Neutralizing antibodies to bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) and its subtypes. Vet Microbiol. 2010, 142: 254-260. 10.1016\/j.vetmic.2009.10.016.","journal-title":"Vet Microbiol"},{"key":"991_CR14","doi-asserted-by":"publisher","first-page":"8","DOI":"10.1186\/1746-6148-7-8","volume":"7","author":"SS Maidana","year":"2011","unstructured":"Maidana SS, Ladelfa MF, P\u00e9rez SE, Lom\u00f3naco PM, Del M\u00e9dico Zajac MP, Ode\u00f3n A, Blanco Viera J, Combessies G, Fondevila N, Palacios M, Thiry J, Muylkens B, Thiry E, Romera AS: Characterization of BoHV-5 field strains circulation and report of transient specific subtype of bovine herpesvirus 5 in Argentina. BMC Vet Res. 2011, 7: 8-10.1186\/1746-6148-7-8.","journal-title":"BMC Vet Res"},{"key":"991_CR15","doi-asserted-by":"publisher","first-page":"509","DOI":"10.1016\/j.jviromet.2009.11.013","volume":"163","author":"TC Cardoso","year":"2010","unstructured":"Cardoso TC, Gomes DE, Ferrari HF, Silva-Frade C, Rosa ACG, Andrade AL, Luvizotto MCR: A novel in situ polymerase chain reaction hybridisation assay for the direct detection of bovine herpesvirus type 5 in formalin-fixed, paraffin-embedded tissues. J Virol Meth. 2010, 163: 509-512. 10.1016\/j.jviromet.2009.11.013.","journal-title":"J Virol Meth"},{"key":"991_CR16","doi-asserted-by":"publisher","first-page":"1139","DOI":"10.1016\/j.theriogenology.2010.11.025","volume":"75","author":"MT Oliveira","year":"2011","unstructured":"Oliveira MT, Campos FS, Dias MM, Velho FA, Freneau GE, Brito WM, Rijsewijk FA, Franco AC, Roehe PM: Detection of bovine herpesvirus 1 and 5 in semen from Brazilian bulls. Theriogenology. 2011, 75: 1139-1145. 10.1016\/j.theriogenology.2010.11.025.","journal-title":"Theriogenology"},{"key":"991_CR17","doi-asserted-by":"publisher","first-page":"247","DOI":"10.1016\/j.theriogenology.2005.05.043","volume":"65","author":"AE Warthall","year":"2006","unstructured":"Warthall AE, Simmons HA, Van Soom A: Evaluation of risks of viral transmission to recipients of bovine embryos arising from fertilization with virus-infected semen. Theriogenology. 2006, 65: 247-274. 10.1016\/j.theriogenology.2005.05.043.","journal-title":"Theriogenology"},{"key":"991_CR18","doi-asserted-by":"publisher","first-page":"111","DOI":"10.1136\/vetrec.165.4.111","volume":"165","author":"PD Kirkland","year":"2009","unstructured":"Kirkland PD, Poynting AJ, Gu X, Davis RJ: Infertility and venereal disease in cattle inseminated with semen containing bovine Herpesvirus type 5. Vet Rec. 2009, 165: 111-113. 10.1136\/vetrec.165.4.111.","journal-title":"Vet Rec"},{"key":"991_CR19","doi-asserted-by":"publisher","first-page":"93","DOI":"10.1111\/j.1751-0813.2009.00545.x","volume":"88","author":"IS Diallo","year":"2010","unstructured":"Diallo IS, Hewitson GR, Hoad J, Turner S, Corney BG, Rodwell BJ: Isolation of bovine herpesvirus type 5 from the semen of a healthy bull in Australia. Aust Vet J. 2010, 88: 93-95. 10.1111\/j.1751-0813.2009.00545.x.","journal-title":"Aust Vet J"},{"key":"991_CR20","doi-asserted-by":"publisher","first-page":"55","DOI":"10.1016\/j.anireprosci.2011.01.004","volume":"124","author":"DG Souza","year":"2011","unstructured":"Souza DG, Silva-Frade C, Martins A: Cardoso TC: Bull semen variables after experimental exposure with Bovine Herpesvirus type 5. Anim Reprod Sci. 2011, 124: 55-60. 10.1016\/j.anireprosci.2011.01.004.","journal-title":"Anim Reprod Sci"},{"key":"991_CR21","volume-title":"Cell Death Dis","author":"LM Tiede","year":"2011","unstructured":"Tiede LM, Cook EA, Morsey B, Fox HS: Oxygen matters: tissue culture oxygen levels affect mitochondrial function and structure as well as responses to HIV viroproteins. Cell Death Dis. 2011, 2: 10.1038\/cddis.2011. Macmillan Publishers limited"},{"key":"991_CR22","doi-asserted-by":"publisher","first-page":"e246","DOI":"10.1038\/cddis.2011.128","volume":"2","author":"LM Tiede","year":"2011","unstructured":"Tiede LM, Cook EA, Morsey B, Fox HS: Oxygen matters: tissue culture oxygen levels affect mitochondrial function and structure as well as responses to HIV viroproteins. Cell Death Dis. 2011, 2: e246-10.1038\/cddis.2011.128.","journal-title":"Cell Death Dis"},{"key":"991_CR23","doi-asserted-by":"publisher","first-page":"1547","DOI":"10.1099\/0022-1317-83-7-1547","volume":"83","author":"S Hay","year":"2002","unstructured":"Hay S, Kannourakis G: A time to kill: viral manipulation of the cell death program. J Gen Virol. 2002, 83: 1547-1564.","journal-title":"J Gen Virol"},{"key":"991_CR24","doi-asserted-by":"publisher","first-page":"46","DOI":"10.1016\/j.jviromet.2011.04.013","volume":"175","author":"IS Diallo","year":"2011","unstructured":"Diallo IS, Corney BG, Rodwell BJ: Detection and differentiation of bovine herpesvirus 1 and 5 using a multiplex real-time polymerase chain reaction. J Virol Meth. 2011, 175: 46-52. 10.1016\/j.jviromet.2011.04.013.","journal-title":"J Virol Meth"},{"key":"991_CR25","doi-asserted-by":"publisher","first-page":"19","DOI":"10.1186\/1746-6148-7-19","volume":"7","author":"MP Del Medico Zajac","year":"2011","unstructured":"Del Medico Zajac MP, Romera SA, Ladelfa MF, Kotsias F, Delgado F, Thiry J, Meurens F, Keil G, Thiry E, Muylkens B: In vitro-generated interspecific recombinants between bovine herpesviruses 1 and 5 show attenuated replication characteristics and establish latency in the natural host. BMC Vet Res. 2011, 7: 19-10.1186\/1746-6148-7-19.","journal-title":"BMC Vet Res"},{"key":"991_CR26","doi-asserted-by":"publisher","first-page":"335","DOI":"10.1016\/j.jviromet.2007.07.018","volume":"143","author":"HF Ferrari","year":"2007","unstructured":"Ferrari HF, Luvizotto MCR, Rahal P, Cardoso TC: Detection of bovineHespesvirus type5 in formalin-fixed, paraffin-embedded bovine brain by PCR: a useful adjunct to conventional tissue based diagnostic test of bovine encephalitis. J Virol Meth. 2007, 143: 335-340.","journal-title":"J Virol Meth"},{"key":"991_CR27","doi-asserted-by":"publisher","first-page":"379","DOI":"10.1007\/s11250-011-9929-7","volume":"44","author":"TC Cardoso","year":"2011","unstructured":"Cardoso TC, Antello TF, Vieira FV, Machado ST, de Campos Nogueira AH: Estimation of the diagnostic accuracy of the glyco-C and US9 gene-based polymerase chain reaction technique for the detection of bovine Herpesvirus type 5 DNA in decomposed brain suspension from a slaughter house using Bayesian analysis, Brazil. Trop Anim Health Prod. 2011, 44: 379-382.","journal-title":"Trop Anim Health Prod"},{"key":"991_CR28","doi-asserted-by":"publisher","first-page":"89","DOI":"10.1016\/j.vetmic.2010.02.017","volume":"143","author":"BD Griffin","year":"2010","unstructured":"Griffin BD, Verweij MC, Wiertz EJHJ: Herpesvirus and immunity: art of invasion. Vet Microbiol. 2010, 143: 89-100. 10.1016\/j.vetmic.2010.02.017.","journal-title":"Vet Microbiol"},{"key":"991_CR29","doi-asserted-by":"publisher","first-page":"527","DOI":"10.1071\/RD10268","volume":"23","author":"P Celi","year":"2011","unstructured":"Celi P, Merlo M, Da Dalt L, Stefani A, Barbato O, Gabai G: Relationship between late embryonic mortality and the increase in plasma advanced oxidased protein products (AOPP) in dairy cows. Reprod Fert Develop. 2011, 23: 527-533. 10.1071\/RD10268.","journal-title":"Reprod Fert Develop"},{"key":"991_CR30","doi-asserted-by":"publisher","first-page":"41","DOI":"10.5603\/FHC.2011.0007","volume":"49","author":"LC Bregano","year":"2011","unstructured":"Bregano LC, Agostinho SD, Roncatti FLBT, Pires MC, Riva HG, Luvizotto MCR, Cardoso TC: Immunohistochemical detection of metalloproteinase-9 (MMP-9), anti-oxidant like 1 protein (AOP-1) and synaptosomal-associated protein (SNAP-25) in the cerebella of dogs naturally infected with spontaneous canine distemper. Folia Histochem Cytobiol. 2011, 49: 41-48.","journal-title":"Folia Histochem Cytobiol"},{"key":"991_CR31","doi-asserted-by":"publisher","first-page":"57","DOI":"10.1186\/1746-1596-5-57","volume":"5","author":"TC Cardoso","year":"2010","unstructured":"Cardoso TC, Ferrari HF, Garcia AF, Bregano LC, Andrade AL, Nogueira AH: Immunohistochemical approach to the pathogenesis of clinical cases of bovine Herpesvirus type 5 infections. Diagn Pathol. 2010, 5: 57-10.1186\/1746-1596-5-57.","journal-title":"Diagn Pathol"},{"key":"991_CR32","first-page":"97","volume":"131","author":"M Derakhshan","year":"2010","unstructured":"Derakhshan M: Effect of measles virus (MV) on mitochondrial respiration. Indian J Med Res. 2010, 131: 97-100.","journal-title":"Indian J Med Res"}],"container-title":["Reproductive Biology and Endocrinology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/1477-7827-10-53.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,9,1]],"date-time":"2021-09-01T19:43:02Z","timestamp":1630525382000},"score":1,"resource":{"primary":{"URL":"https:\/\/rbej.biomedcentral.com\/articles\/10.1186\/1477-7827-10-53"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2012,7,23]]},"references-count":32,"journal-issue":{"issue":"1","published-print":{"date-parts":[[2012,12]]}},"alternative-id":["991"],"URL":"https:\/\/doi.org\/10.1186\/1477-7827-10-53","relation":{},"ISSN":["1477-7827"],"issn-type":[{"value":"1477-7827","type":"electronic"}],"subject":[],"published":{"date-parts":[[2012,7,23]]},"assertion":[{"value":"12 April 2012","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"31 May 2012","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"23 July 2012","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}}],"article-number":"53"}}