RT Journal Article
T1 Recombinant rabbit beta nerve growth factor production and its biological effects on sperm and ovulation in rabbits
A1 Sánchez Rodríguez, Ana
A1 Abad, Paloma
A1 Arias Álvarez, María
A1 Millán Pastor, María Pilar
A1 García Rebollar, Pilar
A1 Bautista Santa Cruz, José Manuel
A1 Lorenzo González, Pedro Luis
A1 García García, Rosa María
AB In some induced-ovulating species, beta nerve growth factor (β-NGF) has important roles in ovulation, though data for rabbits are still inconclusive. In this study we first synthesized functional recombinant β-NGF from rabbit tissue (rrβ-NGF) to address the following objectives: 1) to compare rabbit β-NGF amino acid sequence with those of other induced- or spontaneous-ovulating species; 2) to assess the effects of rrβ-NGF on rabbit sperm viability and motility, and 3) to examine the in vivo ovulation inducing effect of rrβ-NGF added to the seminal dose in rabbit does. The NGF gene in rabbit prostate tissue was sequenced by Rapid Amplification of cDNA Ends and annotated in GenBank (KX528686). Recombinant rβ-NGF was produced in CHO cells and purified by affinity chromatography. Once confirmed by Western blotting and mass spectrometry (MALDI-TOF) that the amino acid sequence of the recombinant protein corresponded to β-NGF, its functionality was validated in PC12 cells in a successful dose-response study over 8 days. The amino acid sequence of prostate rabbit NGF differed to that of other species mainly in its receptor binding sites. In all the spontaneous ovulating species examined, compared with rabbit, alanine and proline residues, which interact with the high-affinity receptor, were replaced by a serine. In rabbits, asparagine and methionine were substituted by lysine at the low-affinity receptor binding site. In time- and dose-response experiments, the in vitro addition of rrβ-NGF to the ejaculate did not affect sperm viability whereas sperm motility parameters were enhanced by the addition of 1 μg/mL of the neuropeptide. Addition of this same concentration of rrβ-NGF to the seminal dose administered via the intravaginal route in does induced ovulation with a delayed LH peak, leading to a plasma progesterone increase, gestation and delivery. Our findings suggest that rrβ-NGF could be a useful option for biotechnological and reproduction assisted techniques in rabbits but further studies are needed.
PB Public Library of Science
SN 1932-6203
YR 2019
FD 2019-07-18
LK https://hdl.handle.net/20.500.14352/93717
UL https://hdl.handle.net/20.500.14352/93717
LA eng
NO Sanchez-Rodriguez A, Abad P, AriasAlvarez M, Rebollar PG, Bautista JM, Lorenzo PL, etal.(2019) Recombinant rabbit beta nerve growth factor production and its biological effects on sperm and ovulation in rabbits. PLoSONE 14(7): e0219780.https://doi.org/10.1371/journal. pone.0219780
NO Ministerio de Economía y Competitividad de España [subvención AGL2015-65572-C2]
NO Contrato Predoctoral UCM-Santander de ASR
NO Contrato de Empleo Joven de la Consejería de Educación, Juventud y Deporte de la Comunidad de Madrid y el Fondo Social Europeo de PA
DS Docta Complutense
RD 29 abr 2025