Enhanced water and cryoprotectant permeability of porcine oocytes after artificial expression of human and zebrafish aquaporin-3 channels
- Authors
- Morató, R., Chauvigné, F., Novo, S., Bonet, S., and Cerdà, J.
- ID
- ZDB-PUB-140410-14
- Date
- 2014
- Source
- Molecular reproduction and development 81(5): 450-61 (Journal)
- Registered Authors
- Cerdà, Joan
- Keywords
- none
- MeSH Terms
-
- Animals
- Aquaporin 3/biosynthesis*
- Aquaporin 3/genetics
- Cryoprotective Agents/pharmacokinetics*
- Cryoprotective Agents/pharmacology
- Ethylene Glycol/pharmacokinetics*
- Ethylene Glycol/pharmacology
- Female
- Gene Expression*
- Humans
- Oocytes/cytology
- Oocytes/metabolism*
- Osmotic Pressure/drug effects
- Permeability
- Sucrose/pharmacokinetics
- Sucrose/pharmacology
- Sweetening Agents/pharmacokinetics
- Sweetening Agents/pharmacology
- Zebrafish/genetics*
- Zebrafish Proteins/biosynthesis*
- Zebrafish Proteins/genetics
- PubMed
- 24488947 Full text @ Mol. Reprod. Dev.
One of the major obstacles for the vitrification of mature porcine oocytes with ethylene glycol is their low permeability to this cryoprotectant, which results in osmotic stress-induced cell damage and low survival. Pig blastocysts, on the other hand, show enhanced water and cryoprotectant permeability, which has been related to the transcriptional activation of aquaporin-3 (AQP3) channels at this stage of development. In this study, we asked if expression of cRNAs encoding two aquaglyceroporins, human AQP3 (hAQP3) or the zebrafish Aqp3b-T85A mutant, in porcine oocytes can increase their permeability. Microinjection of germinal-vesicle-stage oocytes with enhanced green fluorescent protein (EGFP) or AQP3 cRNAs resulted in the expression of the corresponding proteins in <26% of the metaphase-II stage oocytes at 40–44 hr of in vitro culture; co-injection of EGFP cRNA appeared to be a suitable marker for oocyte selection since all EGFP-positive oocytes also expressed the corresponding aquaporin. Using this method, we found that mature oocytes co-expressing EGFP and hAQP3 or EGFP and Aqp3b-T85A showed approximately a twofold increase of the hydraulic conductivity (Lp) with respect non-injected or EGFP alone-injected oocytes in a 0.43 M sucrose or 1.3 M ethylene glycol solution, whereas the ethylene glycol permeability (PEG) of EGFP + hAQP3 and EGFP + Aqp3b-T85A oocytes was 6.7- and 12-fold higher, respectively, than control oocytes. These data demonstrate that the artificial expression of aquaglyceroporins in porcine metaphase-II oocytes improves their permeability, and that the zebrafish Aqp3b-T85A mutant is more efficient than the human channel at increasing the oocyte permeability to ethylene glycol..