Generation of genetically modified mice by spermatozoa transfection in vivo: Preliminary results

Huguet, Elisabeth; Esponda, Pedro

doi:10.1002/(sici)1098-2795(200006)56:2+<243::aid-mrd6>3.0.co;2-c

Cited by 19 publications

(14 citation statements)

References 26 publications

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“…For example, 60-70% of sperm were reported to carry the transgene following injection of naked DNA into the vas deferens, (44) with a follow-up report claiming detection of the transgene in the cells of 7.5% of offspring animals produced following fertilisation with the transgene-bearing sperm. (105) Similar results were reported using liposome-encapsulated transgene molecules injected close to the epididymis. (48,49,65) Another recent development has been the combination of SMGT with microinjection (transgenICSI) in which sperm exposed to naked or liposome-encapsulated transgene molecules are microinjected into oocytes.…”

Section: Recent Technological Advances From Smgtsupporting

confidence: 70%

Sperm‐mediated gene transfer: Applications and implications

2005

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Recent developments in studies of sperm-mediated gene transfer (SMGT) now provide solid ground for the notion that sperm cells can act as vectors for exogenous genetic sequences. A substantive body of evidence indicates that SMGT is potentially useable in animal transgenesis, but also suggests that the final fate of the exogenous sequences transferred by sperm is not always predictable. The analysis of SMGT-derived offspring has shown the existence of integrated foreign sequences in some cases, while in others stable modifications of the genome are difficult to detect. The appearance of SMGT-derived modified offspring on the one hand and, on the other hand, the rarity of actual modification of the genome, suggest inheritance as extrachromosomal structures. Several specific factors have been identified that mediate distinct steps in SMGT. Among those, a prominent role is played by an endogenous reverse transcriptase of retrotransposon origin. Mature spermatozoa are naturally protected against the intrusion of foreign nucleic acid molecules; however, particular environmental conditions, such as those occurring during human assisted reproduction, can abolish this protection. The possibility that sperm cells under these conditions carry genetic sequences affecting the integrity or identity of the host genome should be critically considered. These considerations further suggest the possibility that SMGT events may occasionally take place in nature, with profound implications for evolutionary processes.

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Section: Recent Technological Advances From Smgtsupporting

confidence: 70%

Sperm‐mediated gene transfer: Applications and implications

2005

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“…Transfection techniques of spermatozoa are still in a preliminary stage [54] therefore our investigations on IP 3 accumulation have been limited to the use of putative PLC inhibitor U73122. The initial aim of this study was to block selectively the G i -mediated interaction of A 1 receptors with PLC.…”

Section: Discussionmentioning

confidence: 99%

Stimulation by N6-Cyclopentyladenosine of A1 Adenosine Receptors, Coupled to Gαi2 Protein Subunit, Has a Capacitative Effect on Human Spermatozoa1

Allegrucci

Liguori

Minelli

2001

Biology of Reproduction

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The effects of selective A(1) receptor agonist on human spermatozoa were examined to verify physiological responses and to investigate the signal transduction pathway. N6-Cyclopentyladenosine on uncapacitated spermatozoa did not induce spontaneous acrosome reaction after 5 h capacitation, whereas the number of capacitated spermatozoa, assessed by lysophosphatidylcholine-induced acrosome reaction with Pisum sativum agglutinin staining, was significantly increased. N6-Cyclopentyladenosine was also added to capacitated human spermatozoa to find out whether the agonist could induce the acrosome reaction. Results, although statistically significant, could not be considered biologically significant. A1-Mediated capacitation was followed by the increase of tyrosine phosphorylation of a protein subset ranging between M(r) = 200 000 and 30 000. Stimulation of A1 receptor with the selective agonist elicited an agonist-induced inositol phospholipid hydrolysis leading to a transient rise of inositol triphosphate (IP3). This increase was not induced by A(1) receptor antagonist and was blocked by phospholipase C inhibitor. Coimmunoprecipitation experiments showed that the A(1) receptor is coupled to Galphai2 subunit suggesting that the activation of phospholipase C is mediated by betagamma subunits. In conclusion, the A(1) adenosine receptor in human spermatozoa is coupled to Galphai2, signals via IP3, and affects the capacitative status of ejaculated spermatozoa.

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“…These male mice injected with exogenous DNA encoding green fluorescent protein (GFP) were mated with normal estrus females, and 7.5% newborns were found positive by PCR for exogenous DNA. In these positive animals, some tissues showed GFP expression (Huguet, 2000).…”

Section: Naked Dnamentioning

confidence: 96%