Biomedical subjects

M R Avarbock

Publications and source records attributed to M R Avarbock.

25 records · Page 2Linked to original sources

PubMed1997

Transplantation of testis germinal cells into mouse seminiferous tubules.

In the adult male, germ cell differentiation takes place in the seminiferous tubules of the testis by a complex, highly organized and very efficient process. A population of diploid stem-cell spermatogonia that lie on the basement membrane of the tubule continuously undergoes self-renewal and produces progeny cells, which initiate the process of cellular differentiation to generate mature spermatozoa. Each testis contains many seminiferous tubules, which are connected at both ends to a collecting system called the rete testis. The mature spermatozoa pass from the tubules into the rete and are then carried through efferent ducts to the epididymis for final maturation before they are ready to fertilize an egg. In previous studies, we have demonstrated that donor testis cells collected from a fertile mouse are able to generate spermatogenesis when transplanted to the seminiferous tubules of an infertile male. The spermatozoa produced by the recipient from the donor-derived spermatogonial stem cells are able to fertilize eggs and produce progeny carrying the donor male haplotype. Furthermore, donor testis stem cells from a rat will generate normal rat spermatozoa following transplantation to a mouse testis. The spermatogonial transplantation technique is clearly valuable and applicable to many species, but it is difficult. Therefore, several procedures to introduce donor cells into the seminiferous tubules of a recipient have been developed using the mouse as a model, and they are described here in detail. The results indicate that microinjection of cell suspensions into the seminiferous tubules, efferent ducts or rete testis are equally effective in generating donor cell-derived spermatogenesis in recipients. Each approach is likely to be useful for different experimental purposes in a variety of species.

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PubMed1996

Rat spermatogenesis in mouse testis.

Recently, transplantation of mouse donor spermatogonial stem cells from a fertile testis to an infertile recipient mouse testis was described. The donor cells established spermatogenesis in the seminiferous tubules of the host, and normal spermatozoa were produced. In the most successful transplants, the recipient mice were fertile and sired up to 80 per cent of progeny from donor cells. Here we examine the feasibility of transplanting spermatogonial stem cells from other species to the mouse seminiferous tubule to generate spermatogenesis. Marked testis cells from transgenic rats were transplanted to the testes of immunodeficient mice, and in all of 10 recipient mice (in 19 of 20 testes), rat spermatogenesis occurred. Epididymides of eight mice were examined, and the three from mice with the longest transplants (> or = 110 days) contained rat spermatozoa with normal morphology. The generation of rat spermatogenesis in mouse testes suggests that spermatogonial stem cells of many species could be transplanted, and opens the possibility of xenogeneic spermatogenesis for other species.

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PubMed1996

Reconstitution of spermatogenesis from frozen spermatogonial stem cells.

Spermatozoa from a number of species can be cryopreserved and then subsequently used to fertilize eggs. However, this technique has several limitations. First, the freezing protocol varies for each species and must be determined empirically, and for some species appropriate methods have not yet been identified. Second, because these cells are fully differentiated, they will not undergo replication when thawed, and recombination of genetic information cannot occur. We now demonstrate, by using the recently developed spermatogonial transplantation technique, that male germline stem cells can be successfully cryopreserved. Donor testis cells isolated from prepubertal or adult mice and frozen from 4 to 156 days at -196 degrees C were able to generate spermatogenesis in recipient seminiferous tubules. Relatively standard preservation techniques were used, suggesting that male germ cells from other species can also be stored for long periods. Because transplanted testis stem cells will ultimately undergo replication and meiotic recombination during spermatogenesis, one might consider these preserved male germ lines as biologically immortal.

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PubMed1994

Germline transmission of donor haplotype following spermatogonial transplantation.

Spermatogenesis is a complex, highly organized, very efficient process that is based upon the capacity of stem cell spermatogonia simultaneously to undergo self-renewal and to provide progeny that differentiate into mature spermatozoa. We report here that testis-derived cells transplanted into the testis of an infertile mouse will colonize seminiferous tubules and initiate spermatogenesis in > 70% of recipients. Testis-derived cells from newborn mice were less effective in colonizing recipient testes than cells from 5- to 15- or 21- to 28-day-old mice. Increasing the number of Sertoli cells in the donor cell population did not increase the efficiency of colonization. Unmodified embryonic stem cells were not able to substitute for testis-derived cells in colonizing testes but instead formed tumors in syngeneic as well as nonsyngeneic hosts. Finally, with recipients that maintained endogenous spermatogenesis, testis cell transplantation yielded mice in which up to 80% of progeny were sired by donor-derived spermatozoa. The technique of spermatogonial cell transplantation should provide a means to generate germline modifications in a variety of species following development of spermatogonial culture techniques and should have additional applications in biology, medicine, and agriculture.

Age Factors↗

PubMed1991

Heterologous introns can enhance expression of transgenes in mice.

In a previous study we showed that genomic constructs were expressed more efficiently in transgenic mice than constructs that were identical except for the lack of introns. Using the mouse metallothionein promoter-rat growth hormone gene construct as a model, we show that the first intron of the rat growth hormone gene is essential for high-level expression, whereas the other three introns are less effective. Several heterologous introns placed 3' of the coding region of an intronless rat growth hormone gene are also ineffective. However, insertion of some heterologous introns between the metallothionein promoter and the growth hormone gene improves expression. To determine whether addition of heterologous introns would provide a general strategy for improving expression, we have tested them in conjunction with other intronless genes and with different promoters.

Animals↗

PubMed1989

Targeted correction of a major histocompatibility class II E alpha gene by DNA microinjected into mouse eggs.

DNA molecules containing the 5' end of a functional major histocompatibility class II E alpha gene were injected into mouse eggs bearing E alpha genes with 630-base-pair (bp) deletions encompassing the promoter and first exon. The deletion was corrected by homologous recombination in 1 of about 500 transgenic mice that incorporated the injected DNA. The corrected E alpha gene was transmitted to progeny, which were bred to homozygosity. Southern blot analysis, polymerase chain reaction amplification of the DNA spanning the deletion, and sequence analysis revealed that the corrected allele resembles the wild-type E alpha gene. At sites of single-base-pair polymorphisms, there was apparently random conversion to either the donor or recipient sequence. In addition, many point mutations were introduced. mRNAs were produced from the corrected allele in a tissue-specific manner, but their sizes were different from the wild-type allele, and they did not produce detectable E alpha protein. This experiment demonstrates the feasibility of targeting foreign DNA to a gene that is completely inactive in fertilized mouse eggs.

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PubMed1980

Translation of globin messenger RNA by the mouse ovum.

It has been demonstrated that the Xenopus oocyte can translate rabbit haemoglobin messenger RNA (mRNA) following microinjection of the message into the cell. The Xenopus oocyte has since been shown to be capable of translating a variety of messenger RNAs from different species. This system has proved useful in un-erstanding the mechanism of message translation and has also provided information about the translation capability of the Xenopus oocyte. Several other cell types, including HeLa cells and fibroblasts, can also translate exogenous message injected into the cell. However, there have been no reports of injection of mRNA into oocytes or fertilised one-cell ova of mammalian species. Nevertheless, the latter system could be of considerable use in studying the processing of exogenous messages in a mammalian system undergoing development, as well as providing insight into the way the early embryo processes injected messages and the protein products of such messages. We report here the results of injecting message into the fertilised one-cell mouse ovum and show that both mouse and rabbit globin mRNA are translated in this system.

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