RESUMO
The aim of this study was to investigate gonadal-protective properties of thalidomide in a mouse model of chemotherapy-induced ovarian failure. Administration of thalidomide before alkylating chemotherapy preserved ovarian follicles and lead to the resumption of normal estrous cyclicity.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Ovário/efeitos dos fármacos , Insuficiência Ovariana Primária/induzido quimicamente , Insuficiência Ovariana Primária/prevenção & controle , Talidomida/uso terapêutico , Inibidores da Angiogênese/administração & dosagem , Inibidores da Angiogênese/uso terapêutico , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Contagem de Células , Citoproteção/efeitos dos fármacos , Modelos Animais de Doenças , Regulação para Baixo , Avaliação Pré-Clínica de Medicamentos , Feminino , Gônadas/efeitos dos fármacos , Gônadas/patologia , Camundongos , Camundongos Endogâmicos C57BL , Ovário/patologia , Insuficiência Ovariana Primária/patologia , Talidomida/administração & dosagemRESUMO
BACKGROUND: The tetracycline-inducible gene regulation system is a powerful tool that allows temporal and dose-dependent regulation of target transgene expression in vitro and in vivo. Several tetracycline-inducible transgenic mouse models have been described with ubiquitous or tissue-specific expression of tetracycline-transactivator (tTA), reverse tetracycline-transactivator (rtTA) or Tet repressor (TetR). Here we describe a Tet-On transgenic rat that ubiquitously expresses rtTA-M2 driven by the murine ROSA 26 promoter. RESULTS: The homozygous rat line (ROSA-rtTA-M2) generated by lentiviral vector injection, has a single integration site and was derived from the offspring of a genetic mosaic founder with multiple transgene integrations. The rtTA-M2 transgene integrated into an intron of a putative gene on chromosome 2 and does not appear to affect the tissue-specificity or expression of that gene. Fibroblasts from the ROSA-rtTA-M2 rats were transduced with a TetO7/CMV-EGFP lentivirus and exhibited doxycycline dose-dependent expression of the EGFP reporter transgene, in vitro. In addition, doxycycline-inducible EGFP expression was observed, in vivo, when the TetO7/CMV-EGFP lentivirus was injected into testis, kidney and muscle tissues of ROSA-rtTA-M2 rats. CONCLUSIONS: This conditional expression rat model may have application for transgenic overexpression or knockdown studies of gene function in development, disease and gene therapy.
Assuntos
Técnicas Genéticas , Ratos Transgênicos , Ratos/genética , Animais , Linhagem Celular , Fibroblastos , Camundongos , Regiões Promotoras Genéticas , Tetraciclina , Transativadores/genéticaRESUMO
Spermatogonial stem cells (SSCs) are at the foundation of mammalian spermatogenesis. Whereas rare A(single) spermatogonia comprise the rodent SSC pool, primate spermatogenesis arises from more abundant A(dark) and A(pale) spermatogonia, and the identity of the stem cell is subject to debate. The fundamental differences between these models highlight the need to investigate the biology of primate SSCs, which have greater relevance to human physiology. The alkylating chemotherapeutic agent, busulfan, ablates spermatogenesis in rodents and causes infertility in humans. We treated adult rhesus macaques with busulfan to gain insights about its effects on SSCs and spermatogenesis. Busulfan treatment caused acute declines in testis volume and sperm counts, indicating a disruption of spermatogenesis. One year following high-dose busulfan treatment, sperm counts remained undetectable, and testes were depleted of germ cells. Similar to rodents, rhesus spermatogonia expressed markers of germ cells (VASA, DAZL) and stem/progenitor spermatogonia (PLZF and GFRalpha1), and cells expressing these markers were depleted following high-dose busulfan treatment. Furthermore, fresh or cryopreserved germ cells from normal rhesus testes produced colonies of spermatogonia, which persisted as chains on the basement membrane of mouse seminiferous tubules in the primate to nude mouse xenotransplant assay. In contrast, testis cells from animals that received high-dose busulfan produced no colonies. These studies provide basic information about rhesus SSC activity and the impact of busulfan on the stem cell pool. In addition, the germ cell-depleted testis model will enable autologous/homologous transplantation to study stem cell/niche interactions in nonhuman primate testes.