“…We also observed few floating germ cell clusters which reflect clonal expansion of progenitors. Similar germ cells clusters were also reported in adult mouse ovary [ 13 ]. Figure 2 Testicular cells from chemoablated mouse in culture.…”
Section: Resultssupporting
confidence: 87%
“…The underlying reasoning behind our belief that VSELs give rise to SSCs is the different OCT-4 staining pattern observed in the two cell types (nuclear vs cytoplasmic). Nuclear OCT-4 is a transcription factor required to maintain pluripotent state and when differentiation is initiated, OCT-4 is no longer required, shifts to cytoplasm and gets eventually degraded [ 13 - 15 ]. Thus in our present study we expected VSELs will always give rise to SSCs which will further differentiate, undergo meiosis and produce sperm.…”
Section: Resultsmentioning
confidence: 99%
“…Against this background, there is yet another novel PSCs population which exists in various adult organs [ 11 ] including gonads [ 12 , 13 ]. These are termed very small embryonic-like stem cells (VSELs) and have been well-characterized in mouse and human testicular tissue [ 11 , 14 , 15 ].…”
BackgroundExtensive research is ongoing to empower cancer survivors to have biological parenthood. For this, sperm are cryopreserved prior to therapy and in younger children testicular biopsies are cryopreserved with a hope to mature the germ cells into sperm later on for assisted reproduction. In addition, lot of hope was bestowed on pluripotent embryonic and induced pluripotent stem cells to differentiate into sperm and oocytes. However, obtaining functional gametes from pluripotent stem cells still remains a distant dream and major bottle-neck appears to be their inefficient differentiation into primordial germ cells (PGCs). There exists yet another population of pluripotent stem cells termed very small embryonic-like stem cells (VSELs) in adult body organs including gonads. We have earlier reported that busulphan (25 mg/Kg) treatment to 4 weeks old mice destroys actively dividing cells and sperm but VSELs survive and differentiate into sperm when a healthy niche is provided in vivo.MethodsMouse testicular VSELs that survived busulphan treatment were cultured for 3 weeks. A mix of surviving cells in seminiferous tubules (VSELs, possibly few spermatogonial stem cells and Sertoli cells) were cultured using Sertoli cells conditioned medium containing fetal bovine serum, follicle stimulating hormone and with no additional growth factors.ResultsStem cells underwent proliferation and clonal expansion in culture and spontaneously differentiated into sperm whereas Sertoli cells attached and provided a somatic support. Transcripts specific for various stages of spermatogenesis were up-regulated by qRT-PCR studies on day 7 suggesting VSELs (Sca1) and SSCs (Gfra) proliferate (Pcna), undergo spermatogenesis (spermatocyte specific marker prohibitin), meiosis (Scp3) and differentiate into sperm (post-meiotic marker protamine).ConclusionsProcess of spermatogenesis and spermiogenesis was replicated in vitro starting with testicular cells that survived busulphan treatment. We have earlier reported similar ability of ovarian VSELs enriched in the ovary surface epithelial cells to form oocyte-like structures in vitro. This striking potential of spontaneous differentiation of primitive testicular cells including VSELs that survive chemotherapy is being described for the first time in the present study.Electronic supplementary materialThe online version of this article (doi:10.1186/s12958-015-0031-2) contains supplementary material, which is available to authorized users.
“…We also observed few floating germ cell clusters which reflect clonal expansion of progenitors. Similar germ cells clusters were also reported in adult mouse ovary [ 13 ]. Figure 2 Testicular cells from chemoablated mouse in culture.…”
Section: Resultssupporting
confidence: 87%
“…The underlying reasoning behind our belief that VSELs give rise to SSCs is the different OCT-4 staining pattern observed in the two cell types (nuclear vs cytoplasmic). Nuclear OCT-4 is a transcription factor required to maintain pluripotent state and when differentiation is initiated, OCT-4 is no longer required, shifts to cytoplasm and gets eventually degraded [ 13 - 15 ]. Thus in our present study we expected VSELs will always give rise to SSCs which will further differentiate, undergo meiosis and produce sperm.…”
Section: Resultsmentioning
confidence: 99%
“…Against this background, there is yet another novel PSCs population which exists in various adult organs [ 11 ] including gonads [ 12 , 13 ]. These are termed very small embryonic-like stem cells (VSELs) and have been well-characterized in mouse and human testicular tissue [ 11 , 14 , 15 ].…”
BackgroundExtensive research is ongoing to empower cancer survivors to have biological parenthood. For this, sperm are cryopreserved prior to therapy and in younger children testicular biopsies are cryopreserved with a hope to mature the germ cells into sperm later on for assisted reproduction. In addition, lot of hope was bestowed on pluripotent embryonic and induced pluripotent stem cells to differentiate into sperm and oocytes. However, obtaining functional gametes from pluripotent stem cells still remains a distant dream and major bottle-neck appears to be their inefficient differentiation into primordial germ cells (PGCs). There exists yet another population of pluripotent stem cells termed very small embryonic-like stem cells (VSELs) in adult body organs including gonads. We have earlier reported that busulphan (25 mg/Kg) treatment to 4 weeks old mice destroys actively dividing cells and sperm but VSELs survive and differentiate into sperm when a healthy niche is provided in vivo.MethodsMouse testicular VSELs that survived busulphan treatment were cultured for 3 weeks. A mix of surviving cells in seminiferous tubules (VSELs, possibly few spermatogonial stem cells and Sertoli cells) were cultured using Sertoli cells conditioned medium containing fetal bovine serum, follicle stimulating hormone and with no additional growth factors.ResultsStem cells underwent proliferation and clonal expansion in culture and spontaneously differentiated into sperm whereas Sertoli cells attached and provided a somatic support. Transcripts specific for various stages of spermatogenesis were up-regulated by qRT-PCR studies on day 7 suggesting VSELs (Sca1) and SSCs (Gfra) proliferate (Pcna), undergo spermatogenesis (spermatocyte specific marker prohibitin), meiosis (Scp3) and differentiate into sperm (post-meiotic marker protamine).ConclusionsProcess of spermatogenesis and spermiogenesis was replicated in vitro starting with testicular cells that survived busulphan treatment. We have earlier reported similar ability of ovarian VSELs enriched in the ovary surface epithelial cells to form oocyte-like structures in vitro. This striking potential of spontaneous differentiation of primitive testicular cells including VSELs that survive chemotherapy is being described for the first time in the present study.Electronic supplementary materialThe online version of this article (doi:10.1186/s12958-015-0031-2) contains supplementary material, which is available to authorized users.
“…However, the role in vivo of these stem cells in adult mammalian ovaries and the intrinsic factors that regulate their biology are still unclear. Bhartiya and co-workers postulated that VSELs are the most primitive, pluripotent stem cells in ovary that give rise to committed tissue-specific progenitors including OSCs, expressing OCT-4 in cytoplasm in association to other germ cell markers [22]. It has been also reported that VSELs are relatively quiescent in vitro, in contrast with OSCs that rapidly undergo division and generate germ cell nests before their differentiation to oocytes.…”
Section: Differentiation Of Oscs In Oocytes: Hypothesis or Reality?mentioning
The existence of ovarian stem cells (OSCs) in women as well as their physiological role in post-menopausal age are disputed. However, accumulating evidence demonstrated that, besides the animal models including primarily mice, even in adult women putative OSCs obtained from ovarian cortex are capable to differentiate in vitro into oocyte-like cells (OLCs) expressing molecular markers typical of terminal stage of oogonial cell lineage. Recent studies describe that, similarly to mature oocytes, the OSC-derived OLCs also contain haploid karyotype. As proof of concept of their stem commitment, OSCs from mice differentiated to oocytes in vitro are suitable to be fertilized and implanted in sterilized animals resulting in embryo development. Despite enthusiasm for these data, which definitely require extended confirmation before considering potential application in humans for treatment of ovarian insufficiency, OSCs appear suitable for other clinical uses, restoring the endocrine derangements in premature ovarian failure or for fertility preservation in oncologic patients after anti-cancer treatments. In this context, the selection of viable oocytes generated from OSCs before chemotherapy protocols would overcome the potential adjunct oncogenic risk in women bearing hormone-dependent tumors who are repeatedly stimulated with high dose estrogens to induce oocyte maturation for their egg recruitment and cryopreservation.
“…We have reported relatively quiescent, pluripotent VSELs with nuclear OCT-4 in adult mammalian testis and ovary [ 9 , 10 ]. Besides VSELs, there exists another population of tissue specific progenitors derived from the VSELs which are slightly bigger in size, have cytoplasmic OCT-4 and are more active including spermatogonial stem cells (SSCs) in testis and ovarian germ stem cells (OGSCs) in ovary.…”
BackgroundEndometrium undergoes dramatic growth, breakdown and regeneration throughout reproductive period in mammals. Stem cells have been implicated in the process however their origin, nature, anatomical localization and characterization still remain obscure. Classical concept of presence of stem cells in the basal layer of endometrium was recently challenged when side population and label retaining cells were found to be distributed throughout endometrium. We have earlier reported very small embryonic-like stem cells (VSELs) in adult mammalian ovary and testis as a small population of cells with nuclear OCT-4 along with progenitors (spermatogonial stem cells and ovarian germ stem cells) with cytoplasmic OCT-4. Present study was undertaken to gauge presence of VSELs in bilaterally ovariectomized mouse uterus and their modulation by hormones.MethodsBilaterally ovariectomized mice were subjected to sequential estradiol and progesterone treatment in order to induce proliferation, differentiation and remodeling (regeneration). Stem cells were studied in tissue smears after H & E staining and after sorting using SCA-1 by immuno-localization and qRT-PCR studies (Oct-4A, Nanog and Sca-1). Flow cytometry studies were also undertaken to confirm the presence of VSELs in mouse uterus.ResultsTwo distinct populations of stem cells with dark stained nucleus and high nucleo-cytoplasmic ratio were detected in ovariectomized mouse uterus. These cells were sorted using SCA-1 and comprised smaller VSELs with nuclear expression of OCT-4 and slightly bigger, more abundant progenitors termed as endometrial stem cells (EnSCs) with cytoplasmic OCT-4. RT-PCR studies showed presence of pluripotent transcripts (Oct-4, Sca-1) and flow cytometry confirmed the presence of 0.069% of LIN-/CD45-/SCA-1+ VSELs. These stem cells were distinctly regulated during endometrial growth, differentiation and regeneration as evidenced by qRT-PCR results.ConclusionsVSELs are present in normal uterus and also under conditions of atrophy induced by bilateral ovariectomy. Marked increase in EnSCs is associated with endometrial growth and regeneration. Further studies are warranted to define the niche for these stem cells and whether EnSCs arising from the pluripotent VSELs are common progenitors for epithelial and stromal cells or not remains to be addressed. Results of the present study will help in better understanding of endometrial pathologies and their management in the future.Electronic supplementary materialThe online version of this article (doi:10.1186/s13048-015-0138-2) contains supplementary material, which is available to authorized users.
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