Limits...
Characterization of Nestin-positive stem Leydig cells as a potential source for the treatment of testicular Leydig cell dysfunction.

Jiang MH, Cai B, Tuo Y, Wang J, Zang ZJ, Tu X, Gao Y, Su Z, Li W, Li G, Zhang M, Jiao J, Wan Z, Deng C, Lahn BT, Xiang AP - Cell Res. (2014)

Bottom Line: We showed that these Nes-GFP+ cells expressed LIFR and PDGFR-α, but not LC lineage markers.We further observed that these cells were capable of clonogenic self-renewal and extensive proliferation in vitro and could differentiate into neural or mesenchymal cell lineages, as well as LCs, with the ability to produce testosterone, under defined conditions.In addition, we further demonstrated that CD51 might be a putative cell surface marker for SLCs, similar with Nestin.

View Article: PubMed Central - PubMed

Affiliation: 1] Cell-gene Therapy Translational Medicine Research Center, The Third Affiliated Hospital, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, Guangdong 510630, China [2] Key Laboratory for Stem Cells and Tissue Engineering, Center for Stem Cell Biology and Tissue Engineering, Ministry of Education, Sun Yat-sen University, Guangzhou, Guangdong 510080, China [3] Department of Anatomy, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, Guangdong 510080, China.

ABSTRACT
The ability to identify and isolate lineage-specific stem cells from adult tissues could facilitate cell replacement therapy. Leydig cells (LCs) are the primary source of androgen in the mammalian testis, and the prospective identification of stem Leydig cells (SLCs) may offer new opportunities for treating testosterone deficiency. Here, in a transgenic mouse model expressing GFP driven by the Nestin (Nes) promoter, we observed Nes-GFP+ cells located in the testicular interstitial compartment where SLCs normally reside. We showed that these Nes-GFP+ cells expressed LIFR and PDGFR-α, but not LC lineage markers. We further observed that these cells were capable of clonogenic self-renewal and extensive proliferation in vitro and could differentiate into neural or mesenchymal cell lineages, as well as LCs, with the ability to produce testosterone, under defined conditions. Moreover, when transplanted into the testes of LC-disrupted or aging models, the Nes-GFP+ cells colonized the interstitium and partially increased testosterone production, and then accelerated meiotic and post-meiotic germ cell recovery. In addition, we further demonstrated that CD51 might be a putative cell surface marker for SLCs, similar with Nestin. Taken together, these results suggest that Nes-GFP+ cells from the testis have the characteristics of SLCs, and our study would shed new light on developing stem cell replacement therapy for testosterone deficiency.

Show MeSH

Related in: MedlinePlus

Isolation and phenotypic analysis of Nes-GFP+ cells derived from the testis. (A) Flow cytometry was used to isolate Nes-GFP+ cells from the testes of Nes-GFP transgenic mice on postnatal days 7, 14, and 28. The representative histogram overlays showed the isotype controls (black region) and the stained samples (red lines). n = 5. (B, C) Representative flow cytometric profiles of Nes-GFP+ cells stained with LHR (B) and PDGFR-α (C); these cells were derived from the testes of postnatal day 7 Nes-GFP transgenic mice. FSC, Forward-scattered light; SSC, Side-scattered light; PSP, PDGFR-α single positive; DP, Nes-GFP and PDGFR-α double positive; DN, double negative; NSP, Nes-GFP+ single positive. (D) Phase-contrast micrographs of primary (P0, 4 days after plating) and P1 (7 days after plating) cells in each population derived from the PSP, DP, DN, and NSP cells. Scale bar, 50 μm. (E) Cytogenetic analysis showed that the Nes-GFP+ cells at P25 have a diploid karyotype.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4260348&req=5

fig2: Isolation and phenotypic analysis of Nes-GFP+ cells derived from the testis. (A) Flow cytometry was used to isolate Nes-GFP+ cells from the testes of Nes-GFP transgenic mice on postnatal days 7, 14, and 28. The representative histogram overlays showed the isotype controls (black region) and the stained samples (red lines). n = 5. (B, C) Representative flow cytometric profiles of Nes-GFP+ cells stained with LHR (B) and PDGFR-α (C); these cells were derived from the testes of postnatal day 7 Nes-GFP transgenic mice. FSC, Forward-scattered light; SSC, Side-scattered light; PSP, PDGFR-α single positive; DP, Nes-GFP and PDGFR-α double positive; DN, double negative; NSP, Nes-GFP+ single positive. (D) Phase-contrast micrographs of primary (P0, 4 days after plating) and P1 (7 days after plating) cells in each population derived from the PSP, DP, DN, and NSP cells. Scale bar, 50 μm. (E) Cytogenetic analysis showed that the Nes-GFP+ cells at P25 have a diploid karyotype.

Mentions: Histological data demonstrated that Nestin represents a potential marker for SLCs, suggesting the possibility of isolating SLCs on the basis of Nes-GFP expression. Towards this end, we sorted cells from the testes of postnatal Nes-GFP transgenic mice using fluorescence-activated cell sorting (FACS) and found that Nes-GFP+ cells constituted 52.1% ± 5.6%, 29.5% ± 6.3%, and 1.3% ± 0.4% of the total testicular interstitial cell population on postnatal days 7, 14, and 28, respectively. Consistent with the histological data the frequency of GFP+ cells decreased with age (Figure 2A).


Characterization of Nestin-positive stem Leydig cells as a potential source for the treatment of testicular Leydig cell dysfunction.

Jiang MH, Cai B, Tuo Y, Wang J, Zang ZJ, Tu X, Gao Y, Su Z, Li W, Li G, Zhang M, Jiao J, Wan Z, Deng C, Lahn BT, Xiang AP - Cell Res. (2014)

Isolation and phenotypic analysis of Nes-GFP+ cells derived from the testis. (A) Flow cytometry was used to isolate Nes-GFP+ cells from the testes of Nes-GFP transgenic mice on postnatal days 7, 14, and 28. The representative histogram overlays showed the isotype controls (black region) and the stained samples (red lines). n = 5. (B, C) Representative flow cytometric profiles of Nes-GFP+ cells stained with LHR (B) and PDGFR-α (C); these cells were derived from the testes of postnatal day 7 Nes-GFP transgenic mice. FSC, Forward-scattered light; SSC, Side-scattered light; PSP, PDGFR-α single positive; DP, Nes-GFP and PDGFR-α double positive; DN, double negative; NSP, Nes-GFP+ single positive. (D) Phase-contrast micrographs of primary (P0, 4 days after plating) and P1 (7 days after plating) cells in each population derived from the PSP, DP, DN, and NSP cells. Scale bar, 50 μm. (E) Cytogenetic analysis showed that the Nes-GFP+ cells at P25 have a diploid karyotype.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4260348&req=5

fig2: Isolation and phenotypic analysis of Nes-GFP+ cells derived from the testis. (A) Flow cytometry was used to isolate Nes-GFP+ cells from the testes of Nes-GFP transgenic mice on postnatal days 7, 14, and 28. The representative histogram overlays showed the isotype controls (black region) and the stained samples (red lines). n = 5. (B, C) Representative flow cytometric profiles of Nes-GFP+ cells stained with LHR (B) and PDGFR-α (C); these cells were derived from the testes of postnatal day 7 Nes-GFP transgenic mice. FSC, Forward-scattered light; SSC, Side-scattered light; PSP, PDGFR-α single positive; DP, Nes-GFP and PDGFR-α double positive; DN, double negative; NSP, Nes-GFP+ single positive. (D) Phase-contrast micrographs of primary (P0, 4 days after plating) and P1 (7 days after plating) cells in each population derived from the PSP, DP, DN, and NSP cells. Scale bar, 50 μm. (E) Cytogenetic analysis showed that the Nes-GFP+ cells at P25 have a diploid karyotype.
Mentions: Histological data demonstrated that Nestin represents a potential marker for SLCs, suggesting the possibility of isolating SLCs on the basis of Nes-GFP expression. Towards this end, we sorted cells from the testes of postnatal Nes-GFP transgenic mice using fluorescence-activated cell sorting (FACS) and found that Nes-GFP+ cells constituted 52.1% ± 5.6%, 29.5% ± 6.3%, and 1.3% ± 0.4% of the total testicular interstitial cell population on postnatal days 7, 14, and 28, respectively. Consistent with the histological data the frequency of GFP+ cells decreased with age (Figure 2A).

Bottom Line: We showed that these Nes-GFP+ cells expressed LIFR and PDGFR-α, but not LC lineage markers.We further observed that these cells were capable of clonogenic self-renewal and extensive proliferation in vitro and could differentiate into neural or mesenchymal cell lineages, as well as LCs, with the ability to produce testosterone, under defined conditions.In addition, we further demonstrated that CD51 might be a putative cell surface marker for SLCs, similar with Nestin.

View Article: PubMed Central - PubMed

Affiliation: 1] Cell-gene Therapy Translational Medicine Research Center, The Third Affiliated Hospital, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, Guangdong 510630, China [2] Key Laboratory for Stem Cells and Tissue Engineering, Center for Stem Cell Biology and Tissue Engineering, Ministry of Education, Sun Yat-sen University, Guangzhou, Guangdong 510080, China [3] Department of Anatomy, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, Guangdong 510080, China.

ABSTRACT
The ability to identify and isolate lineage-specific stem cells from adult tissues could facilitate cell replacement therapy. Leydig cells (LCs) are the primary source of androgen in the mammalian testis, and the prospective identification of stem Leydig cells (SLCs) may offer new opportunities for treating testosterone deficiency. Here, in a transgenic mouse model expressing GFP driven by the Nestin (Nes) promoter, we observed Nes-GFP+ cells located in the testicular interstitial compartment where SLCs normally reside. We showed that these Nes-GFP+ cells expressed LIFR and PDGFR-α, but not LC lineage markers. We further observed that these cells were capable of clonogenic self-renewal and extensive proliferation in vitro and could differentiate into neural or mesenchymal cell lineages, as well as LCs, with the ability to produce testosterone, under defined conditions. Moreover, when transplanted into the testes of LC-disrupted or aging models, the Nes-GFP+ cells colonized the interstitium and partially increased testosterone production, and then accelerated meiotic and post-meiotic germ cell recovery. In addition, we further demonstrated that CD51 might be a putative cell surface marker for SLCs, similar with Nestin. Taken together, these results suggest that Nes-GFP+ cells from the testis have the characteristics of SLCs, and our study would shed new light on developing stem cell replacement therapy for testosterone deficiency.

Show MeSH
Related in: MedlinePlus