Telomerase inactivation causes loss of the male germline in worms, fish, and mice, indicating a conserved dependence on telomere maintenance in this cell lineage. Here, using telomerase reverse transcriptase (Tert) reporter mice, we found that very high telomerase expression is a hallmark of undifferentiated spermatogonia, the mitotic population where germline stem cells reside. We exploited these high telomerase levels as a basis for purifying undifferentiated spermatogonia using fluorescence-activated cell sorting. Telomerase levels in undifferentiated spermatogonia and embryonic stem cells are comparable and much greater than in somatic progenitor compartments. Within the germline, we uncovered an unanticipated gradient of telomerase activity that also enables isolation of more mature populations. Transcriptomic comparisons of Tert High undifferentiated spermatogonia and Tert Low differentiated spermatogonia by RNA sequencing reveals marked differences in cell cycle and key molecular features of each compartment. Transplantation studies show that germline stem cell activity is confined to the Tert High cKit − population. Telomere shortening in telomerase knockout strains causes depletion of undifferentiated spermatogonia and eventual loss of all germ cells after undifferentiated spermatogonia drop below a critical threshold. These data reveal that high telomerase expression is a fundamental characteristic of germline stem cells, thus explaining the broad dependence on telomerase for germline immortality in metazoans.

SummaryUndifferentiated spermatogonia comprise a pool of stem cells and progenitor cells that show heterogeneous expression of markers, including the cell surface receptor GFRα1. Technical challenges in isolation of GFRα1+ versus GFRα1– undifferentiated spermatogonia have precluded the comparative molecular characterization of these subpopulations and their functional evaluation as stem cells. Here, we develop a method to purify these subpopulations by fluorescence-activated cell sorting and show that GFRα1+ and GFRα1– undifferentiated spermatogonia both demonstrate elevated transplantation activity, while differing principally in receptor tyrosine kinase signaling and cell cycle. We identify the cell surface molecule melanocyte cell adhesion molecule (MCAM) as differentially expressed in these populations and show that antibodies to MCAM allow isolation of highly enriched populations of GFRα1+ and GFRα1– spermatogonia from adult, wild-type mice. In germ cell culture, GFRα1– cells upregulate MCAM expression in response to glial cell line-derived neurotrophic factor (GDNF)/fibroblast growth factor (FGF) stimulation. In transplanted hosts, GFRα1– spermatogonia yield GFRα1+ spermatogonia and restore spermatogenesis, albeit at lower rates than their GFRα1+ counterparts. Together, these data provide support for a model of a stem cell pool in which the GFRα1+ and GFRα1– cells are closely related but show key cell-intrinsic differences and can interconvert between the two states based, in part, on access to niche factors.

PURPOSE Cervical cancer remains the eighth leading cause of cancer mortality among women in Taiwan. Despite availability of a National Health Insurance program with free yearly screenings, Papanicolaou testing and human papillomavirus (HPV) vaccination rates have been historically low in Taiwan. Even in the United States, cervical cancer screening rates for Asian women are significantly lower than other ethnic groups. The goal is to direct future interventions by providing insight on barriers leading to low screening and vaccination rates among Taiwanese and Asian women. METHODS Anonymous surveys without patient identifiers were randomly administered to patients at a traditional Chinese medicine clinic in Hualien City in June 2018. Inclusion criterion was females. No exclusion criteria were defined. Participants provided written consent. Sixty-three completed surveys were received. A χ2 test was used to determine statistical significance (α = 0.05). RESULTS Formal education level correlated with increased knowledge of HPV ( P = .001), its route of transmission ( P = .044), its link to genital cancer ( P = .0024), and HPV vaccination ( P = .0039). Women were more likely to have Papanicolaou smears if they were older than 30 years of age ( P = .0033), visited the gynecologist ( P < .001), or were recommended one by their physicians ( P < .001). Although 57% of respondents knew of the HPV vaccine, only 19% were vaccinated. Among those not vaccinated, most cited reasons included an inability to find a physician providing it (23.5%), safety concerns (16.4%), belief that it encourages sexual behavior (14.5%), and high out-of-pocket expense (9.1%). CONCLUSION Knowledge of HPV does not predict a higher adherence to cervical screening guidelines. Instead, diligent physician recommendations on Papanicolaou smears can elevate adherence rates among patients. Significant contributors to low HPV vaccination rates in rural Taiwan include lack of awareness and access to the vaccine. Our study emphasizes the physician-patient relationship as a means to target vulnerable populations and increase rates of cervical cancer screening and HPV vaccination.