Background
Male germ cells are unique because they express a substantial number of variants of the general DNA binding proteins, known as histones, yet the biological significance of these variants ...is still unknown. In the present study, we aimed to address the expression pattern of the testis‐specific histone H2B variant (TH2B) and the testis‐specific histone H2A variant (TH2A) within the neonatal mouse testis.
Results
We demonstrate that TH2B and TH2A are present in a testis‐enriched for undifferentiated spermatogonia. Co‐localization studies with an undifferentiated marker, ZBTB16, revealed that TH2B and ZBTB16 co‐localize in the neonatal testis. Upon the appearance of the primary spermatocytes, TH2B no longer co‐localized with the ZBTB16 positive spermatogonia but were instead detected within the differentiating spermatogonia. This pattern of expression where TH2B and ZBTB16 no longer co‐localize was maintained in the adult testis.
Conclusion
These findings are in contrast to previous studies, which demonstrated that TH2B and TH2A were found only in adult spermatocytes. Our data are in support of a switch in the expression of these variants following the first round of spermatogonial differentiation. These studies reinforce current understandings that spermatogonia within the neonatal mouse testis are inherently different from those residing within the adult testis.
Key Findings
Contrary to previous beliefs, testis specific histone variants TH2B and TH2A are also expressed expressed in undifferentiated spermatogonia in the neonatal mouse testis. Upon the appearance of the primary spermatocytes, TH2B switches its expression from spermatogonia to the spermatocyte population. This study reinforces the idea that spermatogonia in the neonatal mouse testis is inherently different than those residing within the adult.
In the female reproductive tract, the innate immune system is modulated by two sex steroid hormones, estrogen and progesterone. A cyclical wave of neutrophils in the vaginal lumen is triggered by ...chemokines and correlates with circulating estrogen levels. Classical estrogen signaling in the female reproductive tract is activated through estrogen receptor α (encoded by the Esr1 gene). To study the role of estrogen action in the vagina, we used a mouse model in which Esr1 was conditionally ablated from the epithelial cells (Wnt7a
; Esr1
). Histological evidence showed that in response to a physical stress, the lack of ESR1 caused the vaginal epithelium to deteriorate due to the absence of a protective cornified layer and a reduction in keratin production. In the absence of ESR1 in the vaginal epithelial tissue, we also observed an excess of neutrophil infiltration, regardless of the estrous cycle stage. The histological presence of neutrophils was found to correlate with persistent enzymatic activity in the cervical-vaginal fluid. Together, these findings suggest that ESR1 activity in the vaginal epithelial cells is required to maintain proper structural integrity of the vagina and immune response, both of which are necessary for protecting the vagina against physical damage and resetting the vaginal environment.
Despite the essential role of the active metabolite of vitamin A, all-trans retinoic acid (atRA) in spermatogenesis, the enzymes, and cellular populations responsible for its synthesis in the ...postnatal testis remain largely unknown. The aldehyde dehydrogenase 1A (ALDH1A) family of enzymes residing within Sertoli cells is responsible for the synthesis of atRA, driving the first round of spermatogenesis. Those studies also revealed that the atRA required to drive subsequent rounds of spermatogenesis is possibly derived from the ALDH1A enzymes residing within the meiotic and post-meiotic germ cells. Three ALDH1A isozymes (ALDH1A1, ALDH1A2, and ALDH1A3) are present in the testis. Although, ALDH1A1 is expressed in adult Sertoli cells and is suggested to contribute to the atRA required for the pre-meiotic transitions, ALDH1A2 is proposed to be the essential isomer involved in testicular atRA biosynthesis. In this report, we first examine the requirement for ALDH1A2 via the generation and analysis of a conditional Aldh1a2 germ cell knockout and a tamoxifen-induced Aldh1a2 knockout model. We then utilized the pan-ALDH1A inhibitor (WIN 18446) to test the collective contribution of the ALDH1A enzymes to atRA biosynthesis following the first round of spermatogenesis. Collectively, our data provide the first in vivo evidence demonstrating that animals severely deficient in ALDH1A2 postnatally proceed normally through spermatogenesis. Our studies with a pan-ALDH1A inhibitor (WIN 18446) also suggest that an alternative source of atRA biosynthesis independent of the ALDH1A enzymes becomes available to maintain atRA levels for several spermatogenic cycles following an initial atRA injection. Summary Sentence Elimination of ALDH1A enzymatic activity following a single pulse of retinoic acid does not immediately ablate spermatogenesis due to the presence of an additional source of atRetinal oxidation.
The importance of all-trans-retinoic acid ( at-RA; hereafter referred to as RA) in normal sperm production has been recognized for many years. Consistent with this, an excess or deficiency of RA ...results in a range of male reproductive abnormalities. It has been widely postulated that the aldehyde dehydrogenase 1A (ALDH1A) family of enzymes, specifically the isozyme ALDH1A2 residing within the meiotic and post-meiotic germ cells, are responsible for RA synthesis after the first round of spermatogenesis. However, the studies presented within this dissertation demonstrate that the genetic elimination of ALDH1A2 is not sufficient to arrest spermatogenesis. Surprisingly the potent inhibition of the ALDH1A enzyme family (i.e. ALDH1A1, ALDH1A2, and ALDH1A3) after a single RA injection was not able to immediately arrest spermatogenesis. We identified the aldehyde oxidases as potential contributors to RA synthesis, thus compensating for the loss of ALDH1A enzymes within experimental testes. Collectively, these results suggest that an additional source of RA synthesis exists in the postnatal testis. Additionally, a large number of testis-specific histone variants have been identified, however little is known about the role that these variants play during male germ cell development. Unpublished studies in our laboratory suggest that two histone variants, Hist1h2b (TH2B) and Hist1h2a (TH2A), may be differentially expressed between the first round and subsequent rounds of spermatogenesis. To test this, we employed a unique double immunofluorescence protocol to illustrate that TH2B and TH2A are localized to the undifferentiated spermatogonia within the neonatal testis. However, once the first preleptotene spermatocytes appear, we identified a unique switch in the expression of TH2B from the undifferentiated spermatogonia to the spermatocyte population. These studies contribute to the growing body of evidence demonstrating that spermatogonia within the neonate are epigenetically distinct from spermatogonia residing within the adult.
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