The mammalian fallopian tube mucosa consists primarily of two cell types, ciliated cells and secretory cells. Cilia are usually classified into primary cilia and motile cilia according to the components of their axonemal microtubules (22). Motile cilia extend from the basal bodies in multi-ciliated cells of the ciliated epithelium in the fallopian tube. Ciliated cells occur throughout the fallopian tube but predominant at the apex of mucosal folds. Physiologically, the coordinated beating of multiple motile cilia is thought to provide the fluid flow needed to move an oocyte through the fallopian tube to the uterus. Multiple motile cilia have a canonical “9 + 2” ultrastructure of microtubules in the ciliary axoneme, consisting of 9 peripheral doublet microtubules surrounding 2 single central microtubules (23). In this study, we examined motile cilia of the tubal epithelia, which can be identified microscopically and immunohistochemically by tubulin stain.
Tubal ciliated cells are thought in general to represent terminally differentiated cells from either tubal secretory cells or stem-like cells within the fallopian tube and induced by estrogen stimulation (24–26). Thus, it is not surprising that there is a reduction of ciliated cells with age, in particular over age 40. In reality, tubal ciliated cells in vitro may reversely differentiate into non-ciliated cells (morphologically show no difference from the secretory cells), which is commonly observed in tubal primary culture experiments (Zheng et al., unpublished data). It is unclear how this happens and whether those ciliated cells have potential to initiate serous neoplasia after becoming non-ciliated cells. One early study about the role of tubal epithelia in supporting early embryo development showed that tubal motile cilia is lost in vitro when estrodial is withdrawn (24). But overall, there is limited understanding of the tubal ciliated cells on the aspects of cellular differentiation and neoplasia. As the tubal mucosa is mainly composed of ciliated and secretory cells, people may have a general impression that the increased density of tubal secretory cells reflects the decreased number of ciliated cells. In reality, such relationship has never been systemically studied.
In this study, we have examined the global change of tubal ciliated cells and its relationship with the aging process by counting the number of ciliated cells in patients with low-risk (benign group), high-risk, and O/PSC. Although the study is descriptive in nature, it explores a novel approach to evaluate the risk for ovarian or pelvic serous carcinogenesis. This is the first study to describe the change in number of tubal ciliated cells in relation to age. For patients in the low-risk (control group) for ovarian or pelvic serous cancers, the number of tubal ciliated cells decreased with age, starting at age 30 s and decreasing by 94% after age 80. It appears that the reduction of ciliated cells is consistent with the increment of tubal secretory cells with aging as we demonstrated earlier (2, 15). However, it is unclear if the reduction of tubal ciliated cells and increased the number of secretory cells are independent or reciprocal. Our observations of loss of tubal ciliated cells with aging also go along with the well-known epidemiologic findings that O/PSC increases with age and shows a peak incidence after menopause (27–29). A more dramatic decrease of tubal ciliated cells is observed in patients with high-risk factors, such as BRCA mutations or family history of ovarian cancer, and in patients with O/PSC. This decreased the number of tubal ciliated cells is also closely associated with age in both high-risk and O/PSC groups, indicating that decreased number of tubal ciliated cells is linked to “ovarian” or pelvic serous neoplasia.
Since decreased number of tubal ciliated cells was associated with age and more strikingly with high-risk as well as with O/PSC patients, we examined if the decreased number of ciliated cells in high-risk or O/PSC patients are independent of age in the study. By linear regression analysis, when both the cases and controls were combined, we found that all three factors namely age, high-risk status, and patients with O/PSC, are independent risk factors for the decreased number of tubal ciliated cells. Approximately 10–17% of decreased ciliated cells in the fallopian tube are attributed to age, while 83–90% attributed to O/PSC and high-risk status, respectively. Patients with BRCA mutations show the strongest association with decreased number of ciliated cells in the fallopian tube, consistent with its known risk for O/PSC development (28, 30–32). While there is still an association found of reduction of ciliated cells as well in the O/PSC group, this group is not restricted to cases with serous neoplasia. Thus, diverse tumor microenvironmental changes exist which could impact number of neighboring ciliated cells by other mechanisms. From these findings we speculate that patients with high-risk factors and/or with O/PSCs have an unidentified mechanism to cause reduction of tubal ciliated cells in addition to the aging process and such changes are not influenced by hormonal change since our study is age-matched.
Fallopian tube serves as the cellular source for the majority of O/PSC, while tubal fimbria is considered as the anatomical site of origin of these non-uterine high-grade serous cancers in women (33). Tubal ciliated cells are usually distributed evenly within the tubal fimbria and ampulla segments. In this study, we examined the number of ciliated cells and their distributions in both tubal segments. We found that there is no significant difference between the two tubal segments for the number of ciliated cells. However, the number of tubal ciliated cells are significantly less in high-risk group that that in O/PSC group within the tubal fimbria (Fig. 2A). It is unclear how to explain this phenomenon. But the findings are consistent with that patients with BRCA mutations develop HGSCs at an earlier age than those who develop sporadic cancer (34) as well as with the well accepted concept of tubal fimbria as the main anatomic site for ovarian or pelvic serous neoplasia (35, 36).
The functional role of cilia in human carcinogenesis is unclear. There are no studies on the role of multiple motile cilia for the tubal ciliated cells in the process of ovarian carcinogenesis. Through literature search, however, a few studies on the biologic function of primary cilia of the fallopian tube in the process of cancer development come to our attention (23, 37). Egeberg et al., suggested that defects of primary cilium in ovarian tumorigenesis may be related to deregulation of cilia signaling pathways such as Hedgehog, platelet-derived growth factor, aurora A kinase signaling (37). Some other earlier studies showed that primary cilia may play a critical role in tumorigenesis and cancer progression by functioning as a tumor suppressor organelle that regulates cell cycle/proliferation, differentiation, polarity, and migration (38, 39). On the other hand, loss of tubal ciliated cells may also reduce the capacity of removing follicular fluid induced genotoxicity within the fallopian tube (40). In one of our studies about ovarian serous carcinogenesis, we also notice that gradual motile cilia loss from serous cystadenoma to serous borderline tumor and finally complete loss of cilia in low-grade serous carcinoma (5). More recently, we have studied tubal epithelial cells from BRCA1 mutation carriers and benign controls without known history of BRCA1 mutation by using single cell sequencing technology. We found that tubal ciliated cells express SOX2 biomarker, which is known to be one of the stem cell markers, while secretory cells do not (manuscript in preparation). It would be interesting to study biologic function of tubal ciliated cells and their motile cilia in the process of serous carcinogenesis.
Tubal mucosa consists of both secretory and ciliated cells, arranged in a recurring pattern of alternating each other in normal fallopian tube of reproductive aged women. PAX-8 is a member of the pair-box (PAX) family of transcription factor genes. Studies have shown that PAX8 is a biomarker of tubal secretory cells and is used to distinguish gynecologic cancers from non-gynecologic malignancies (41). Therefore, PAX-8 has been widely used in the clinic. In this study, we used PAX-8 to distinguish tubal secretory cells from tubal ciliated cells. However, not infrequently we have found that not only secretory cells, but also some of the ciliated cells are positively stained by PAX-8 (Fig. 5). This is the reason for us to use tubulin highlighting the ciliated cells in the study. Although morphologically tubal ciliated cells are easily distinguished from secretory cells because of the presence of multi-motile cilia on the cellular apex, these two cell types may be interchangeable, which is supported by ciliation changes of the tubal epithelia in the menstrual cycle (42). That can explain why some of the ciliated cells are positive for PAX-8 expression in current study. Studies to identify regulatory factors for the transitions between ciliated and secretory cells are needed to help us uncover the role and functions of these tubal epithelial cells.