Contributed by John A Ozolek, MD, Ronald Jaffe, MB, BCh, and Maria Z Parizhskaya, MD FINAL DIAGNOSIS:
DISCUSSION:
Case 1 is a 15 year-old girl who presented for evaluation of primary amenorrhea and delayed puberty. Results of this work-up revealed a 46 XY genotype. Pelvic ultrasound revealed a normal vagina, cervix, uterus and fallopian tubes, and absent left adnexa and a right adnexal structure (ovary versus ovotestis). Laboratory evaluation included a markedly low estradiol level of less than 10 picograms per milliliter, and elevated luteinizing hormone and follicle stimulating hormone levels (48.1 milli-international units per milliliter and 32.4 mill-international units per milliliter, respectively). A 1.3 x 1.3 x 1.0 centimeter finely granular pink-tan tissue was submitted for pathological examination. Sections revealed remnants of ovarian stroma and capsule without ova. The remainder of the tissue was replaced by tumor composed of sharply circumscribed nodules composed of small, dark cells interspersed with larger, paler cells often centered around hyaline bodies. Accompanying this morphology were sheets of large pale cells (dysgerminoma or seminomatous like) with interspersed giant cells and epithelioid granulomatous reaction. Many of the cells in the latter population were positive for placental alkaline phosphatase (PLAP), while cells within the granulomatous areas were CD68 positive.
Gonadoblastoma is classified as a mixed germ cell-sex cord-stromal tumor and is rare even in the pediatric age group. This tumor is usually found in patients with an underlying gonadal disorder; pure or mixed gonadal dysgenesis and may exhibit varied karyotypes including 46XY or 45X, 46XY (1) Approximately 12% of individuals with Turner syndrome have Y chromosome material, but the incidence of gonadoblastoma in these individuals is about 7-10% (2). These tumors can also arise rarely in apparently normal women with a history of pregnancy (3). Histologic hallmarks are nests containing germ cells (larger cells with clear cytoplasm) and stromal cells of granulosa-Sertoli type. Dysgerminoma in a patient with dysgenetic gonads and a Y chromosome should prompt search for gonadoblastoma. Foci of calcification and/or a small nest of typical gonadoblastoma may be the only histologic evidence of previous gonadoblastoma. Pure gonadoblastomas are benign, but are frequently associated with malignant germ cell tumors. The differential diagnosis includes a pure dysgerminoma, sex cord tumor with annular tubules (SCTAT), unclassified germ cell-sex-cord-stromal tumors, and normal fetal and neonatal ovary (4).
Case 2 is a 13-year-old girl with an immature teratoma with yolk sac carcinoma. Portions of this tumor histologically demonstrated mature elements of skin, bone and mature neural tissue including choroid plexus, neuroglia and cerebellar tissue. The immature components (demonstrated here) consisted of a highly cellular neuroglia that in areas formed neuroepithelial tubes and rosettes as described. Yolk sac carcinoma was seen in some areas directly contiguous with immature neural elements.
Immature teratomas are the third most common germ cell tumor in the adolescent ovary. They are graded histologically by the amount of immature neuroectoderm present (Grades 1, 2, and 3). In the pediatric age group, one must search carefully for a malignant component typically endodermal sinus or yolk sac tumor. As in the demonstrated case here, these foci can be small, blend imperceptibly with immature neuroectoderm, and take a variety of histologic patterns as is well described for yolk sac tumors. Our case here demonstrated cystic, glandular and endometrioid patterns (4).
A recent paper in Science Online demonstrates the ability to differentiate mouse embryonic stem cells (ES cells) into oocytes and possibly trophoblastic cells in vitro (5). These were accomplished with both male and female ES cells presumably outside of the influence of SRY expression. This finding is of paramount interest since a priori cultured ES cells were not thought to be totipotent (able to contribute to all tissues of the embryo including extraembryonic tissue), but pluripotent (able to differentiate into the majority of somatic embryonic tissue excluding the germline and extraembryonic tissue). This finding is exciting since little is known about the direct signals that allow early ES cells to differentiate into not only somatic tissue but especially germline tissue. Knowledge of the precise molecular interactions and signaling events of differentiation from ES cells will allow great insights into disease mechanism, phenotype and phenotypic variations in syndromic conditions, tissue repair and regeneration, and infertility, all areas of interest and application in Pediatrics and Pediatric Pathology.
These cases are illustrations of the fine and yet unknown molecular balance between normal and abnormal and potentially malignant embryological development. These cases, while unusual in the adult population, may signify at a molecular level processes that occur in malignant transformation and adult stem cell renewal in various tissues. Indeed, the rarity of certain germ cells tumors in the adult population may be a consequence of early signaling pathways in the formation and migration of germline destined cells. At the same time, many questions arise while finalizing the diagnosis and final report. Why does gonadal dysgenesis occur and why does this usually result in a malignant tumor with cells that in certain molecular respects resemble embryonic stem cells? Why do teratomas and other germ cell tumors arise in other anatomic locations (CNS, mediastinum, tongue)? How do germ cells become germ cells and how do they get where they should be? Why do these "stem cells" sometimes form all three germ layers in a benign fashion? Why do they sometimes take a more immature path and form predominantly neural tissues? Why is the malignant element in immature teratomas in the form of the endodermal sinus tumor? A monograph of questions could be constructed that would be exhaustive. This balance and these answers will have roots in the molecular character of the embryonic stem cell and events that occur early in its cellular life.
Primordial germ cells (PGCs) can be isolated from the mouse embryo at gestational day 7, and be maintained in culture for long periods of time and do exhibit the potential to contribute to all three embryonic germ layers and are considered pluripotent (6). Intratubular germ cell neoplastic cells (carcinoma in situ) of the testis can have varied phenotypes within the same tubule expressing antigens commonly seen in seminomas, non-seminomatous germ cell tumors and both (7). It is proposed that PGCs are under the influence of multiple polypeptide growth factors that affect both survival and proliferation. Evidence suggests that certain factors such as Steel factor (SLF) may be necessary for proper migration of PGCs into the fetal gonad and lack of the transmembrane factor may allow for migration of PGCs into other areas of the body. If other areas can produce SLF then these PGCs can survive without undergoing apoptosis. This may in part explain the existence of teratomas and other germ cell tumors in other anatomic areas besides the gonad, i.e. mediastinum, brain. Evidence exists that the c-kit-signaling pathway (and other factors) is critical for the proliferation, migration, differentiation, and survival of PGCs. Combinations of these factors (SLF, leukemia inhibitory factor (LIF), fibroblast growth factor (bFGF)) in vitro can have dramatic effects on PGC growth effectively immortalizing these cells. This model proposes that inappropriate activation of the c-kit signaling pathway can lead to continued proliferation of PGCs. Interestingly, c-kit levels are low or undetectable in human non-seminoma and teratoma, but high in intratubular germ cell neoplasia (carcinoma in situ) and seminoma (6) signifying continued activation of this pathway. The process of malignant transformation may also be reversible and flexible. Single teratocarcinoma cells cloned into marked mouse blastocysts can be shown to contribute to a wide range of somatic tissues in the chimeric mice. With a near normal chromosome complement and the proper environment, orderly gene expression can be restored (8).
Very recent data demonstrate the presence of OCT3/4 protein by immunohistochemical methods in carcinoma in situ/gonadoblastoma, seminoma/germinoma/dysgerminoma and embryonal carcinoma, but not in differentiated non-seminomatous germ cell tumors (9). OCT3/4 is a member of the POU family of transcription factors that are expressed in pluripotent mouse and human embryonic stem and germ cells including PGCs. This octamer activates transcription by binding to sites near transcriptional initiation sites and is a key element in the regulation of embryonic cell differentiation (10,11).
This brief, focal, and incomplete review of stem cell technology reveals the future directions of molecular embryology. Human and non-human primate embryonic stem cell technology is still in its infancy and may hold the key to transforming the advances of the partnership between molecular biology, sophisticated non-invasive imaging, and embryology into therapeutic benefit and understanding childhood diseases, developmental anomalies, and malignant transformation.
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Contributed by John A Ozolek, MD, Ronald Jaffe, MB, BCh, and Maria Z Parizhskaya, MD
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