Germ cell proliferation and apoptosis in the developing human ovary

CONTEXT: The regulation of germ cell proliferation and loss during human ovarian development is poorly understood. This is of particular interest at the time leading up to the formation of primordial follicles, at 18 wk gestation onward. OBJECTIVE: The objective of the study was to identify and quantify germ cell proliferation and apoptosis and expression of caspases in the human fetal ovary. DESIGN: This study was a laboratory investigation. SETTING: The study was conducted at a research institute. METHODS: Cell proliferation and apoptosis were detected using immunohistochemical localization of phosphorylated histone H3 and cleaved caspase-3, respectively. Caspases were also detected by immunoblotting. RESULTS: The overall proportion of germ cells in mitosis remained constant between 14 and 19 wk but showed increasing clustering. Caspase-2, -3, -7, -8, and -9 were detected by immunoblotting. There was a significant increase in germ cell apoptosis. A specimen of 20 wk gestation showed similar phosphorylated histone H3 but markedly lower cleaved caspase-3 expression than earlier gestations. Cleaved caspase-3 was not expressed in oocytes that had formed primordial follicles. CONCLUSIONS: These results indicate that as primordial follicle formation is initiated and progresses, there is an increase in both mitotic activity and apoptosis of those germ cells that have not reached the apparently protective environment of the primordial follicle.     

Germ cells are essential for sexual dimorphism in the medaka gonad

To further elucidate the roles of germ cells in the sex differentiation of gonads, we have used the medaka, a teleost fish, to generate mutants that lack germ cells from the onset of gonadogenesis by the morpholino-mediated knockdown of cxcr4. The resulting germ-cell-deficient medaka show female-to-male sex reversal of their secondary sex characteristics, accompanied by increased levels of androgen and reduced levels of estrogen. A failure to maintain granulosa cells or estrogen-producing cells also occurs at early stages of sex differentiation in the cxcr4 morphants, before the initiation of gonadal morphogenesis. In contrast, androgen-producing cells are unaffected in germ-cell-deficient medaka of either sex. In addition, a single tube-like gonad that expresses male-specific genes is formed in these mutants irrespective of the genetic sex. Significantly, each of these mutant phenotypes occurs in a somatic cell-autonomous manner, suggesting that gonadal somatic cells are predisposed toward male development in the absence of germ cells. This highlights the importance of germ cells in the sexual dimorphism of the gonads.     

Expression of steroidogenic factor-1 in frog embryo and developing gonad

The effects of a dinoflagellate parasite (Hematodinium sp.) on carbohydrate metabolism were examined in the Norway lobster, Nephrops norvegicus. Five stages of infection were observed. These included uninfected (Stage 0), subpatently infected (SP), and patently infected (Stage 1-4) lobsters. During patent infection, the concentration of glucose in the hemolymph was reduced significantly from its value of 180 microg ml(-1) in uninfected (Stage 0) lobsters to 25.3 microg ml(-1) in Stage 3-4. These changes were accompanied by significantly lower levels of hepatopancreatic glycogen in lobsters at Stage 2 (2.01 mg g(-1)) and Stage 3-4 (0.84 mg g(-1)) of infection than in those at Stage 0 (16.19 mg g(-1)) and Stage 1 (14.71 mg g(-1)). Due to disruption of the normal feedback loops which control the release of crustacean hyperglycemic hormone (CHH), plasma concentrations increased with the severity of infection from 32.2 fmol ml(-1) in Stage 0 to 106.6 fmol ml(-1) in Stage 3-4. The increased CHH concentrations occurred concomitantly with reduced concentrations of plasma glucose and tissue glycogen. A significantly increased hemolymph CHH titer (107.7 fmol ml(-1)) was also observed during SP infection. It is concluded that the parasite places a heavy metabolic load on the host lobster.     

Germ cell tumours of the testis

Cancer of the testis is a relatively rare disease, accounting for about 1% of all cancers in men. Cryptorchidism is the only confirmed risk factor for testicular germ cell tumour. The majority of GCT are clinically detectable at initial presentation. Any nodular, hard, or fixed area discovered in the testis, must be considered neoplastic until proved otherwise. The appropriate surgical procedure to make the diagnosis is a radical orchidectomy through an inguinal incision. Many GCT produce tumoural markers (AFP, HCG, LDH), who are useful in the diagnosis and staging of disease; to monitor the therapeutic response and to detect tumour recurrence. In 1997 a prognostic factor-based classification for the metastatic germ cell tumours was developed by the IGCCCG: good, intermediate and poor prognosis, with 5-year survival of 91, 79 and 48%, respectively. GCT of the testis is a highly table, often curable, cancer. Germ cell testicular cancers are divided into seminoma and non-seminoma types for treatment planning because seminomatous testicular cancers are more sensitive to radiotherapy. Seminoma (all stages combined) has a cure rate of greater than 90%. For patients with low-stage disease, the cure approaches 100%. For patients with non-seminoma tumours, the cure rate is >95% in stages I and II; it is approximately 70% with standard chemotherapy and resection of residual disease, if necessary, in stages III and IV. Minimum guidelines for clinical, biochemical, and radiological follow-up have been reported by ESMO in 2001.     

Mechanisms and cell lineages in lymphatic vascular development

Angiogenesis - Lymphatic vessels have critical roles in both health and disease and their study is a rapidly evolving area of vascular biology. The consensus on how the first lymphatic vessels...     

Germ cell cancer risk in DSD patients

The risk of germ cell cancer is elevated in many DSD patients, although not to the same extent. A number of risk factors have been identified recently, but their interplay and relative impact is currently not fully clear. Until the advent of reliable screening tools for the detection of pre-invasive cancer lesions, managing germ cell tumour risk focuses on the question if and when to perform biopsy or gonadectomy in most patients and how to interpret the histological findings.     

Germ cell stimulation of Sertoli cell protein phosphorylation

Cultures of Sertoli cells were treated with freshly isolated, intact germ cells to determine if germ cells were capable of influencing Sertoli cell function. By using two-dimensional polyacrylamide gel electrophoresis and autoradiography, germ cells were found to increase several-fold the incorporation of [32P]orthophosphate into two phosphoproteins (which we term germ cell-dependent phosphoproteins 1 and 2 or GC1 and GC2) of Sertoli cells. Increased phosphorylation of GC1 and GC2 was rapid, germ cell dose dependent, and calcium dependent. The increased phosphorylation of GC1 appears to involve calmodulin-dependent protein kinase, while phosphorylation of GC2 appears to involve the activation of calcium/phospholipid-dependent protein kinase (protein kinase C). Medium conditioned by germ cells was capable of eliciting the same response in Sertoli cells. Germ cells had no effect on the phosphorylation of proteins in Chinese hamster ovarian cells, but did result in increased phosphorylation of a protein in TR-ST cells, which migrated similarly to GC1 of Sertoli cells. Neither Chinese hamster ovarian nor TR-ST cells had any effect on Sertoli cell protein phosphorylation. These results indicate that germ cells may be directly involved in the local regulation of Sertoli cell function within the seminiferous epithelium. The results further suggest that the mechanism of germ cell-Sertoli cell interaction involves the mobilization of intracellular calcium, activation of Ca2+/calmodulin-dependent protein kinase, and protein kinase C. We infer from these results that germ cell-Sertoli cell interaction may operate via hydrolysis of Sertoli cell membrane phophatidylinositols.     

Germ cell transplantation and the study of testicular function.

Spermatogonial stem cell transplantation is a novel technique in which donor testicular cells are transferred into recipient testes. A population of germ cells from a transgenic or mutant donor is introduced into the seminiferous tubules of recipient testes by microinjection. Following injections, spermatogonial stem cells can colonize the recipient testis, initiate spermatogenesis and produce sperm capable of fertilization. This technique will allow scientists to: (1) investigate fundamental aspects of spermatogenesis; (2) provide a method to regenerate spermatogenesis in infertile individuals; and (3) genetically manipulate spermatogonial stem cells to develop transgenic animals.     

Differential expression of microRNAs in hematopoietic cell lineages

Objectives: MicroRNAs (miRNAs) play key roles in a wide variety of normal and pathological cellular processes. A number of studies identified hematopoietic‐specific miRNAs that are necessary for correct function of blood cells. Out of our microarray data, we chose 13 miRNAs that showed differential expression in peripheral blood cells (miR‐15b, miR‐16, miR‐24, miR‐30c, miR‐106b, miR‐142‐3p, miR‐142‐5p, miR‐150, miR‐155, miR‐181, miR‐223, miR‐342, and miR‐451) and examined their expression in separated hematopoietic cell lineages. Methods: Using quantitative real‐time polymerase chain reaction, we measured relative expression of the miRNAs in fractions of reticulocytes, platelets, granulocytes, monocytes, B‐ and T‐lymphocytes as well as in several hematopoietic cell lines. Results: We observed that miR‐16 and miR‐142‐3p were highly expressed in all native cell lineages, miR‐451 reached the maximal expression in reticulocytes, miR‐223 in platelets, granulocytes and monocytes, and miR‐150 in B‐ and T‐lymphocytes. Hierarchical clustering analysis grouped the lineage samples according to their origin based on the expression of these miRNAs. To validate discrimination power of the miRNAs, we quantified expression of the 13 miRNAs in several immortalized cell lines. Although the cell lines showed miRNA expression patterns considerably different from those of native cell lineages, clustering analysis distinguished between myeloid, lymphoid and non‐hematopoietic cells. Conclusions: In conclusion, the study reports the expression levels of 13 miRNAs in particular blood cell lineages as well as immortalized cell lines. We demonstrate that the expression profiles of these miRNAs may be used for discrimination of the hematopoietic cell lineages.     

Heterogeneity of cell lineages in L3 leukemias

Five cases of adult leukemia with L3 morphology in bone marrow were studied for the presence of immunologic, metabolic, and enzymatic markers. Among the five patients, there were four males and female. Median age was 66 with a range of 16-80 yr. Median survival was only 5 mo. Serum lactate dehydrogenase (LDH) levels, 3H-thymidine labeling indices, and DNA/RNA content of the L3 lymphoblasts were markedly elevated. B-cell markers were found in three cases, two exhibiting surface membrane IgM-lambda, and one IgG-K. Terminal deoxynucleotidyl transferase (TdT) enzymatic activity was consistently low in this group. In one case, the L3 lymphoblasts displayed only surface Fc receptors demonstrated by the binding of aggregated IgG. TdT activity was found to be significantly increased. In another instance, the lymphoblasts formed spontaneous rosettes with sheep erythrocytes and exhibited paranuclear staining with acid phosphatase. TdT activity was found to be low. Although most of the L3 leukemias are neoplasias of B lymphocytes, other lineages may also express this morphology.