Granulosa cell-oocyte interactions: the phosphorylation of specific proteins in mouse oocytes at the germinal vesicle stage is dependent upon the differentiative state of companion somatic cells.

The role of granulosa cells in the regulation of mouse ovarian oocyte metabolism was investigated. Fully grown antral oocytes, isolated from surrounding cumulus cells, were cultured on monolayers of preantral granulosa cells in the presence of dbcAMP to prevent the resumption of meiosis. Under these conditions metabolic cooperativity was established between the two cell types as early as 1 hr after seeding. Moreover, cocultured oocytes phosphorylated two polypeptides of 74 and 21 kDa which are normally phosphorylated in follicle-enclosed growing oocytes but not in cumulus cell-enclosed fully grown oocytes at the germinal vesicle stage. When cocultured oocytes were allowed to resume meiosis, the 74 and 21 kDa proteins were synthesized but no longer phosphorylated even though intercellular coupling between the two cell types was maintained during radiolabeling. It appears therefore: a) that the different protein kinase activity of growing and fully grown germinal vesicle-stage mouse oocytes is related to the differentiative state of granulosa cells, and b) that the regulation of oocyte protein phosphorylation activity by granulosa cells is dependent on the meiotic stage of the oocyte

Somatic cell-oocyte interactions in mouse oogenesis: stage specific regulation of mouse oocyte protein phosphorylation by granulosa cells.

The relative rate of synthesis of a number of proteins and the protein phosphorylation pattern of growing and fully grown oocytes were influenced by the presence of granulosa cells. In particular, a 74-kDa phosphorylated protein was detected only in granulosa cell-enclosed growing mouse oocytes. When reaggregated with granulosa cells, the growing oocyte displayed the phosphorylated form of the 74-kDa protein but when oocytes were cultured on Sertoli cell monolayers or in granulosa cell-conditioned medium the 74-kDa protein was not phosphorylated. We propose that (1) granulosa cells regulate protein phosphorylation in mouse oocytes; (2) a 74-kDa protein is phosphorylated only in growing oocytes when surrounded by granulosa cells; and (3) granulosa cells, but not Sertoli cells, are competent to send the appropriate "signal" to the growing oocyte

Mammalian oocyte growth in vitro is stimulated by soluble factor(s) produced by preantral granulosa cells and by Sertoli cells

We have evaluated the possibility that mouse oocyte growth in vitro could be achieved under the influence of soluble compound(s) released by different somatic cell types. For this purpose, zona-free denuded oocytes from 12-day-old mice were cultured on monolayers of NIH-3T3 fibroblasts, which are able to establish gap junctional communications with them, in the presence or absence of media conditioned by preantral granulosa cells or by Sertoli cells, plated at increasing concentrations from 0.3-1 x 10(6) ml-1 cells. After 3 days, no increase in vitellus diameter was recorded from fibroblast-coupled oocytes maintained in culture medium or in the presence of media conditioned by 0.3 x 10(6) ml-1 Sertoli cells. By contrast, increasing proportions of coupled oocytes grew, provided the continuous presence of media conditioned by 0.5 or 1 x 10(5) ml-1 Sertoli cells, or by 0.3, 0.5, and 1 x 10(5) ml-1 preantral granulosa cells. Since the ligand of c-kit, the growth factor KL, promotes the growth in vitro of oocytes cultured in follicles from 8-day-old mice, an antibody against mouse KL was used to evaluate whether in our culture conditions KL might also be responsible for the growth of oocytes from 12-day-old mice. No inhibition of growth was evident in oocytes cultured directly on preantral granulosa or Sertoli-cell monolayers. Furthermore, the growth of fibroblast-coupled oocytes cultured in media conditioned by preantral granulosa cells was not significantly affected by the presence of this antibody during culture. By contrast, a high percentage of oocytes cultured on fibroblasts in the presence of media conditioned by Sertoli cells showed a significant inhibition of growth and no metabolic cooperativity. It was concluded that, besides KL, other bioactive factor(s) released by either preantral granulosa or Sertoli cells can induce a significant stimulation of mouse oocyte growth in vitro

Follicle-stimulating hormone induced-phospholipase A2 activity and -eicosanoid generation in rat Sertoli cells.

The possibility that FSH stimulates the phospholipase A(2) (PLA(2)) pathway was studied in cultured immature Sertoli cells. FSH induced [H-3]-arachidonic acid (AA) release from prelabeled cells in a time- and concentration-dependent fashion (ED(50) = 21.8 +/- 1.9 ng/ml). This response could be fully prevented by pretreatment of cells with the PLA(2) inhibitor, mepacrine. That PLA(2) was the main enzyme responsible for cleavage of AA from membrane phospholipids was directly shown by PLA(2) activity assay using vesicles of radiolabeled phosphatidylcholine (PC) as substrate. Furthermore, FSH stimulated eicosanoid generation in a time-dependent manner through the cyclooxygenase but not the lipoxygenase pathway. In fact, higher levels of prostaglandin (PG) E(2), F-2 alpha,F- and the stable products of PGI(2) and thromboxane A(2) (6-keto PGF(1 alpha) and thromboxrane B-2, respectively) were generated by the gonadotropin-treated cells as compared to control cells. The effect was inhibited by mepacrine, further supporting the pivotal role of PLA(2) in the release of the eicosanoid precursor, AA. Finally, the effect of the main product of FSH-induced AA metabolism, i.e., PGE(2), was studied. Intracellular cAMP accumulation in Sertoli cells was stimulated by the prostanoid in a dose-dependent manner (ED(50) = 2.3 +/- 0.37 nM). PGE(2) also significantly stimulated aromatase activity, a specific marker of Sertoli cell functions, measured as 17 beta-estradiol production (ED(50) = 4.7 +/- 0.8 nM). Similar results were obtained with PGF(2 alpha). Our findings show that FSH, through the activation of PLA(2), leads to AA release with consequent metabolism by the cyclooxygenase pathway. Prostanoids produced by Sertoli cells upon FSH stimulation can control, in an autocrine or paracrine manner, the functions of the somatic cells in the seminiferous epithelium

Aortic dimensions in patients with bicuspid aortic valve without significant valve dysfunction

The dimensions of the entire aorta at different anatomic levels were measured by transthoracic 2-dimensional echocardiography in 162 consecutive patients with iso- lated bicuspid aortic valves (BAVs) without significant aortic valve dysfunction. Aortic dilation involved the aortic root and the ascending aorta but was not present in the descending and abdominal aorta. A significant increase in the dimensions of the aortic arch was found in patients with BAVs aged >40 years. Ascending aortic diameter and the extension of aortic dilation were sig- nificantly correlated with age, but no correlation was found between aortic dimensions and aortic valve morpholog