Serum progesterone at human chorionic gonadotropin injection significantly correlate with female age.

AIM: In in vitro fertilization-embryo transfer (IVF-ET) higher age and low responses are associated with accelerated luteinization of mature follicles rather than diminished responsiveness. The aim of this study was to determine whether an elevated serum progesterone (P) on the day of human chorionic gonadotropin (hCG) administration during gonadotropin stimulation for IVF-ET is associated with age. METHODS: E2 (17beta estradiol) and P concentrations on the day of hCG administration, number and quality of oocytes and embryos, and clinical pregnancies were retrospectively analyzed in 460 women undergoing IVF-ET. We evaluated patients according to age; the 25-30 age group (n=140), the 31-35 age group (n=100), the 36-40 (n=90), and the 41-45 age group (n=130). RESULTS: In the 25-30 age group (n=140) P was 0.67+/-0.3 ng/mL, in the 31-35 age group (n=100) P was 0.87+/-0.2 ng/mL, in the 36-40 age group (n=90) P was 0.95+/-0.2 ng/mL, in the 41-45 age group (n=130) P was 1+/-0.2 ng/mL. The difference between the 25-30 age group and the 41-45 age group was statistically significant (P<0.05). CONCLUSIONS: Periovulatory levels of serum P vary according to ovarian response to controlled ovarian hyperstimulation. Periovulatory P may reflect inadequate steroidogenesis. In women stimulated with recombinant follicle stimulating hormone for IVF, the serum P on the day of hCG administration increases with age.

Cabergoline influences ovarian stimulation in hyperprolactinaemic patients with polycystic ovary syndrome.

BACKGROUND: Polycystic ovary syndrome (PCOS) is characterized by abnormal gonadotrophin secretion, in particular an elevated serum concentration of LH, depressed FSH, and an LH/FSH ratio of >or =2. Mild, transient hyperprolactinaemia is frequently associated with PCOS (30% of patients); furthermore, it can be observed during the late follicular and luteal phases of both natural and stimulated cycles. It is suggested that a reduction of the dopamine inhibitory effect might raise both prolactin (PRL) and LH. METHODS AND RESULTS: We compared ovarian stimulation in two groups of hyperprolactinaemic (hyperPRL)-PCOS patients; one group was treated with cabergoline, reducing PRL plasma concentrations to the range normally observed during ovulation induction. In the untreated hyperPRL-PCOS group, we noted a reduced total number of ampoules of recombinant FSH (P < 0.04), fewer days to reach HCG administration (P < 0.04), and significantly higher peak oestrogen plasma concentrations (P < 0.03) compared with the treated group. By ultrasound examination the same group showed significantly higher ovarian volume and an increased total number of follicles of every size. In untreated hyperPRL-PCOS patients, four cycles out of 65 were cancelled due to mild ovarian hyperstimulation syndrome (OHSS) that occurred during ovulation induction. Only one cycle out of 42 in the patients treated with cabergoline was cancelled. No significant differences in pregnancy rate nor in multiple pregnancy were found. CONCLUSION: Our data suggest a dopaminergic control of LH release and support the use of cabergoline in the management of such patients, in order to provide better clinical control of ovarian response and consequently a reduction of the risk of OHSS, with no decrease in pregnancy rate.

Polycystic ovary syndrome: evidence for reduced 3 beta-hydroxysteroid dehydrogenase gene expression in human luteinizing granulosa cells.

In a preview study we found that luteinizing granulosa cells from follicles of patients with polycystic ovary syndrome (PCOS) have a reduced capacity to synthesize progesterone in vitro. Because the 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4-isomerase (3 beta-HSD) is an important enzyme for the biosynthesis of progesterone, the reduced capacity of PCO luteinizing granulosa cells to synthesize progesterone in vitro may be due to reduced 3 beta-HSD gene expression. A reverse transcriptase polymerase chain reaction for 3 beta-HSD was performed and the relative intensity of signals for 3 beta-HSD was evaluated using computer-assisted densitometry. Cells from polycystic ovaries expressed less 3 beta-HSD in follicles < or 10 mm (p < 0.05) and in follicles > or 16 mm (p < 0.05) than cells from normal ovaries. Furthermore, after human chorionic gonadotropin stimulus (50 ng/ml), cells from polycystic ovaries expressed less 3 beta-HSD in follicles > or = 16 mm (p < 0.01) than cells from normal ovaries. The data show that there is a specific change in the gene expression of 3 beta-HSD in PCO granulosa cells resulting in a suppressed capacity to secrete progesterone.

Treatment versus no treatment of transient hyperprolactinemia in patients undergoing intracytoplasmic sperm injection programs.

The aim of our study was to investigate the effect of increased plasma prolactin levels on oocyte and fertilization rate in patients undergoing in vitro fertilization (IVF) intracytoplasmic sperm injection (ICSI) treatment. We identified 135 patients with transient or borderline hyperprolactinemia, measured in the mid and late follicular phase and in the mid-luteal phase of the cycle before ovarian stimulation. The patients were assigned to either the no treatment group (76 patients) or the treatment group (59 patients). The treated group underwent treatment with cabergoline or bromocriptine before ovarian stimulation, until there was a decrease of plasma prolactin levels, and the therapy was continued also during the ICSI programme. Both groups received a gonadotropin-releasing hormone (GnRH) agonist and were subsequently stimulated with follicle-stimulating hormone (FSH) up to the day of human chorionic gonadotropin (hCG) administration. The untreated group needed a significantly lower number of FSH ampoules than the treated group to reach the day of hCG administration (38.1 +/- 18.2 versus 43.9 +/- 28.5; p < 0.05). No correlation was found between the two groups on the peak estradiol level achieved, the progesterone level at hCG administration and the numbers of oocytes retrieved. The number of oocytes with superior morphology (87.9% versus 80.4%; p < 0.05), the fertilization rate (70.8 +/- 28.0 versus 60.8 +/- 28.5; p < 0.03), and the mean number of embryos transferred (3.6 +/- 1.6 versus 3.2 +/- 1.5; p < 0.05) were significantly higher in the patients whose hyperprolactinemia was left untreated. In conclusion, we found that transient hyperprolactinemia is positively associated with ICSI outcome, in particularly with oocyte quality and fertilization rate.

The use of Matrigel at low concentration enhances in vitro blastocyst formation and hatching in a mouse embryo model.

OBJECTIVE: To determine the effect of Matrigel at a low concentration on the growth of mouse embryos in culture. DESIGN: Randomized case-control study of mouse embryos. SETTING: An academic research environment. ANIMALS: Mouse embryos. INTERVENTION(S): Embryos were cultured in Quinn's or Celbio's human tubal fluid (HTF) enriched with 1.5% bovine serum albumin and 0.8% liquid Matrigel. Each HTF was compared with the same medium devoid of Matrigel. Afterward, Quinn's and Celbio's HTF, both containing Matrigel, were compared directly. Embryos were cultured in four-well dishes, and their morphology and viability were assessed at 96 hours. MAIN OUTCOME MEASURE(S): Level of interleukin-1alpha in media collected at the end of culture. RESULT(S): In both types of HTF, the presence of Matrigel allowed a larger number of embryos to reach the blastocyst stage and to hatch; blastocyst morphology also was improved. These positive effects were enhanced in Quinn's HTF: embryos cultured in its Matrigel-enriched version secreted a higher level of interleukin-1alpha than those in Celbio's HTF plus Matrigel and also showed a better morphology. CONCLUSION(S): In the mouse embryo model, Matrigel improves culture conditions in terms of both embryo viability and morphology, and these effects are enhanced in Quinn's HTF.

Human albumin enhances expression of vascular endothelial growth factor in cultured human luteinizing granulosa cells: importance in ovarian hyperstimulation syndrome.

Ovarian hyperstimulation syndrome (OHSS) is a severe complication of ovarian stimulation for assisted reproductive techniques. Clinical manifestations are massive extravascular fluid accumulation and haemoconcentration. Vascular endothelial growth factor (VEGF) has been demonstrated to mediate the development of OHSS. Intravenous albumin at the time of oocyte aspiration has been suggested as an effective prophylactic treatment against the occurrence of severe OHSS. Here it is reported that in cultured human luteinizing granulosa cells, VEGF mRNA expression was enhanced by human albumin and maximum expression was observed in cultured granulosa cells obtained from patients with serum oestradiol concentrations >2000 pg/ml on the day of human chorionic gonadotrophin injection (P < 0. 05).

Evidence that a functional fertilin-like ADAM plays a role in human sperm-oolemmal interactions.

Fertilin is a protein initially identified in guinea pig spermatozoa; it is the prototype of a larger family of conserved, proteins designated as a disintegrin and a metalloproteinase (ADAM). These heterodimers which consist of alpha and beta subunits, containing metalloproteinase-like and disintegrin-like domains, appear to play a role in mammalian fertilization. Peptides derived from the disintegrin domains of two ADAMs, fertilin and cyritestin, interfere with gamete adhesion and sperm-egg membrane fusion in non-human species. It has been suggested that fertilin-beta binds to an oolemmal integrin, and it is proposed that the tripeptide FEE (Phe-Glu-Glu) is the integrin recognition sequence in human fertilin-beta. We evaluated whether fertilin beta plays a role in human fertilization by studying the effects of a linear octapeptide containing the FEE sequence, SFEECDLP, and a scrambled octapeptide with the same amino acids, SFPCEDEL, on the incorporation of human spermatozoa by human zona-free eggs. The effects of G4120, a potent RGD-containing (Arg-Gly-Asp) thioether-bridged cyclic peptide which blocks both fibronectin and vitronectin receptors, and the relationship between FEE- and RGD-receptor interactions on sperm-egg interactions were also studied. The FEE-containing peptide, but not the scrampled peptide, inhibited sperm adhesion to oocytes and their penetration, over the range 1-5 microM. The inhibition induced by SFEECDLP was reversible and occurred only in the presence of peptide itself. The G4120 peptide exhibited 10-fold less inhibitory effects on sperm adhesion and penetration than did SFEECDLP. When combined, SFEECDLP and G4120 exhibited strong inhibition of both adhesion and penetration at concentrations that individually had been ineffective, suggesting co-operation between the two receptor-ligand interactions during fertilization. We propose that a fertilin-like molecule is functionally active on human spermatozoa and that its interaction with an oolemmal integrin receptor plays a role in fertilization in humans.

Elevated serum progesterone on the day of HCG administration in IVF is associated with a higher pregnancy rate in polycystic ovary syndrome.

Our study compared 84 patients with polycystic ovary syndrome (PCOS) with 84 control patients who had normal ovaries and who were matched for the main determinants of success in in-vitro fertilization (IVF) and embryo transfer. Serum concentrations of oestradiol and progesterone on the day of human chorionic gonadotrophin (HCG) injection were significantly higher in PCOS than in normal patients (oestradiol 2016 +/- 1.8 pg/ml versus 1456 +/- 40.9 pg/ml, P < 0.01; progesterone 1.6 +/- 0.1 ng/ml versus 1.2 +/- 0.1 ng/ml, P = 0.03). Furthermore despite oocytes from PCOS patients having a reduced fertilization rate compared with normal patients (61.8 +/- 4.1% versus 73.5 +/- 4.3%, P = 0.03), the differences in pregnancy rate (22.6 versus 19%) and miscarriage (31.5 versus 18.7%) were not statistically significant. In PCOS patients, a critical breakpoint was identified at serum progesterone concentrations of 1.2 ng/ml on the day of HCG injection. The PCOS patients with progesterone > or = 1.2 ng/ml showed a higher pregnancy and miscarriage rate than PCOS patients with progesterone < 1.2 ng/ml (26.6 versus 17.9%, P < 0.01; and 41.7% versus 14.3%, P < 0.01 respectively). These findings suggest that premature progesterone production does not have an adverse effect on pregnancy rate in PCOS, but on the contrary, may be a predictor for success in IVF/embryo transfer.

Polycystic ovary syndrome: anomalies in progesterone production.

The underlying cause of anovulation and miscarriage in polycystic ovary syndrome (PCOS) is unknown. Progesterone may play an important role in oocyte fertilization and embryo implantation. Therefore, in this study we analyse the endocrine function of luteinizing granulosa cells to synthesize progesterone in vivo and in vitro in PCOS and normal patients participating in an in-vitro fertilization programme. Human luteinizing granulosa cells were obtained from 10 patients with normal ovaries (controls) and 10 patients with PCOS by follicular aspiration of individual follicles of each patient and pooled in an attempt to obtain three groups: cells from follicle sizes < or =10,>10< or =15 and > or =16. Serum concentrations of oestradiol and progesterone on the day of human chorionic gonadotrophin (HCG) injection were significantly higher (P < 0.01 and P < 0.05) in PCOS patients than in controls. After HCG stimulation, in-vitro progesterone production was enhanced in granulosa cells of the control group and concentrations increased with follicular size as expected. However, the concentration of progesterone of PCOS patients did not increase with follicular size and there was a significant difference between normal and PCOS groups in follicles >10< or =15 mm (P < 0.05) and > or =16 mm (P < 0.01). Oestradiol production was increased in follicles > or =16 mm in both groups, although this did not reach significance. In summary, it seems that PCOS granulosa cells demonstrate an abnormal capacity to synthesize progesterone in vivo and in vitro. The understanding of granulosa cell function in PCOS may explain the anovulation and miscarriage that occurs in these patients.

Premature ovarian failure: steroid synthesis and autoimmunity.

The androgen biosynthesis and autoimmunity of 25 patients with premature ovarian failure (POF) and 18 control subjects with normal cycles were examined. Serum levels of dehydroepiandrosterone sulfate (DHEAS), 17-hydroxyprogesterone (17-OHP), androstenedione, and testosterone were analyzed in POF patients with or without organ-specific autoimmunity, and the results compared with those of women with normal ovarian function. The comparative analysis of DHEAS, 17-OHP, androstenedione and testosterone showed that POF patients had significantly lower values than normal women (DHEAS, androstenedione and testosterone p < 0.01, 17-OHP p < 0.05). Furthermore, we found one or more organ-specific autoantibodies in 11 patients with POF (44%), while only one woman in the control group showed autoimmunity (antithyroid microsome) (5.5%). Only one patient had both anti-ovarian and anti-adrenal antibodies (4%). The comparison of androgen levels in POF patients with or without autoimmunity revealed a statistically significant reduction of DHEAS levels in POF patients with organ-specific autoimmunity (p < 0.01). These data reveal the reduction in androgen synthesis in POF patients, particularly in those with organ-specific autoimmunity.

Hashimoto's disease in a papillary carcinoma of the thyroid originating in a teratoma of the ovary (malignant struma ovarii).

Adenocarcinomas represent a relatively rare complication of a cystic teratoma of the ovary. Those of thyroid origin have been reported in only a few cases. In this paper we report a case of papillary carcinoma of the thyroid arising from a cystic teratoma. The patient had no thyroid symptoms, but because of the presence of antimicrosomal and antithyroglobulin antibodies the diagnosis of Hashimoto's disease was made.

Expression of vascular endothelial growth factor in human luteinizing granulosa cells and its correlation with the response to controlled ovarian hyperstimulation.

Ovulation induction represents one of the most important steps for the success of assisted reproductive technology (ART) procedures. To better understand the mechanisms that regulate follicle growth, oocyte maturation, and ovarian steroidogenesis, we investigated the correlations between vascular endothelial growth factor (VEGF) gene expression in human luteinizing granulosa cells, steroid production and oocyte retrieval in patients undergoing controlled ovarian hyperstimulation. We evaluated the messenger ribonucleic acid (mRNA) for VEGF in human luteinizing granulosa cells obtained at the time of oocyte retrieval from 24 women participating in an in vitro fertilization program at the Reproductive Endocrinology Center of our Department of Obstetrics and Gynecology. We found a positive linear correlation of VEGF mRNA with estradiol and progesterone serum levels at the day of oocyte retrieval (p < 0.05). Furthermore, VEGF mRNA expression was significantly higher in granulosa cells obtained from patients with an elevated number of oocytes and high fertilization rate (p < 0.05). Our data confirm that VEGF may play an important role in the regulation of vascular development during follicular growth and luteal differentiation.

In controlled ovarian hyperstimulation, steroid production, oocyte retrieval, and pregnancy rate correlate with gene expression of vascular endothelial growth factor.

PURPOSE: Whether the gene expression of vascular endothelial growth factor (VEGF) in human granulosa cells is a predictor of fertilization was evaluated in patients participating in an in vitro fertilization program. METHODS: Fifty patients with normal ovaries who were participating in an in vitro fertilization program at the University of Milan, San Raffaele Scientific Institute, were included in the study. We correlated E2 and P serum levels on the day of oocyte collection, the number of follicles, oocytes collected, and fertilized, and pregnancies with mRNA for VEGF of luteinizing granulosa cells obtained at the time of oocyte retrieval. RESULTS: Comparing E2 and P serum levels, the number of follicles, oocytes collected and fertilized, and pregnancies with gene expression for VEGF, we found a positive correlation. E2 and P serum levels were higher in patients with increased VEGF (P < 0.01). Furthermore, there were more follicles, oocytes collected and fertilized, and pregnancies in patients with maximum expression of VEGF, and the difference was statistically significant (P < 0.05). CONCLUSIONS: Our results suggest that VEGF may be important for vascular development during follicular growth and luteal differentiation, oocyte maturation, and fertilization.

Vascular endothelial growth factor messenger ribonucleic acid expression in human ovarian and endometrial cancer.

Vascular endothelial growth factor (VEGF) is a previously discovered angiogenic factor that seems to influence the neoangiogenesis of neoplastic and non-neoplastic tissues. Substantial experimental evidence links tumor growth and metastasis with blood vessel formation. Tumor angiogenesis can be induced by factors released by the tumor cells themselves. A variety of transformed cell lines expresses the VEGF transcript and secretes an EGF-like protein, suggesting that this angiogenic factor may be one of the mediators of tumor angiogenesis. By Northern blot analysis and in situ hybridization, we investigated the expression of VEGF transcript in human ovarian and endometrial neoplasms. Messenger RNA encoding VEGF was detected in all tissues studied and was more densely expressed in endometrial carcinoma. VEGF expression was also identified in cells obtained from ovarian and endometrial ascitic fluid. VEGF mRNA, detected by in situ hybridization, was identified in the epithelial cells of endometrial adenocarcinoma. This distribution was localized primarily in the apices of the papillae. The prominence of VEGF mRNA levels in human ovarian and endometrial tumors demonstrates that VEGF may be involved in promoting tumor angiogenesis and stroma generation, acting as an endothelial cell mitogen.

Vascular endothelial growth factor. Expression in human vulvar neoplastic and nonneoplastic tissues.

OBJECTIVE: To investigate the expression of vascular endothelial growth factor (VEGF) in human vulvar neoplastic and nonneoplastic tissues. STUDY DESIGN: Specimens were collected at the Vulvovaginal Clinic, Department of Obstetrics and Gynecology, University of Milan. Human vulvar neoplastic and nonneoplastic tissues were dissected and frozen immediately at -80 degrees C until RNA extraction. Five micrograms of total RNA from each sample was denatured and transferred to nitrocellulose and nylon membranes for dot blot hybridization with labeled [alpha-32P]dCTP cDNA probe for VEGF. RESULTS: Messenger RNA encoding VEGF was detected in all tissues studied. VEGF mRNA was highly expressed in vulvar epithelial neoplasia (VIN) associated with human papillomavirus infection and minimally expressed in invasive squamous cells carcinoma of the vulva. Nonneoplastic lesions, such as chronic inflammation, lichen sclerosus, lichen planus, squamous hyperplasia and squamous papilloma, were also assessed, and none had a significant difference in VEGF mRNA expression. CONCLUSION: The prominence of VEGF mRNA levels in particular cases of VIN demonstrated that VEGF may be involved in promoting a new vascular network as a basic condition for the progression or at least self-maintenance of those lesions.

Expression of 17 alpha-hydroxylase and aromatase in the syndrome of androgen resistance: a case report.

A complete form of testicular feminization with normal gonadotropin and high testosterone levels is described. The testicular histology of tubular atrophy and hyperplastic Leydig cells accords with the high testosterone levels. We evaluated the expression of the enzymes involved in the production of testosterone and estrogens. An increase in P450c17 (17 alpha-hydroxylase/17,20-lyase) messenger RNA expression has been shown in testis with androgen resistance compared with in normal testis. More P450 aromatase was expressed in normal testis than in testis with androgen resistance.

Success in inducing ovulation in a case of premature ovarian failure using growth hormone-releasing hormone.

Ovulation was obtained in a 29-year-old woman affected by premature ovarian failure who had previously failed to respond to two attempts performed administering human menopausal gonadotropin or follicle-stimulating hormone after the spontaneous gonadotropin production was suppressed using a gonadotropin-releasing hormone analog (buserelin). Induction of ovulation succeeded when 1000 mg/day growth hormone-releasing hormone was added to the induction scheme. Five mature follicles were obtained after 27 days therapy and the serum level of 17 beta-estradiol was 975 pg/ml (195 pg/ml per follicle) at the time of human chorionic gonadotropin administration.

Effects of growth hormone and growth hormone-releasing hormone on steroid synthesis in cultured human luteinizing granulosa cells.

To assess the direct effect of growth hormone and growth hormone-releasing hormone on gene expression of steroidogenic enzymes and production of progesterone, 17-hydroxyprogesterone (17-OHP) and estradiol, we cultured luteinizing granulosa cells with or without follicle-stimulating hormone (FSH), growth hormone and growth hormone-releasing hormone at different concentrations. Luteinizing granulosa cells were obtained from women undergoing an in vitro fertilization program in the Department of Obstetrics and Gynecology, S. Raffaele Scientific Institute, Milan, Italy. At a concentration of 1 microgram/ml, FSH significantly increased estradiol production (2.1 +/- 0.7-fold the control value; p < 0.05 vs. control) and progesterone production (3.5 +/- 2.0-fold the control value; p < 0.05 vs. control). Growth hormone was effective on estradiol, progesterone and 17-OHP at 1 microgram/ml, enhancing estradiol production (1.3 +/- 0.2-fold the control value; p < 0.05 vs. control), progesterone production (2.5 +/- 1.0-fold the control value; p < 0.05 vs. control), and 17-OHP (1.4 +/- 0.2-fold the control value; p < 0.05 vs. control). Growth hormone-releasing hormone increased estradiol production (1.5 +/- 1.2-fold the control value) and progesterone production (1.3 +/- 0.8-fold the control value), but not significantly. No effects by growth hormone-releasing hormone were seen on 17-OHP production. FSH, growth hormone and growth hormone-releasing hormone did not increase P450scc and P450 aromatase mRNAs, whereas FSH increased P450c17 mRNA to 150% at 100 ng/ml and 1 microgram/ml, growth hormone increased it to 230% at 100 ng/ml and to 200% at 1 microgram/ml, and growth hormone-releasing hormone increased it to 140% at 100 ng/ml and to 190% of control values at 1 microgram/ml. These results indicate a direct effect of growth hormone on steroidogenesis by increasing P450c17 mRNA accumulation and progesterone, 17-OHP and estradiol production.

Complete testicular feminization with normal gonadotropin and high androgen secretion: a case report.

A patient with a complete form of testicular feminization with normal gonadotropin and high testosterone levels is described here. These findings are in contrast to previous reports that have shown high circulating luteinizing hormone (LH) levels in adults affected by this syndrome. Testicular histology revealed tubular atrophy and hyperplastic Leydig cells which are in accordance with the high testosterone levels and the difficulty in visualization of the testes. Because of the increased risk of gonadal malignancy, a laparoscopic gonadectomy was performed.

In the human fetus, vascular endothelial growth factor is expressed in epithelial cells and myocytes, but not vascular endothelium: implications for mode of action.

Vascular endothelial growth factor (VEGF) is potentially an important regulator of angiogenesis, particularly during the extensive tissue growth and remodeling that occur in utero. In the present study, we have investigated the role of VEGF during human fetal development by analyzing the distribution of VEGF messenger RNA as well as the tissue- and cell-specific localization of VEGF peptide in the human midgestation (16-22 weeks) fetus. As a comparison, we conducted parallel studies on several human adult tissues. Messenger RNA encoding VEGF was detected in all fetal tissues studied and was most abundant in human fetal lung, kidney, and spleen; moderately abundant in heart, adrenal, pancreas, intestine, liver, testis, skin, muscle, and brain; and minimally detected in thymus and placenta. VEGF peptide, detected by immunohistochemistry, always was intracytoplasmic and localized principally in epithelial cells and myocytes, including the smooth muscle cells lining blood vessels. VEGF was not detected in vascular endothelial cells. As the cellular localization of VEGF in several human adult tissues was similar to that found in the cognate fetal tissues, VEGF is probably important not only in angiogenesis, but also in the maintenance of existing vessels. As VEGF was localized primarily in epithelial cells and myocytes and not in endothelial cells, these data are consistent with a paracrine mechanism of action whereby VEGF secreted by nonendothelial cells modulates activities in adjacent vascular endothelium.