Uterine smooth muscle cells express functional receptors (flt-1 and KDR) for vascular permeability factor/vascular endothelial growth factor.

Vascular permeability factor (VPF), also known as vascular endothelial growth factor (VEGF), is an angiogenic factor with important roles in tumor growth, wound healing, and inflammation. VPF/VEGF interacts with endothelial cells by way of two high-affinity receptor tyrosine kinases: flt-1 and KDR. The vast majority of published studies have described expression of the VPF/VEGF receptors only in endothelial cells, and the statement is frequently made that these receptors are endothelial-cell-specific. In this study, we detected mRNA for flt-1 and KDR by in situ hybridization in smooth muscle cells in sections of the wall of the uterus. To confirm these unexpected findings, smooth muscle cells from the uterus and, as a control, from the colon were isolated, characterized, and cultured. Both uterine and colonic smooth muscle cells in culture expressed VPF/VEGF, but only smooth muscle cells from the uterus expressed mRNA for flt-1 and KDR by Northern analysis and in situ hybridization. Cell culture extracts of uterine but not colonic smooth muscle cells were also positive for flt-1 by Western analysis. Moreover, cultured uterine but not clonic smooth muscle cells phosphorylated the flt-1 receptor and proliferated strongly in response to added VPF/VEGF. This is one of the first rigorous demonstrations that a normal cell type other than endothelial cells can express functional receptors for VPF/VEGF in vivo and in vitro, suggesting that VPF/VEGF may have important, previously unsuspected roles on cell types other than endothelium.

Brown LF ，Detmar M ，Tognazzi K ，Abu-Jawdeh G ，Iruela-Arispe ML ... -  《LABORATORY INVESTIGATION》

Strong expression of vascular permeability factor (vascular endothelial growth factor) and its receptors in ovarian borderline and malignant neoplasms.

Angiogenesis is a critical factor in the growth, progression, and metastatic spread of solid tumors. Furthermore, angiogenesis has been correlated with prognosis in patients with ovarian cancer. The pathogenesis of the angiogenic events in ovarian cancer, however, are not well defined. Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) is a multifunctional cytokine that has been shown to be an important regulator of tumor angiogenesis. The purpose of the present study was to define the expression of VPF/VEGF and its receptors flt-1 and KDR in ovarian tumors. Four specimens of normal ovarian cortex and 41 specimens of benign (4), borderline (8), and malignant (29) ovarian tumors were studied by in situ hybridization, and in some cases by immunohistochemical analysis. VPF/VEGF protein was also determined by an immunofluorometric assay in cyst fluids obtained from 11 patients, including 7 benign, 2 borderline, and 2 malignant tumors. VPF/VEGF mRNA and protein were expressed by the neoplastic cells in all of the malignant tumors evaluated, with the majority of tumors (28 of 29) showing strong expression of mRNA. Serous borderline tumors had variable VPF/VEGF mRNA expression, with two of six cases showing focal strong expression and four showing low-level expression. No definite expression of VPF/VEGF was seen in two cases of mucinous borderline tumors. No strong expression of VPF/VEGF mRNA was observed in normal ovarian cortex, including surface epithelium, or benign tumors. Substantially higher VPF protein concentrations were detected in cyst fluids of the two malignant (60, 440 pM) and two borderline tumors (210, 590 pM) than in the seven benign serous cysts (mean, 10 +/- 3 pM). In addition, microvascular endothelial cells strongly expressed mRNA of the VPF/VEGF receptors flt-1 and KDR and immunostained for VPF/VEGF protein in the majority of malignant and borderline tumors examined. These findings suggest that VPF/VEGF plays an important role in the angiogenesis associated with ovarian neoplasms.

Abu-Jawdeh GM ，Faix JD ，Niloff J ，Tognazzi K ，Manseau E ，Dvorak HF ，Brown LF ... -  《LABORATORY INVESTIGATION》

Expression of vascular permeability factor (vascular endothelial growth factor) and its receptors in adenocarcinomas of the gastrointestinal tract.

Vascular permeability factor (VPF) is one of the most potent known inducers of microvascular hyperpermeability; in addition, it is a selective endothelial cell growth factor, hence its alternate name, vascular endothelial growth factor. VPF exerts its actions on the microvasculature by interacting with specific endothelial cell receptors. VPF is expressed by many transplantable animal tumors, by tumor cell lines in culture, and by certain normal cells in situ. The purpose of the present investigation was to determine whether and with what consistency VPF and its endothelial cell receptors are expressed in primary autochthonous human tumor of gastrointestinal tract origin, as determined by in situ hybridization and immunohistochemistry. Twenty-one primary adenocarcinomas (17 colon, 2 stomach, 1 small bowel, and 1 pancreas) were studied. The malignant epithelial cells expressed VPF mRNA strongly, in contrast to normal epithelium, hyperplastic polyps, and adenomas, which expressed little or no VPF mRNA. VPF expression was further increased in tumor cells immediately adjacent to zones of tumor necrosis; in such areas, occasional stromal cells also expressed VPF mRNA. In the ten colon carcinomas studied, tumor cells stained for VPF protein by immunohistochemistry. The endothelial cells of nearby stromal blood vessels also stained for VPF by immunohistochemistry and in addition expressed mRNAs encoding the VPF receptors flt-1 and kdr as determined by in situ hybridization. Endothelial cells away from the tumor did not stain for VPF and no definite mRNA expression for flt-1 or kdr was detected by in situ hybridization. The ganglion cells of the myenteric plexus of normal bowel expressed VPF mRNA and protein. These data indicate that primary autochthonous human tumors of gastrointestinal origin regularly express both VPF mRNA and VPF protein and that adjacent stromal vessels express mRNAs for both known VPF receptors. VPF is likely to contribute to tumor growth by promoting angiogenesis and stroma formation, both directly, through its action as an endothelial cell growth factor, and indirectly, by increasing vascular permeability, thereby leading to plasma protein extravasation, fibrin deposition, and the eventual replacement of the resulting matrix with vascularized stroma.

Brown LF ，Berse B ，Jackman RW ，Tognazzi K ，Manseau EJ ，Senger DR ，Dvorak HF ... -  《CANCER RESEARCH》

Immunohistochemical localization of vascular endothelial growth factor (VEGF) and its two specific receptors, Flt-1 and KDR, in the porcine placenta and non-pregnant uterus.

Placental angiogenesis and growth are crucial elements in embryonic and later fetal development. Vascular endothelial growth factor (VEGF) and its specific receptors Flt-1 (VEGFR-1) and KDR (VEGFR-2) compose potent ligand receptor systems involved in angiogenesis and microvascular hyperpermeability. In the present immunohistochemical study, VEGF, Flt-1 and KDR were localized in uterus of cyclic non-pregnant pigs and in the porcine epitheliochorial placenta throughout gestation. Emphasis was placed on early gestational stages, where morphological studies have demonstrated extensive angiogenesis during initial placentation. The results revealed a high correlation in spatiotemporal distribution between the ligand and its receptors and a surprising demonstration of VEGF receptors in several non-endothelial cells. In non-pregnant pigs, VEGF, Flt-1 and KDR exhibited moderate to intense staining in uterine luminal epithelium and smooth muscle cells of the vessel walls. Endothelial cells of arteries and arterioles revealed labelling for Flt-1 and KDR, whereas the uterine glandular epithelium displayed intense KDR immunoreactivity at the late luteal phase. During gestation the uterine luminal epithelium demonstrated weak ligand and receptor immunostaining in the first half of early gestation [< or = 21 days post coitus (p.c.)], whereas later stages (> or = 21 days p.c.) displayed intense immunolabelling. Endothelial cells, smooth muscle cells of the vessel walls and uterine glandular epithelium revealed intense ligand and receptor immunoreactivity throughout gestation. In the fetal part of the placenta, VEGF, Flt-1 and KDR immunostaining displayed moderate to intense reactivity in the trophoblast throughout gestation, except during the second half of early gestation (days 21-30 p.c.). Fetal vessel walls were also immunopositive for VEGF, Flt-1 and KDR. Taken together, the results imply that the VEGF, Flt-1 and KDR ligand receptor system participate in the regulations of porcine placentation and that it in addition to its angiogenic properties also may influence the cellular differentiation and transport capabilities in uterine luminal as well as glandular epithelium and the trophoblast.

Winther H ，Ahmed A ，Dantzer V 《PLACENTA》

Expression of vascular permeability factor (vascular endothelial growth factor) and its receptors in endometrial carcinoma.

Solid tumors, including endometrial carcinomas, must induce a vascular stroma to grow beyond a minimal size. The mechanisms responsible for angiogenesis in endometrial carcinoma, however, are not well defined. Vascular permeability factor (VPF), also known as vascular endothelial growth factor (VEGF), is a multifunctional cytokine that is an important regulator of tumor angiogenesis. We evaluated VPF/VEGF mRNA and protein expression, as well as VPF/VEGF receptor mRNA expression, in endometrial carcinoma. Fourteen examples of endometrial carcinoma were evaluated by in situ hybridization; in 7 cases, benign atrophic endometrium from the same patient was also examined. Histologic sections were subjected to in situ hybridization using 35S-labeled riboprobes specific for VPF/VEGF and, in a subset of cases, riboprobes specific for the VPF/VEGF receptors flt-1 and KDR. In addition, ten examples of endometrial carcinoma were evaluated for VPE/VEGF protein expression by immunohistochemistry. All 14 examples of endometrial carcinoma studied by in situ hybridization exhibited focal strong VPF/VEGF mRNA expression by tumor cells. In addition, the endothelial cells of surrounding microvessels strongly expressed flt-1 and KDR mRNAs in all ten cases examined. In contrast, no strong expression of VPF/VEGF, flt-1, or KDR mRNA was observed in the seven examples of benign atrophic endometrium studied. All ten cases of endometrial carcinoma studied by immunohistochemistry exhibited strong VPF/VEGF protein expression by tumor cells. These observations suggest that VPF/VEGF is an important angiogenic factor in endometrial carcinoma.

Guidi AJ ，Abu-Jawdeh G ，Tognazzi K ，Dvorak HF ，Brown LF ... -  《CANCER》