Management of monoamniotic twin pregnancies: Where, when, how?

Management difficulties for monochorionic monoamniotic (MCMA) twin pregnancy reflect the absence of high-quality research into optimal types of monitoring, essential as MCMA twins have a high risk of intrauterine and neonatal death with perinatal mortality. D'Antonio et al's meta-analysis and the MonoMono study published in 2019, investigated the impact of monitoring location, out- or in-patient, of MCMA pregnancies and concluded that no specific management location is associated with improvement in prognosis. To evaluate the optimal timing for delivery of MCMA pregnancies, Van Mieghem and Chitrit carried out retrospective studies comparing gestational age of intrauterine death and risk of neonatal complication. The crossover point between the propective risk of intrauterine fetal death and neonatal complication was found at 32,33 weeks of gestation (WG), in accordance with American College of Obstetricians and Gynecologists and Royal College of Obstetricians and Gynaecologists recommendations but inclusion of complicated pregnancies and analysis of fetuses individually may be regarded as a bias. The majority of studies of MCMA pregnancies focused on elective scheduled cesareans, with only rare retrospective studies reporting on vaginal delivery. Of these, two recent studies carried out by French teams suggest that vaginal deliveries may be as safe as cesarean births for MCMA twin pregnancies when specific criteria are met. In summary, concerning MCMA pregnancies, prognosis is not found to improve with inpatient management, optimal timing for delivery is at approximately 33 GW and vaginal delivery should not be excluded.

Successful outcome after spontaneous first trimester intra-amniotic haematoma and early preterm premature rupture of membranes.

A spontaneous intra-amniotic haematoma is a rare cause of preterm premature rupture of the membranes (PPROM) but can have significant fetal and maternal consequences. It has previously been reported to occur in the second and third trimesters but not in an earlier gestation. We present a case that presented acutely in the first trimester of pregnancy, which leads to early PPROM at 15 weeks and spontaneous preterm delivery at 28 weeks of gestation. There were no maternal complications during the pregnancy.

Intra-Amniotic Hemorrhage Imitating Gastroschisis: A Case Report and Review of the Literature.

BACKGROUND A spontaneous intra-amniotic hemorrhage is rarely encountered during pregnancy. The correct diagnosis and management are problematic because of the infrequency of this condition and the high likelihood of a misdiagnosis. CASE REPORT A primigravida with an uncomplicated pregnancy and a normal targeted ultrasound presented late in the second trimester of pregnancy with antepartum bleeding of unknown origin. A repeat ultrasound was suggestive of an abdominal wall defect (gastroschisis). The patient continued to have antepartum bleeding and developed uterine contractions and abdominal pain necessitating frequent visits to labor and delivery. An MRI ruled out gastroschisis and diagnosed intra-amniotic hematoma. The patient presented with acute abdominal pain and was clinically considered to be having an abruption, and was delivered by cesarean. Old blood was noted in the abdominal cavity and within the uterine cavity. At the time of the cesarean, an area of intra-amniotic hematoma was identified, as well as a retroplacental blood clot. CONCLUSIONS An intra-amniotic hematoma is unusual and may be misdiagnosed. MRI may be helpful in determining the correct diagnosis and subsequent management.

Interleukin-33 in the human placenta.

OBJECTIVE: Interleukin-33 (IL-33) is the newest member of the IL-1 cytokine family, a group of key regulators of inflammation. The purpose of this study was to determine whether IL-33 is expressed in the human placenta and to investigate its expression in the context of acute and chronic chorioamnionitis. METHODS: Placental tissues were obtained from five groups of patients: 1) normal pregnancy at term without labor (n = 10); 2) normal pregnancy at term in labor (n = 10); 3) preterm labor without inflammation (n = 10); 4) preterm labor with acute chorioamnionitis and funisitis (n = 10); and 5) preterm labor with chronic chorioamnionitis (n = 10). Immunostaining was performed to determine IL-33 protein expression patterns in the placental disk, chorioamniotic membranes, and umbilical cord. mRNA expression of IL-33 and its receptor IL1RL1 (ST2) was measured in primary amnion epithelial and mesenchymal cells (AECs and AMCs, n = 4) and human umbilical vein endothelial cells (HUVECs, n = 4) treated with IL-1ß (1 and 10 ng/ml) and CXCL10 (0.5 and 1 or 5 ng/ml). RESULTS: 1) Nuclear IL-33 expression was found in endothelial and smooth muscle cells in the placenta, chorioamniotic membranes, and umbilical cord; 2) IL-33 was detected in the nucleus of CD14+ macrophages in the chorioamniotic membranes, chorionic plate, and umbilical cord, and in the cytoplasm of myofibroblasts in the Wharton's jelly; 3) acute (but not chronic) chorioamnionitis was associated with the presence of IL-33+ macrophages in the chorioamniotic membranes and umbilical cord; 4) expression of IL-33 or IL1RL1 (ST2) mRNA in AECs was undetectable; 5) IL-33 mRNA expression increased in AMCs and HUVECs after IL-1ß treatment but did not change with CXCL10 treatment; and 6) IL1RL1 (ST2) expression decreased in AMCs and increased in HUVECs after IL-1ß but not CXCL10 treatment. CONCLUSIONS: IL-33 is expressed in the nucleus of placental endothelial cells, CD14+ macrophages, and myofibroblasts in the Wharton's jelly. IL-1ß can induce the expression of IL-33 and its receptor. Protein expression of IL-33 is detectable in macrophages of the chorioamniotic membranes in acute (but not chronic) chorioamnionitis.

Placental characteristics of monochorionic diamniotic twin pregnancies in relation to perinatal outcome.

To study placental characteristics in relation to perinatal outcome in 150 pairs of monochorionic diamniotic (MCDA) twins. Between January 1998 and January 2007 150 pairs of MCDA twins were delivered in the University Medical Center, Utrecht, The Netherlands. Mortality, neonatal morbidity and birth weight discordancy were studied in relation to type of anastomoses, type and distance between cord insertions and placental sharing. From 14 weeks onwards, there were 45 (15.0%) perinatal deaths. We found no clear relationship between perinatal mortality and type of anastomoses, distance between cord insertions and placental sharing. Perinatal mortality was significantly increased in the presence of velamentous cord insertion (OR 3.65, 95% CI 1.83-7.28). Data concerning neonatal morbidity were similar. TTTS occurred predominantly in the presence of AV-anastomoses without compensating superficial AA-anastomoses (p=0.005) and occurred more frequently in the presence of velamentous cord insertion (OR 1.79, 95% CI 0.94-3.44). Twins with unequal shared placentas had significantly more often severe birth weight discordancy, although only in the presence of AA-anastomoses (OR 4.09, 95% CI 1.74-9.63). If AA-anastomoses were absent in the unequally shared placenta, there was no relation between severe birth weight discordancy and unequal sharing of the placenta (OR 1.06, 95% CI 0.08-13.52). In MCDA twins, placental characteristics determine perinatal outcome, occurrence of TTTS and fetal growth. Prenatal identification of these characteristics by ultrasound may alter counselling and intensity of pregnancy surveillance.

Hysteroscopic management of cervical pregnancy with intrauterine irrigation with H2O2.

The objective of our study was to introduce a safe and effective method for the treatment of cervical pregnancy. This was a case series and took place at the minimal invasive surgery clinic at Soonchunhyang University Bucheon Hospital, Bucheon, Korea. From April 2005 through October 2007, a total of 10 patients with cervical pregnancy underwent hysteroscopic surgery. An 18 F foley catheter was inserted through the cervical canal after dilation, and the uterine cavity was irrigated with 3.5% H2O2 300 mL for prevention of unrelenting bleeding. Hysteroscopic removal of cervical gestation under ultrasonographic guidance was performed uneventfully. All patients had an unremarkable postoperative course. Serum beta-human chorionic gonadotropin levels were monitored after surgery and exhibited optimal decrease. All cases had histologic confirmation of products of conception removed from the cervix. The estimated blood loss was less than 50 mL. All patients were discharged after the first or second postoperative day. Hysteroscopic removal of cervical pregnancy with H2O2 is very useful and safe and preserves fertility.

Placentation in dolphins from the Amazon River Basin: the Boto, Inia geoffrensis, and the Tucuxi, Sotalia fluviatilis.

A recent reassessment of the phylogenetic affinities of cetaceans makes it timely to compare their placentation with that of the artiodactyls. We studied the placentae of two sympatric species of dolphin from the Amazon River Basin, representing two distinct families. The umbilical cord branched to supply a bilobed allantoic sac. Small blood vessels and smooth muscle bundles were found within the stroma of the cord. Foci of squamous metaplasia occurred in the allanto-amnion and allantochorion. The interhemal membrane of the placenta was of the epitheliochorial type. Two different types of trophoblastic epithelium were seen. Most was of the simple columnar type and indented by fetal capillaries. However, there were also areolar regions with tall columnar trophoblast and these were more sparsely supplied with capillaries. The endometrium was well vascularised and richly supplied with actively secreting glands. These findings are consistent with the current view that Cetacea are nested within Artiodactyla as sister group to the hippopotamids.

Vascularization is more delayed in amniotic membrane graft than conjunctival autograft after pterygium excision.

PURPOSE: To monitor the development of graft vascularization by indocyanine green angiography (ICGA) after pterygium excision with limbal-conjunctival autograft transplantation (LCAT) and amniotic membrane transplantation (AMT). DESIGN: Randomized prospective interventional case series. METHODS: Anterior segment ICGA findings at one, seven, and 30 days after surgery were evaluated in eyes with primary pterygium treated by excision and LCAT (14 eyes of 14 patients) or AMT (13 eyes of 13 patients), and the results for graft vascularization were compared. RESULTS: All conjunctival autografts showed early vascularization from underlying episcleral vascular bed as multiple hyperfluorescent foci at the graft margin on postoperative day 7; graft vascularization was complete on postoperative day 30. In contrast, all grafts in AMT group showed no graft vascularization at any stage after pterygium surgery. This delay of vascularization with AMT persisted until one month after surgery. CONCLUSIONS: Anterior segment ICGA is useful to monitor graft vascularization after pterygium surgery. Graft health after LCAT may be demonstrated by early graft vascularization and perfusion; however, there is a delay in graft vascularization after AMT that may be related to the antiangiogenic effects of the membrane. Further study is needed to demonstrate the relationship between this delay of graft perfusion and early postoperative pterygium recurrence.

Vascular distribution patterns in monochorionic twin placentas.

Several recent publications have focused on the association between the occurrence of twin-to-twin transfusion syndrome (TTTS) in diamniotic-monochorionic twins and the presence of a number of selected anatomic placental characteristics (distribution of vascular territory, cord insertion, type and number of inter-twin anastomoses). In contrast, the potential importance of the vascular distribution patterns of the individual twins remains to be elucidated. Based on its gross architectural distribution pattern, chorionic vasculature is traditionally described as disperse, magistral or mixed. The aim of this study was (1) to determine the relative prevalence of these vascular distribution patterns in monochorionic twin placentas, and (2) to correlate these patterns with the presence of TTTS and known anatomic placental features linked to TTTS. The placentas of 89 consecutive diamniotic-monochorionic twins (15 with TTTS, 74 without TTTS), examined at Women and Infants Hospital, were studied. Disperse vascular patterns were seen in 53% of twins, and magistral or mixed patterns in 47%. The prevalence of magistral/mixed vascular patterns was significantly higher in TTTS gestations than in non-TTTS gestations (60% versus 44%, P<0.05) and, in TTTS gestations, much higher in donor twins than in recipient twins (87% versus 33%, P<0.005). A strong association was found between the presence of magistral/mixed patterns and marginal/velamentous cord insertion, low number of inter-twin anastomoses, and uneven distribution of the vascular territories. These findings suggest that the magistral/mixed vascular distribution pattern may represent an important placental architectural feature contributing to the complex pathophysiology of TTTS.

Neovascularization of the amniotic membrane as a biological immune barrier.

The clinical application of islet transplantation is limited due to the limited source and the morbidity of systemic immunosuppression to prevent rejection. The two problems can be solved by using encapsulated islets. We have used amniotic membranes as biocompatible natural immune barriers. The objective of this study was to assess the revascularization of the membrane, which is necessary to ensure islet viability when the membrane is used for islet encapsulation. The amniotic membranes, obtained from full-term pregnant female dogs, were molded to form macrocapsules, which were implanted in the peritoneal cavity. The capsules were removed after 3, 10, 15, and 30 days and examined histopathologically using hematoxylin and eosin and by immunohistochemistry for neovascularization using factor VIII to detect angiogenesis. Upon histopathological examination, all specimens showed localized, moderate inflammation and congested blood vessels with no thrombosis or rejection. There was a mild degree of fibroblast proliferation starting from day 10 to day 30. Immunohistochemical staining revealed that the number of blood vessels was 7, 11, 13, 10 per high-power microscopic field on days 3, 10, 15, and 30, respectively. We concluded from this study that implanted amniotic sac capsules were vascularized within the omental tissue from day 10 on with significant blood vessel formation starting on day 3 by immunohistochemical study.

Chorionic plate vessels as an origin of amniotic fluid neutrophils.

The present study was conducted to investigate the potential anatomical source of amniotic fluid neutrophils. Microdissection of neutrophils from the chorioamnion of the fetal membranes and the amnion of the chorionic plates of 10 preterm placentas with acute chorioamnionitis was performed and the genotypes of the neutrophils were compared with those of the mother and fetus using polymerase chain reaction of nine autosomal STR loci. In separate analyses, we reviewed eight cases of fetal autopsies with increased amniotic fluid neutrophils for the presence of neutrophils in the alveoli, and also analyzed the relationship between the amniotic fluid white blood cell (WBC) count and the histological pattern of placental inflammation. The genotypes of all of the neutrophils found in the chorioamnion of the fetal membrane matched those of the mother (n = 10). The genotypes of neutrophils found in the chorionic plate were of mixed maternal and fetal origin (n = 4). In the autopsy series of the fetuses with amniotic fluid WBC (n = 8), only five cases showed neutrophils in the alveolar space, while all the placentas had chorioamnionitis. There was no significant difference in amniotic fluid WBC count between the cases with or without acute membranitis, while among the cases with placental inflammation, those with inflammation of the chorionic plate had a significantly higher amniotic fluid WBC count than both the membranitis-only cases (P < 0.001) and the membranitis and funisitis cases (P < 0.05). These results imply that fetal vasculature at the chorionic plate is the main source of amniotic fluid neutrophils, especially in the cases without funisitis.

Modelling the influence of amnionicity on the severity of twin-twin transfusion syndrome in monochorionic twin pregnancies.

Clinical treatment for diamniotic-monochorionic twin-twin transfusion syndrome (TTTS) may include conversion of diamniotic pregnancies to a monoamniotic-monochorionic state by disrupting the amnion septum. We sought to test the underlying hypothesis, i.e. that a monoamniotic state reduces the severity of TTTS. With use of our previously developed mathematical model of two equal fetoplacental circulatory units connected by various sizes and types of placental anastomoses, we compared the haemodynamic and amniotic fluid dynamics of monoamniotic and diamniotic twins that develop TTTS. We used three anastomotic patterns that produce severe, moderate or mild forms of TTTS, respectively, in our diamniotic-monochorionic twin model. Monoamnionicity was modelled by adding the two amniotic fluid volumes and using the volume-averaged amniotic fluid osmolality. The results were as follows: for severe TTTS, small differences develop between diamniotic and monoamniotic donor twins in fetal urine production, swallowed volume, blood volume, blood pressures, net fetofetal transfusion, and blood and amniotic fluid osmolality. However, the circulatory imbalance between the monoamniotic twins deteriorates similar to that of diamniotic twins. The pathophysiological differences tend to disappear for milder TTTS. In conclusion, our model suggests that the uncommon finding of TTTS in monoamniotic twins is not due to the presence of a single amniotic sac. Rather, clinically significant differences in anastomotic patterns and the delayed or lack of identification of manifestations in monoamniotic twins account for the reduced rate of TTTS diagnosis. Based on these results we expect the clinical disruption of the amnion septum in diamniotic-monochorionic TTTS pregnancies to have only minimal benefits.

Automated evaluation of angiogenic effects mediated by VEGF and PlGF homo- and heterodimers.

The effects of growth factors on the blood vessel pattern of chick chorioallantoic membrane (CAM) were assessed with a fast and automated method (extended counting method, XCM; Sandau, 1996) that measures complexity, without assumptions about a fractal structure. XCM is a reliable measure of complexity not only in theory but also in practice: (1) it is robust with respect to thresholding; (2) it shows reduced variance due to pattern translation and rotation; (3) its properties come close to requirements of fractal geometry. It hence is superior to established fractal methods for distinguishing effects induced by various isoforms of vascular endothelial growth factor (VEGF121 and VEGF165), placenta growth factor (PlGF) isoforms, and control treatment. We here show that VEGF homo- and heterodimers and VEGF121/PlGF1 heterodimers increase vascular complexity, whereas PlGF1 and PlGF2 are not effective. PlGF1 and VEGF121 did not mutually influence each other when applied in adjacent fields on the same CAM. Since blood vessels in the CAM originate via nonfractal growth processes, their growth should be analyzed accordingly.

Measurement of blood flow by the 133Xe clearance technique to grafts of amnion used in vestibuloplasty.

The purpose of this study was to determine the blood supply to lyophilized amniotic membranes when used as graft material in vestibuloplasties. 133Xe clearance technique was used to measure the blood flow to the grafts. A total of 20 patients had either Clark (10) or Kazanjian (10) vestibuloplasties. The blood flow was determined at 2-3 days preoperatively and at 10 and 30 days postoperatively. The preoperative mandibular anterior alveolar mucosal blood flow was 34.4 +/- 10.7 and 23.1 +/- 13.1 ml/100 g/min for the Clark and Kazanjian groups, respectively. Ten days after vestibuloplasty operation with lyophilized amniotic membrane graft application the blood flow to the graft increased to 56.8 +/- 45.4 and 62.6 +/- 30.4 ml/100 g/min for the Clark and Kazanjian groups, respectively. The corresponding values at 30 days postoperatively were 24.6 +/- 10.2 and 22.2 +/- 9.2 ml/100 g/min, indicating the return to normal levels. The changes in blood flow as a function of time were statistically significant in each group (P<0.05). Our results demonstrated the angiogenic effect of lyophilized amniotic membranes until mucoid degeneration after 10-15 days.

Arginine vasopressin-induced changes in blood flow to the ovine chorion, amnion, and placenta across gestation.

OBJECTIVES: To determine whether physiological increases in fetal plasma arginine vasopressin (AVP) concentration alter blood flow rates to the ovine chorion, amnion or placenta, and to determine whether these AVP-induced changes in flow are dependent on gestational age. METHODS: Colored microspheres (15.10 +/- 0.02 microns (standard deviation) were infused into the superior vena cava before and at 30, 75, and 125 minutes of an intravenous AVP infusion (3 ng/min/kg) in 9 chronically catheterized fetal sheep between 103 days to 141 days gestation (term = 147 days). Chorion, amnion, and placental cotyledons were removed, and microspheres were counted to determine blood flow rates. RESULTS: Fetal arterial pressure (FAP) increased (analysis of variance, P < .0001) and heart rate (FHR) decreased (P < .0001) during the infusion, with responses greater in older (> 125 days) compared to younger (< 125 days) fetuses (P < .001). Chorionic blood flow rate increased by 19 +/- 7% at 30 min of the AVP infusion, and declined by 18 +/- 6% at 75 min, and 32 +/- 5% at 125 min (P < .0001). Similarly, fetal placental blood flow rate increased by 20 +/- 7% at 30 min of infusion, and declined, in parallel with chorionic blood flow rates, by 6 +/- 4% at 75 min, and 17 +/- 4% at 125 min (P < .0001). Amniotic blood flow rate did not change significantly during the infusion. The membranous and placental blood flow rate responses to AVP infusion did not depend upon gestational age. CONCLUSIONS: Physiologic increases in plasma AVP concentration induce an early increase in chorionic blood flow followed by a gradual decrease which parallels similar changes in placental blood flow rate. Unlike FAP and FHR, these blood flow changes are not gestational age-dependent. AVP-induced blood flow changes could play an important role in determining abnormalities of amniotic fluid volume observed clinically in some stressed fetuses.

Changes in blood flow to the ovine chorion and amnion across gestation.

OBJECTIVE: The purpose of this study was to determine the developmental changes in blood flow to the network of fetal microvessels in ovine chorion and amnion. METHODS: Colored microspheres (15.10 +/- 0.02 [standard deviation] mu in diameter) were infused into the superior vena cava in nine chronically catheterized fetal sheep with gestational ages ranging from 103-141 days (term 147). After euthanasia, chorion, amnion, and cotyledons were separated and microspheres were counted to determine blood flow rates. Standard correlation and regression analyses were used to analyze the data. RESULTS: Chorionic blood flow rate increased linearly (r = 0.82, P = .006) from 12 mL/minute at 103 days' gestation to 70 mL/minute at 141 days, and averaged 10.3 +/- 1.2% of the total umbilical blood flow. Weight-normalized chorionic flow (18.4 +/- 2.0 [standard error] mL/minute/kg of fetus) did not change significantly across gestation. Absolute and weight-normalized blood flow to the amnion (0.82 +/- 0.31 mL/minute and 0.34 +/- 0.11 mL/minute/kg fetus) increased with advancing gestation until 130 days and declined thereafter. Absolute cotyledonary blood flow rate increased with gestational age (r = 0.81, P = .008), and weight-normalized cotyledonary blood flow decreased with advancing gestation (r = -0.89, P = .002). Absolute but not weight-normalized chorionic and cotyledonary blood flow rates correlated positively. CONCLUSIONS: Blood flow to the microvascular network in the ovine chorion is high and increases with advancing gestation. Blood flow to the amnion is low but not insignificant. Therefore, intramembranous exchange may play an increasingly important role in determining amniotic fluid volume and composition as gestation proceeds.

Color Doppler ultrasonography in the identification of communicating vessels in twin-twin transfusion syndrome and acardiac twins.

This study evaluates the role of color Doppler ultrasonography in the identification of the communicating placental vessels in pregnancies with twin-twin transfusion syndrome and acardiac twins. In 18 pregnancies with twin-twin transfusion syndrome and two with an acardiac twin, color Doppler studies of the placental vasculature were performed before fetoscopy for laser coagulation of the communicating vessels. In six cases of twin-twin transfusion syndrome the placental attachment of the intertwin membrane could be visualized. Pulsatile arterial blood flow was observed from the donor to the recipient twin that disappeared after laser therapy. In both cases of acardiac twins, one communicating vessel with pulsatile and another vessel with nonpulsatile blood flow in the opposite direction could be identified. Color Doppler imaging is unlikely to play a major role in assisting endoscopic laser separation of chorioangiopagus in patients with acute polyhydramnios, but it may prove to be useful in the early identification of pregnancies at risk of developing twin-twin transfusion syndrome.

Vascularization of the ovine amnion and chorion: a morphometric characterization of the surface area of the intramembranous pathway.

OBJECTIVE: The purpose of this study was to characterize the vascularization of the ovine amnion, allantois, and chorion. STUDY DESIGN: A white silicone vascular casting material was infused into both umbilical arteries of nine fetal sheep ranging in age from 58 to 142 days' gestation. A morphometric analysis of photomicrographs of the membranes was then performed with computerized image analysis techniques. RESULTS: After removal of the uterus, the fetus was surrounded by a layer of white silicone-filled microvessels in the chorion. The amniotic membrane after separation from the chorion was covered by a fine mesh of microvessels, whereas the allantois was avascular. The amniotic membrane readily separated into an outer vascularized layer and an inner, avascular layer containing the amnion. Approximately 50% of the surface of the chorionic membrane was covered by microvessels; this appeared independent of gestational age. At midgestation 30% of the surface of the amnion was covered by microvessels, and this decreased to 17% at 142 days. Relative to fetal weight, the amniotic and chorionic vascular surface areas decreased from 6 to 0.3 and from 15 to 1.5 cm2/gm fetal weight, respectively, over the last half of gestation. CONCLUSIONS: There is an extensive network of microscopic fetal blood vessels within the ovine chorion and covering the outer surface of the amnion. These vessels appear to be ideally situated to facilitate a direct exchange of water and solutes between amniotic or allantoic fluids and fetal blood through the intramembranous pathway.

In vitro angiogenesis on the human amniotic membrane: requirement for basic fibroblast growth factor-induced proteinases.

The role of basic fibroblast growth factor-(bFGF) induced proteinases in basement membrane (BM) invasion by bovine capillary endothelial (BCE) cells was studied using a quantitative in vitro assay previously described (Mignatti et al., 1986). 125I-iododeoxyuridine-labeled BCE cells were grown for 72 h on the human amnion BM, and cell invasion was determined by measuring the radioactivity associated with the tissue after removal of the noninvasive cell layer. BCE cells were noninvasive under normal conditions. Addition of human bFGF to either the BM or to the stromal aspect of the amnion induced BCE cell invasion with a dose-dependent response. This effect was maximal in the presence of 70 ng/ml bFGF, and was inhibited by anti-FGF antibody. Transforming growth factor beta, as well as plasmin inhibitors and anti-tissue type plasminogen activator antibody inhibited BCE cell invasion. The tissue inhibitor of metalloproteinases, 1-10 phenanthroline, anti-type IV and anti-interstitial collagenase antibodies had the same effect. On the contrary, anti-stromelysin antibody and Eglin, an inhibitor of elastase, were ineffective. The results obtained show that both the plasminogen activator-plasmin system and specific collagenases are involved in the invasive process occurring during angiogenesis.