Decrease In Vascular Permeability Research Articles

ABT-869, a Multitargeted Receptor Tyrosine Kinase Inhibitor, Reduces Tumor Microvascularity and Improves Vascular Wall Integrity in Preclinical Tumor Models

N-[4-(3-amino-1H-indazol-4-yl)phenyl]-N1-(2-fluoro-5-methyl phenyl)-urea (ABT-869) is a novel multitargeted receptor tyrosine kinase inhibitor that demonstrates single-agent activity in preclinical studies and has undergone phase I and II clinical trials. We characterized the mechanism of action of ABT-869 by examining vascular changes after treatment (25 mg/kg per day) in HT1080 fibrosarcoma and SW620 colon carcinoma cells, using immunohistochemistry, dynamic contrast enhanced-magnetic resonance imaging (DCE-MRI), and hypoxic protein detection. We observed the inhibition of vascular endothelial growth factor receptor 2 and platelet-derived growth factor receptor β phosphorylation in both tumors and changes in tumor vasculature. Reductions in microvessel density and diameter were observed. Vascular-wall integrity was assessed by colocalization of pericytes and basement membrane. Although both microvessel density and total number of pericytes decreased with treatment, the percentage of pericyte coverage on remaining vessels significantly increased. These data suggest the selective ablation of microvessels lacking pericyte coverage. Functional vascular measures DCE-MRI and hypoxia formation were also tested. After 2 days of treatment on the HT1080 model, vascular permeability, K(trans), was reduced by >60% and hypoxic tumor fraction was significantly decreased, which was also seen in the SW620 tumors after 4 days of treatment. Taken together, decreases in vascular permeability and changes in vascular integrity observed in these studies define the mode of action of ABT-869 and may aid in optimizing the timing of therapeutic window for combination therapies.

Targeted Therapy of VEGFR2 and EGFR Significantly Inhibits Growth of Anaplastic Thyroid Cancer in an Orthotopic Murine Model

Anaplastic thyroid carcinoma (ATC) is one of the most lethal human cancers with a median survival of 6 months. The inhibition of epidermal growth factor receptor (EGFR) alone, or with VEGF receptor 2 (VEGFR2), represents an attractive approach for treatment of ATC. Several reports have examined agents that target these receptors. However, with the misidentification of as many as 60% of all commonly used ATC cell lines, the significance of these past findings is unclear. Cell lines authenticated by short tandem repeat profiling were selected to establish xenograft tumors in an orthotopic murine model of ATC. These mice were then treated with vandetanib to evaluate its effects on ATC tumor growth. Dynamic contrast-enhanced (DCE) MRI was utilized to measure the impact of vandetanib on tumor vasculature. Vandetanib inhibited tumor growth of the ATC cell lines Hth83 and 8505C in vivo by 69.3% (P < 0.001) and 66.6% (P < 0.05), respectively, when compared with control. Significant decreases in vascular permeability (P < 0.01) and vascular volume fraction (P < 0.05) were detected by DCE-MRI in the orthotopic xenograft tumors after 1 week of treatment with vandetanib as compared with control. The inhibition of EGFR and VEGFR2 by vandetanib and its tremendous in vivo antitumor activity against ATC make it an attractive candidate for further preclinical and clinical development for the treatment of this particularly virulent cancer, which remains effectively untreatable. Vandetanib disrupts angiogenesis and DCE-MRI is an effective method to quantify changes in vascular function in vivo.

Magnetic Resonance Imaging Defines Cervicovaginal Anatomy, Cancer, and VEGF Trap Antiangiogenic Efficacy in Estrogen-Treated K14-HPV16 Transgenic Mice

Noninvasive detection of dysplasia provides a potential platform for monitoring the efficacy of chemopreventive therapy of premalignancy, imaging the tissue compartments comprising dysplasia: epithelium, microvasculature, and stromal inflammatory cells. Here, using respiratory-gated magnetic resonance imaging (MRI), the anatomy of premalignant and malignant stages of cervical carcinogenesis in estrogen-treated K14-HPV16 transgenic mice was noninvasively defined. Dynamic contrast enhanced (DCE)-MRI was used to quantify leakage across premalignant dysplastic microvasculature. Vascular permeability as measured by DCE-MRI, K(trans), was similar in transgenic (0.053 +/- 0.020 min(-1); n = 32 mice) and nontransgenic (0.056 +/- 0.029 min(-1); n = 17 mice) animals despite a 2-fold increase in microvascular area in the former compared with the latter. DCE-MRI did detect a significant decrease in vascular permeability accompanying diminution of dysplastic microvasculature by the antiangiogenic agent, vascular endothelial growth factor Trap (K(trans) = 0.052 +/- 0.013 min(-1) pretreatment; n = 6 mice versus K(trans) = 0.019 +/- 0.008 min(-1) post-treatment; n = 5 mice). Thus, we determined that the threshold of microvessel leakage associated with cervical dysplasia was <17 kDa and highlighted the potential of DCE-MRI to noninvasively monitor the efficacy of antiangiogenic drugs or chemoprevention regimens targeting the vasculature in premalignant cervical dysplasia.

Bevacizumab and Irinotecan in the Treatment of Recurrent Malignant Gliomas

Rapidly dividing glioma cells maintain adequate oxygen and nutrient delivery through co-opting existing host blood vessels or promoting the formation of new vessels, a process called angiogenesis. Vascular endothelial growth factor is a mediator of hypoxia-induced endothelial cell proliferation and migration and is highly expressed in gliomas, where it acts as a potent regulator of angiogenesis. The use of vascular endothelial growth factor receptor antagonists and vascular endothelial growth factor scavenging antibodies has generated excitement in neuro-oncology because of the rapid but reversible decrease in vascular permeability. This decrease in vascular permeability is marked by a decrease in cerebral edema and a decrease in contrast enhancement visualized on magnetic resonance imaging. These effects on the tumor vasculature are mistakenly referred to as tumor responses because the historical method of measuring tumor response and progression was based on tumor size assessed by contrast permeability through a leaky blood brain barrier. Despite the difficulties in accurately measuring the effect of antivascular endothelial growth factor therapy on tumor viability, several studies confirm that the antivascular endothelial growth factor human monoclonal antibody bevacizumab combined with irinotecan can significantly improve 6-month progression free survival of patients with malignant gliomas compared with historical controls. The impact of cytotoxic chemotherapy on the efficacy of bevacizumab and the effect of this therapy on overall survival are important questions that remain to be answered.

Antiangiogenesis Treatment for Glioblastoma Multiforme: Challenges and Opportunities

Angiogenesis is a major hallmark of cancer cells, and glioblastomas are among the most angiogenic tumors. The cascade of angiogenesis is probably initiated by hypoxia, leading to the production of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). Both VEGF and bFGF have paracrine effects on endothelial cells, pericytes, or both, causing the formation of hyperpermeable tumor blood vessels. Advanced MRI techniques, such as dynamic contrast-enhanced, dynamic susceptibility, and arterial spin labeling MRI, have provided semiquantitative measurements of tumor vascular permeability and perfusion. A decrease in vascular permeability and perfusion can be detected after antiangiogenesis drug treatment, either with monoclonal antibody such as bevacizumab that sequesters VEGF, or small-molecule VEGF receptor tyrosine kinase inhibitors. Therefore, antiangiogenesis therapies are being increasingly adopted for treating glioblastomas. However, caution must be exercised because neural stem cells are also sensitive to antiangiogenesis drugs and the combined effect of ionizing radiation. This article summarizes 30 years of laboratory and clinical research on glioblastoma angiogenesis and discusses its underlying biology, clinical trial results, vascular neuroimaging, and the potential side effects of antiangiogenesis treatment.

Dual targeting of vascular endothelial growth factor (VEGF) with sorafenib and bevacizumab: Clinical and translational results

3542 Background: VEGF is a major mediator of angiogenesis in malignancy. We targeted the VEGF pathway in series by combining the VEGFR2/Raf kinase inhibitor sorafenib (S) with bevacizumab (B), a selective anti-VEGF antibody. We hypothesized that this combination would suppress angiogenesis and tumor growth, with corresponding changes in correlative dynamic imaging. Methods: Eligible pts had non-curable solid tumors, ECOG performance status 0–1, good end organ function, and no prior exposure to S or B. We enrolled pts at S 200 mg po BID and B 5 mg/kg IV q2weeks (dose level [DL] 1) and escalated each drug sequentially. The MTD was DL1; 24 additional pts were enrolled at this dose and randomized to single agent S (Arm 1) or B (Arm 2) for one month, then S+B thereafter. Serum samples for cytokine analysis, dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) and 18FDG- PET scans were performed at 0, 2, and 6 weeks. DCE-MRI and PET scans measured vascular permeability (Ktrans) and metabolic activity (standardized uptake values [SUV]), respectively. The Wilcoxon signed rank test was used to evaluate the differences between the paired imaging values at the test time points in an exploratory fashion. Results: 7/38 pts (6/14 ovarian ca pts and 1/3 renal cell ca pts) had partial responses (8+-20 mo; median 15). Grade 3–4 toxicities included hypertension (10), proteinuria (2, DLT), and thrombocytopenia (1, DLT). Common grade 2 toxicities were hand-foot syndrome (18), fatigue (13), infection (13), and hypertension (12). DCE-MRI Ktrans decreased with S+B compared to baseline (p&lt;0.03, both arms); no significant change occurred with either S or B treatment alone or between arms. No significant changes in PET SUV measurements occurred with treatment. Correlation of dynamic imaging parameters with response was not possible due to the limited patient numbers. Serial cytokine analysis is presently being performed and will be included at the time of presentation. Conclusions: S+B is active in solid tumors with manageable toxicity. DCE-MRI shows a decrease in vascular permeability with S+B treatment. A larger trial is necessary to evaluate if DCE-MRI changes can be predictive of response and is ongoing now in ovarian cancer. No significant financial relationships to disclose.

Lung inflammation and vascular remodeling after repeated allergen challenge detected noninvasively by MRI

Magnetic resonance imaging (MRI) has been used previously to follow noninvasively inflammatory processes in rat acute models of lung inflammation. Here the technique was applied to a model involving repeated intratracheal administration of ovalbumin (OA). Anatomical MRI was performed at different time points with respect to a single or multiple OA challenges in Brown Norway rats actively sensitized to the allergen. Vascular permeability was assessed using dynamic contrast-enhanced MRI (DCE-MRI). Bronchoalveolar lavage (BAL) fluid analysis and histology were performed to validate the MRI data. The time course of MRI signals after a single OA challenge reached a maximum at 48 h and decreased significantly at 96 h. After the second and subsequent challenges, the maximum signal occurred at 6 h with a time-dependent decline over the remainder of the time course. A reduction of the inflammatory response following repeated administration of OA was also detected by BAL fluid analysis. The decrease in vascular permeability assessed by DCE-MRI in repeatedly OA-challenged rats was consistent with the thickening of the vascular wall for vessels of diameter up to 300 microm revealed by histology. Angiogenesis of vessels smaller than 30 microm was also detected histologically. These results suggest that MRI can be used to detect the inflammatory response and vascular remodeling associated with chronic airway inflammation in rat models involving repeated administration of allergen. As the contrast agent used in the DCE-MRI experiments is approved for clinical use, there is potential to translate the approach to patients.

Early Antiangiogenic Activity of SU11248 Evaluated In vivo by Dynamic Contrast-Enhanced Magnetic Resonance Imaging in an Experimental Model of Colon Carcinoma

To compare two dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) techniques in terms of their ability in assessing the early antiangiogenic effect of SU11248, a novel selective multitargeted tyrosine kinase inhibitor, that exhibits direct antitumor and antiangiogenic activity via inhibition of the receptor tyrosine kinases platelet-derived growth factor receptor, vascular endothelial growth factor receptor, KIT, and FLT3. A s.c. tumor model of HT29 human colon carcinoma in athymic mice was used. Two DCE-MRI techniques were used based, respectively, on macromolecular [Gd-diethylenetriaminepentaacetic acid (DTPA)-albumin] and low molecular weight (Gd-DTPA) contrast agents. The first technique provided a quantitative measurement of transendothelial permeability and fractional plasma volume, accepted surrogate markers of tumor angiogenesis. With the second technique, we quantified the initial area under the concentration-time curve, which gives information related to tumor perfusion and vascular permeability. Experiments were done before and 24 hours after a single dose administration of SU11248. The early antiangiogenic effect of SU11248 was detected by DCE-MRI with macromolecular contrast agent as a 42% decrease in vascular permeability measured in the tumor rim. The effect was also detected by DCE-MRI done with Gd-DTPA as a 31% decrease in the initial area under the concentration-time curve. Histologic slices showed a statistically significant difference in mean vessel density between the treated and control groups. The early antiangiogenic activity of SU11248 was detected in vivo by DCE-MRI techniques using either macromolecular or low molecular weight contrast agents. Because DCE-MRI techniques with low molecular weight contrast agents can be used in clinical studies, these results could be relevant for the design of clinical trials based on new paradigms.

KRN633: A selective inhibitor of vascular endothelial growth factor receptor-2 tyrosine kinase that suppresses tumor angiogenesis and growth

Vascular endothelial growth factor (VEGF) and its receptor VEGFR-2 play a central role in angiogenesis, which is necessary for solid tumors to expand and metastasize. Specific inhibitors of VEGFR-2 tyrosine kinase are therefore thought to be useful for treating cancer. We showed that the quinazoline urea derivative KRN633 inhibited tyrosine phosphorylation of VEGFR-2 (IC50 = 1.16 nmol/L) in human umbilical vein endothelial cells. Selectivity profiling with recombinant tyrosine kinases showed that KRN633 was highly selective for VEGFR-1, -2, and -3. KRN633 also blocked the activation of mitogen-activated protein kinases by VEGF, along with human umbilical vein endothelial cell proliferation and tube formation. The propagation of various cancer cell lines in vitro was not inhibited by KRN633. However, p.o. administration of KRN633 inhibited tumor growth in several in vivo tumor xenograft models with diverse tissue origins, including lung, colon, and prostate, in athymic mice and rats. KRN633 also caused the regression of some well-established tumors and those that had regrown after the cessation of treatment. In these models, the trough serum concentration of KRN633 had a more significant effect than the maximum serum concentration on antitumor activity. KRN633 was well tolerated and had no significant effects on body weight or the general health of the animals. Histologic analysis of tumor xenografts treated with KRN633 revealed a reduction in the number of endothelial cells in non-necrotic areas and a decrease in vascular permeability. These data suggest that KRN633 might be useful in the treatment of solid tumors and other diseases that depend on pathologic angiogenesis.

A phase I/II trial of single-agent PTK 787/ZK 222584 (PTK/ZK), a novel, oral angiogenesis inhibitor, in patients with recurrent glioblastoma multiforme (GBM)

1512 Background: VEGF-mediated signaling pathways may be critical to GBM neovascularization, and inhibition of VEGF pathways may be an important therapeutic target in GBM. PTK/ZK is a novel, oral angiogenesis inhibitor that potently inhibits all known vascular endothelial growth factor receptors (VEGFRs), and therefore blocks signaling by VEGF-A, -B, -C, and -D. Methods: PTK/ZK was administered once daily as oral monotherapy starting at 150 mg/day and escalating to 500, 1,000, 1,500, or 2,000 mg/day, with dose expansion at the 1,200- and 1,500-mg/day dose levels. The primary objective was to determine the safety and tolerability of continuous oral PTK/ZK in patients with recurrent GBM. A secondary endpoint was response to treatment (defined as decreases in tumor blood supply by dynamic contrast-enhanced [DCE] and dynamic susceptibility-change [DSC] MRI) and tumor response. Results: To date, 55 patients have been treated at these dose levels: 150 mg/day (n = 3); 500 mg/day (n = 6); 1,000 mg/day (n = 6); 1,200 mg/day (n = 20); 1,500 mg/day (n = 14); and 2,000 mg/day (n = 6). Median age was 54 years (range, 21–73 years), and median prior number of therapies was 4 (range, 1–12 therapies). Dose-limiting toxicities included liver enzyme elevation and deep-vein thrombosis (in 1/6 patients at 1,000 mg/day), insomnia or cerebral edema (in 2/14 patients at 1,500 mg/day), and fatigue or nausea/vomiting (in 2/6 patients at 2,000 mg/day). Among 47 evaluable patients, best responses were 2 (4%) partial responses, 31 (56%) stable disease, and 14 (25%) disease progression. Median duration of stable disease was 12.1 weeks (95% CI = 9.9, 15.1 weeks). DCE- and DSC-MRI scanning showed decreases in vascular permeability and cerebral blood volume, respectively, at days 2 and 30 of treatment; the decreases in both parameters at day 30 appeared dose dependent. Further assessment by blinded reviewers is ongoing. Conclusions: PTK/ZK monotherapy at 1,200 and 1,500 mg/day was well tolerated in patients with recurrent GBM, demonstrated changes in vascular permeability and blood volume, and stabilized disease in the majority of patients. Author Disclosure Employment or Leadership Consultant or Advisory Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Novartis Pharmaceuticals Corp.; Schering AG Imclone; Novartis; Schering-Plough Imclone; Novartis; Regeneron; Schering-Plough; Schering AG Novartis

A pilot study to evaluate response and angiogenesis after treatment with bevacizumab in patients with inflammatory breast cancer

578 Background: Inflammatory breast cancer (IBC) is an aggressive form of breast cancer associated with vascular and lymphatic invasion. Previously untreated IBC patients received bevacizumab (BV), a recombinant humanized monoclonal antibody to VEGF, to evaluate endothelial cell proliferation (CD31/Ki67), tumor cell proliferation (Ki67), tumor microvessel density (MVD), VEGF, and vascular permeability (k21) on dynamic contrast enhanced MRI (dMRI). Methods: Treatment consisted of BV alone for cycle (C) 1 (15mg/m2 on day 1) followed by six cycles of BV (15mg/m2) with doxorubicin (50mg/m2) and docetaxel (75mg/m2) q3wk and G-CSF in C2–7. Following mastectomy and radiation, BV was given for eight more cycles with hormonal therapy for ER+ pts. Tumor core biopsies and dMRI were obtained at baseline, after C1, 4, 7. Results: Sixteen pts enrolled received 125 cycles of treatment. Pts: Stage III 87.5% and IV 12.5%, ER+ 38%, Her2 positive 12.5%. Post-chemo, thirteen pts (3 too early to evaluate): 8PRs, 1PR unconfirmed, 1PD. A decrease in CD31/Ki67 was seen after BV treatment alone (-1% median change, p=0.031). There were no decreases in Ki67 or MVD after BV alone (p=0.97 and 0.70) or BV and chemotherapy (p=0.23 and 0.85). There were significant decreases in k21 after treatment (n=12), with a median change of -53% (p=0.019) after C1, -70% (p=0.001) after C4, and -56% (p=0.008) after C7. In patients with PR there were trends towards a decrease in VEGF after C1(-57% median change, p=0.09) and decrease in Ki67 after C4/C7 (-57% median change, p=0.09). There were no correlations between k21and CD31/Ki67 index, Ki67, or VEGF after BV alone (n=11); but a trend towards a correlation between VEGF and k21 after BV and chemotherapy (r=0.62, p=0.10). Most common grade 3/4 toxicities (n=16): neutropenia (63%), diarrhea (25%), HTN (25%), and febrile neutropenia (19%). Conclusions: Preliminary results revealed a decrease in vascular permeability on dMRI and endothelial cell proliferation with BV treatment alone. In responding pts, there were trends toward a decrease in VEGF after BV and tumor cell proliferation after BV and chemotherapy. No significant financial relationships to disclose.

A pilot study to evaluate response and angiogenesis after treatment with bevacizumab in patients with inflammatory breast cancer

578 Background: Inflammatory breast cancer (IBC) is an aggressive form of breast cancer associated with vascular and lymphatic invasion. Previously untreated IBC patients received bevacizumab (BV), a recombinant humanized monoclonal antibody to VEGF, to evaluate endothelial cell proliferation (CD31/Ki67), tumor cell proliferation (Ki67), tumor microvessel density (MVD), VEGF, and vascular permeability (k21) on dynamic contrast enhanced MRI (dMRI). Methods: Treatment consisted of BV alone for cycle (C) 1 (15mg/m2 on day 1) followed by six cycles of BV (15mg/m2) with doxorubicin (50mg/m2) and docetaxel (75mg/m2) q3wk and G-CSF in C2–7. Following mastectomy and radiation, BV was given for eight more cycles with hormonal therapy for ER+ pts. Tumor core biopsies and dMRI were obtained at baseline, after C1, 4, 7. Results: Sixteen pts enrolled received 125 cycles of treatment. Pts: Stage III 87.5% and IV 12.5%, ER+ 38%, Her2 positive 12.5%. Post-chemo, thirteen pts (3 too early to evaluate): 8PRs, 1PR unconfirmed, 1PD. A decrease in CD31/Ki67 was seen after BV treatment alone (-1% median change, p=0.031). There were no decreases in Ki67 or MVD after BV alone (p=0.97 and 0.70) or BV and chemotherapy (p=0.23 and 0.85). There were significant decreases in k21 after treatment (n=12), with a median change of -53% (p=0.019) after C1, -70% (p=0.001) after C4, and -56% (p=0.008) after C7. In patients with PR there were trends towards a decrease in VEGF after C1(-57% median change, p=0.09) and decrease in Ki67 after C4/C7 (-57% median change, p=0.09). There were no correlations between k21and CD31/Ki67 index, Ki67, or VEGF after BV alone (n=11); but a trend towards a correlation between VEGF and k21 after BV and chemotherapy (r=0.62, p=0.10). Most common grade 3/4 toxicities (n=16): neutropenia (63%), diarrhea (25%), HTN (25%), and febrile neutropenia (19%). Conclusions: Preliminary results revealed a decrease in vascular permeability on dMRI and endothelial cell proliferation with BV treatment alone. In responding pts, there were trends toward a decrease in VEGF after BV and tumor cell proliferation after BV and chemotherapy. No significant financial relationships to disclose.

Tyroxine hydroxylase (TH) downregulation in hyperstimulated ovaries reveals the dopamine agonist bromocriptine (Br2) as an effective and specific method to block increased vascular permeability (VP) in OHSS

Objective: Ovarian hyperstimulation syndrome (OHSS) is induced by the ovarian release of vasoactive-angiogenic substances which increase vascular permeability (VP). Vascular Endothelial Growth Factor (VEGF) plays a major role in this regard. Inhibition of VEGF action has been accompanied by decrease in VP. However, the use of such strategy as antiangiogenic tumoral drugs in women has been abandoned due to toxic effects. Aiming to find a specific-non toxic treatment for OHSS, we have looked at the expression of gene products regulated under OHSS conditions, and developed new approaches employing the D-2 dopamine agonist Bromocriptine (Br2). Design: In the first series of experiments, we compared ovarian gene expression in animals (n = 8 per group) subjected to 3 different regimens of gonadotropins: Pregnant Mare′s Serum Gonadotropin (PMSG) 10 IU/day for 4 days and hCG (30 IU) the fifth day (OHSS group); PMSG (10 IU) plus hCG (10 IU) 2 days later (MILD group); and PMSG (1 IU) plus hCG (1 IU) 48 hrs later (CONTROL group). Fourty-eight hours after hCG, VP was measured, and mRNA from ovaries extracted to perform gene expression profiles in macroarray filters containing 14,000 genes. Only genes with a 3-fold up or down regulation were considered to be significantly regulated. The hybridization results were confirmed by quantitative Fluorescence(QF)-PCR, immunostaining, and cluster analysis. In the second series of experiments, Br2 was employed in OHSS animals (40 IU PMSG+30IU hCG). Different doses of Br2: 1.6, 4, 10 and 25 mg/kg were intraperitoneally administered the day of hCG and 24 hrs later. VP was measured 48 hrs after hCG. Materials and Methods: Immature female Wistar rats were employed. In the macroarray technology, RNA was reverse transcribed and transferred to the nylon membrane filters under radioactive conditions. Clusters of 5×5 spot signals were analyzed using the AIDA software, and its expression paired to the other groups after background signal substraction. To confirm gene expression, we used the same RNA and performed QF-PCR in an ABIPRISM 7700 thermocycler to amplify 3 up-regulated and 3 down-regulated genes. To measure VP, 0.2 ml 5mM Evans blue(EB) were injected via the femoral vein. After 30 min, the peritoneal cavity was irrigated with saline. Light absorption was measured in the recovered fluid at 595 nm and the VP expressed as μg EB/100 g body weight. Results: Gene expression in the 3 groups showed 80 upregulated and 7 downregulated genes in OHSS as compared to MILD and CONTROLS. Upregulated genes were grouped in 5 families: Cholesterol synthesis,VEGF signal transduction, Prostaglandins synthesis, oxidative stress process and Prolactin synthesis. The down-regulation of Tyrosine-hydroxylase (TH, enzyme responsible for Dopamine synthesis) was considered a characteristic of OHSS too. As a result, the D2 dopamine agonist Br2 was employed. Increased VP in OHSS animals (30.7±5.8 μg EB/100 g body weight) was significantly reduced when 1.6 mg Br2 (15.3±3.3; p = 0.04), 4 mg Br2 (14.2±2.2; p = 0.04), 10 mg Br2 (10.1±2.6; p = 0.009), and 25 mg Br2 (8.0±1.9; p = 0.001) were employed. Conclusion: OHSS is associated with the up and down regulation of several families of genes in the ovaries. In addition to VEGF, the up and down regulation of genes encoding for prolactin and dopamine expression, encouraged the successful use of Br2 to prevent increased VP. These experiments provide the rationale to test Br2 as a safe and possible effective medication in the prevention/treatment of OHSS.

Lessons from CADASIL.

Vascular dementia (VaD) includes several different vascular mechanisms and changes in the brain. Among VaD, CADASIL is an inherited angiopathy caused by mutations in the Notch3 gene. The pathological hallmark of CADASIL is a granular osmiophilic material deposit (GOM) that is not only found in the brain, but also in the peripheral vascular tree. Consequently, a window into the brain was opened from a strictly neurological disease with tremendous consequences thanks to a skin biopsy. The latter was and continues to be used as a diagnostic tool for CADASIL, despite an immunohistochemical test that is now available. The skin biopsy first used as a diagnostic tool revealed the existence of numerous other VaDs presenting systemic vascular changes. Later, skin biopsy became a research tool, and a morphological skin vessel change classification was proposed on 300 patients. Interestingly, similar skin vessel lesions appear to be related to the same biological modifications. In addition, an early destruction of the medial muscle cells was noticed in 74% of cases. Because vascular smooth muscle cells secrete a powerful endothelial permeability factor (VEGF), their destruction could lead to a decrease in vascular permeability. Cocultures of endothelial cells with vascular muscle cells showed that their presence doubled vascular permeability. Thus, alteration or the loss of vascular muscle cells likely results in hypopermeability, in addition to vessel wall hypotonia and a watershed hypoperfusion. The wealth of information brought forth by knowledge of CADASIL provided new tools for research and clues for understanding the consequences of vascular impairments in dementia.

Clinical use of antenatal corticosteroids: benefits and risks.

After completing this article, readers should be able to: 1. Describe the rationale for the use of antenatal corticosteroids for human fetal maturation. 2. Delineate recommendations of the 1994 National Institutes of Health Consensus Conference on Antenatal Corticosteroids. 3. Review areas of controversy in the clinical use of antenatal corticosteroids. 4. Describe the potential risks of multiple courses of antenatal corticosteroids. 5. Delineate current recommendations for clinical use of antenatal corticosteroids to promote fetal maturation. Glucocorticoid administration initially was appreciated as a method for accelerating development of the intestine more than 40 years ago. Since that time, developmental effects of these agents have been defined in at least 16 tissues of mammals. More than 30 years ago, while studying the mechanism of initiation of labor in sheep, Liggins made the observation that infusion of corticosteroids into fetal lambs was associated with improved survival and decreased lung disease in animals delivered preterm. Subsequently, numerous studies in cell culture, animal models, and human fetal tissue have delineated the multiple effects of glucocorticoids on the developing lung. These include increases in both the total tissue and alveolar surfactant pools; a decrease in vascular permeability, with less protein leak into the alveolar spaces; enhanced clearance of lung water; maturation of parenchymal structure; increase in lung compliance and maximal lung volume; enhanced response to surfactant treatment; and improvement in respiratory function, outcome, and survival. (See accompanying article on scientific rationale for use of antenatal glucocorticoids.) In 1972, the first randomized clinical trial of administration of antenatal corticosteroids (ANCS) in humans demonstrated decreased mortality and a decreased incidence of respiratory distress syndrome (RDS) in preterm infants born at less than 34 weeks’ gestation, who were treated with betamethasone for at least 24 hours before delivery. Over the ensuing 20 years, multiple clinical investigations continued to document the effectiveness of ANCS on …

Lung oedema induced by Tityus serrulatus scorpion venom in the rat

The effects of drugs were investigated on the induction of acute lung oedema by scorpion Tityus serrulatus venom in male Wistar rats (200—230 g) anaesthetized with sodium pentobarbital (40 mg/kg, i.p.). Intravenous (i.v.) injection of scorpion venom (0.5 mg/kg) into 12 rats induced arterial hypertension and severe lung oedema, whereas i.v. injection of scorpion venom into 16 rats previously injected with commercial heparin induced arterial hypertension, but only a slight lung oedema. It is suggested that the inhibitory effect of commercial heparin on the genesis of lung oedema may be due to a decrease in vascular permeability in the lungs. Previous i.v. injection of aprotinin did not prevent the arterial hypertension and the lung oedema induced by scorpion venom. Previous injections of platelet-activating factor antagonists (BN-52021 and WEB-2170) or of an inhibitor of lipo- and cyclooxygenase (Nordihydroguaiaretic acid) did not prevent the arterial hypertension induced by scorpion venom, but decreased the magnitude of the lung oedema elicited by the venom. Previous injections of inhibitors of 5-lipoxygenase (MK-886) or cyclooxygenase (aspirin or indomethacin) significantly decreased the magnitude of the lung oedema induced by scorpion venom. It is concluded that the release of vascular permeability factors, such as platelet-activating factor, leukotrienes, and prostaglandins may play a role in the induction of acute lung oedema by scorpion venom in rats.

Microvascular responses in the skeletal muscle of the diabetic rat

We examined microvascular responses to acetylcholine (ACh), adenosine (ADO), nitroprusside (SNP), bradykinin (BK), histamine (HIS), and serotonin (5-HT) in control and diabetic rats. Agonists were applied topically to neurovascularly intact and environmentally controlled cremaster muscles of diabetic or control rats 3 weeks after streptozotocin or vehicle injection. Precapillary arteriolar diameters and leakage of fluorescein isothiocyanate conjugated to bovine serum albumin (FITC-BSA) in postcapillary venules were measured by using intravital microscopy techniques. All agents dilated the arterioles, and BK, HIS, and 5-HT caused concentration-dependent leakage of FITC-BSA. Of the vasodilators tested, only ACh-induced dilation was attenuated in diabetic animals. BK and HIS caused a concentration-dependent leakage of FITC-BSA that was similar in diabetic and control groups, but FITC-BSA leakage to 5-HT was significantly attenuated in diabetic animals. To determine whether the attenuated responses were attributable to an alteration of nitric oxide, the same dilational and leakage-promoting agents were tested in normoglycemic animals in the presence of N'-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide synthase. Only ACh-induced dilation in control animals was attenuated in the presence of L-NAME; all other responses were normal. Thus, although the attenuated ACh-induced arteriolar dilation in striated muscle of diabetic rats may be linked to an impairment of nitric oxide function or release, the reduced leakage to serotonin is not linked to nitric oxide. The impaired leakage response to 5-HT is specific for that agonist and is not an indication of a generalized decrease in vascular permeability in these diabetic animals.

Preventive effect of atropine on indomethacin-induced gastrointestinal mucosal and vascular damage in rats.

The protective effect of 'chemical vagotomy' (atropine given in three doses: 0.1, 0.5 and 1.0 mg/kg i.p.) was examined on indomethacin (IND, 20 mg/kg s.c.)-induced macroscopic gastrointestinal (GI) mucosal erosions and changes of vascular permeability in the stomach and three equal parts of small intestine and colon in rats. The different doses of atropine were administered at 0, 5, 10, 15 and 20 h after IND administration, then the number and severity of lesions were noted and the vascular damage was measured by Evans blue extravasation into the mucosa and intraluminal juice at 24 h after the IND treatment. Our results indicate that atropine ('chemical vagotomy') dose-dependently and significantly decreases the IND-induced mucosal erosions and vascular permeability in the vagal nerve-innervated parts of GI tract (i.e. the stomach, small intestine and proximal colon). Atropine in 0.5 and 1.0 mg/kg doses has a significantly higher protective effect on the vascular damage than on the macroscopic mucosal lesions in the stomach, small intestine and proximal colon. The vascular permeability is only one of those factors which have a role in the appearance of the GI mucosal erosions after IND treatment. These results suggest that the decrease of vascular permeability is involved in the protective effect of atropine against IND-induced GI mucosal damage.

Quantitative and qualitative effects of experimental radioimmunotherapy on tumor vascular permeability

Localization of radiolabeled antibodies in the perivascular space of tumors resulted in morphological changes in blood vessel structure and physiological changes in tumor vessel function. Vessel diameter decreased by day 14 and was associated with a significant decline in vascular volume (VV). Upon recovery of VV, the basement membrane surrounding the endothelium had thickened. Tumor vascular permeability (VP) decreased within 7 days of treatment and remained suppressed throughout the 42-day observation period of our study. The decline in VP, which could be visualized by fluorescent microscopy of FITC-dextran extravasation, was dose-related and could be quantified at doses as low as 800 cGy. The radioantibody-induced 50-80% decline in tumor VP was observed in 3 human colonic xenografts (GW-39, LSI74T and MOSER). Decreases in VP have been observed for proteins ranging in size from 20 to 150 kDa. Similar effects on VP were noted when low protein doses (10-30 micrograms), which resulted in heterogeneous antibody distribution, were used or with high protein doses (400-750 micrograms), which resulted in more uniform penetration of antibody. If 188Re or 90Y, radiometals with higher beta-energy and longer path lengths, were substituted for 131I, a similar decrease in VP was observed. The radioimmunotherapy (RAIT)-induced decrease in tumor VP resulted in a 90% decline in accretion of a second dose of radioantibody. The 10% of the second dose that was taken up by the tumor targeted many already non-viable tumor regions and some, but not all, viable tumor cell clusters.

Pharmacological characterization of polycation‐induced rat hind‐paw oedema

1. The inflammatory response induced by poly-L-arginine in the rat hind-paw was studied both by measuring paw oedema and histologically. 2. The paw volume was measured with a hydroplethysmometer at 0.5, 1, 2, 4, 6 and 18 h after the subplantar injection of the polycation. Protein extravasation was evaluated with Evans' blue and the histology studied by light microscopy. 3. Poly-L-arginine (12, 24, 43 and 115kD) caused dose- and molecular weight-dependent oedema which had a rapid onset and long duration. Evans' blue extravasation paralleled the oedema induced by poly-L-arginine. Microscopic examination of the paws at early stages of oedema formation showed exuberant liquid exudate with no inflammatory cells. After 18 h, a cellular infiltrate was present, consisting mainly of mononuclear cells. 4. Indomethacin, dexamethasone, BW755c or the PAF-antagonist WEB 2086 caused no significant inhibition of the poly-L-arginine-induced oedema. Cyproheptadine had inhibitory effects only on the early stages of the polycation-induced oedema. Similar results were observed with rats depleted of histamine and 5-hydroxytryptamine. 5. Heparin, a polyanion, injected in the rat paw caused a marked inhibition of the polycation-induced oedema. NG-monomethyl-L-arginine (LNMMA), an inhibitor of EDRF synthesis, injected locally also produced a marked inhibition, but this inhibition was reversed by iloprost. 6. These results suggest that the oedema induced by polycations was due to their cationic charge. The inhibitory effect of LNMMA is probably due to a decrease in vascular flow rather than a decrease in vascular permeability.