Izumi Hayashi

Host stromal bradykinin B2 receptor signaling facilitates tumor-associated angiogenesis and tumor growth.

We evaluated the significance of the host kallikrein-kinin system in tumor angiogenesis and tumor growth using two rodent models genetically deficient in a kallikrein-kinin system. Inoculation of Walker 256 carcinoma cells into the s.c. tissues of the back of normal Brown Norway Kitasato rats (BN-Ki rats) resulted in the rapid development of solid tumors with marked angiogenesis. By contrast, in kininogen-deficient Brown Norway Katholiek rats (BN-Ka rats), which cannot generate intrinsic bradykinin (BK), the weights of the tumors and the extent of angiogenesis were significantly less than those in BN-Ki rats. Daily administration of B2 receptor antagonists significantly reduced angiogenesis and tumor weights in BN-Ki rats to levels similar to those in BN-Ka rats but did not do so in BN-Ka rats. Angiogenesis and tumor growth were significantly suppressed in B2 receptor knockout mice bearing sarcoma 180 compared with their wild-type counterparts. Immunoreactive vascular endothelial growth factor (VEGF) was localized in Walker tumor stroma more extensively in BN-Ki rats than in BN-Ka rats, although immunoreactive B2 receptor also was detected in the stroma to the same extent in both types of rats. Cultured stromal fibroblasts isolated from BN-Ki rats and BN-Ka rats produced VEGF in response to BK (10−8-10−6 m), and this stimulatory effect of BK was abolished with a B2 receptor antagonist, Hoe140 (10−5 m). These results suggest that BK generated from kininogens supplied from the host may facilitate tumor-associated angiogenesis and tumor growth by stimulating stromal B2 signaling to up-regulate VEGF production mainly in fibroblasts.

Role of thromboxane derived from COX‐1 and ‐2 in hepatic microcirculatory dysfunction during endotoxemia in mice

Although thromboxanes (TXs), whose synthesis is regulated by cyclooxygenase (COX), have been suggested to promote inflammation in the liver, little is known about the role of TXA2 in leukocyte endothelial interaction during endotoxemia. The present study was conducted to investigate the role of TXA2 as well as that of COX in lipopolysaccharide (LPS)‐induced hepatic microcirculatory dysfunction in male C57Bl/6 mice. We observed during in vivo fluorescence microscopic study that LPS caused significant accumulation of leukocytes adhering to the hepatic microvessels and non‐perfused sinusoids. Levels of serum alanine transaminase (ALT) and tumor necrosis factor alpha (TNFα) also increased. LPS raised the TXB2 level in the perfusate from isolated perfused liver. A TXA2 synthase inhibitor, OKY‐046, and a TXA2 receptor antagonist, S‐1452, reduced LPS‐induced hepatic microcirculatory dysfunction by inhibiting TNFα production. OKY‐046 suppressed the expression of an intercellular adhesion molecule (ICAM)‐1 in an LPS‐treated liver. In thromboxane prostanoid receptor‐knockout mice, hepatic responses to LPS were minimized in comparison with those in their wild‐type counterparts. In addition, a selective COX‐1 inhibitor, SC‐560, a selective COX‐2 inhibitor, NS‐398, and indomethacin significantly attenuated hepatic responses to LPS including microcirculatory dysfunction and release of ALT and TNFα. The effects of the COX inhibitors on hepatic responses to LPS exhibited results similar to those obtained with TXA2 synthase inhibitor, and TXA2 receptor antagonist. In conclusion, these results suggest that TXA2 is involved in LPS‐induced hepatic microcirculatory dysfunction partly through the release of TNFα, and that TXA2 derived from COX‐1 and COX‐2 could be responsible for the microcirculatory dysfunction during endotoxemia. (HEPATOLOGY 2004;39:139–150.)

Roles of Prostaglandin E2–EP3/EP4 Receptor Signaling in the Enhancement of Lymphangiogenesis During Fibroblast Growth Factor-2–Induced Granulation Formation

Objective—One of the hallmarks of inflammation is lymphangiogesis that drains the interstitial fluids. During chronic inflammation, angiogenesis is induced by a variety of inflammatory mediators, such as prostaglandins (PGs). However, it remains unknown whether they enhance lymphangiogenesis. We examined the roles of cyclooxygenase-2 (COX-2) and PGE2 receptor signaling in enhancement of lymphangiogenesis during proliferative inflammation. Methods and Results—Lymphangiogenesis estimated by podoplanin/vascular endothelial growth factor (VEGF) receptor-3/LYVE-1 expression was upregulated during proliferative inflammation seen around and into subcutaneous Matrigel plugs containing fibroblast growth factor-2 (125 ng/site). A COX-2 inhibitor (celecoxib) significantly reduced lymphangiogenesis in a dose-dependent manner, whereas topical PGE2 enhanced lymphangiogenesis. Topical injection of fluorescein isothiocyanate–dextran into the Matrigel revealed that lymphatic flow from the Matrigels was COX-2 dependent. Lymphangiogenesis was suppressed in the granulation tissues of mice lacking either EP3 or EP4, suggesting that these molecules are receptors in response to endogenous PGE2. An EP3-selective agonist (ONO-AE-248) increased the expression of VEGF-C and VEGF-D in cultured macrophages, whereas an EP4-selective agonist (ONO-AE1-329) increased VEGF-C expression in cultured macrophages and increased VEGF-D expression in cultured fibroblasts. Conclusion—Our findings suggest that COX-2 and EP3/EP4 signaling contributes to lymphangiogenesis in proliferative inflammation, possibly via induction of VEGF-C and VEGF-D, and may become a therapeutic target for controlling lymphangiogenesis.

Calcitonin gene‐related peptide released by capsaicin suppresses myoelectrical activity of gastric smooth muscle

Background:u2002 It is widely accepted that the inhibition of gastric motor activity as well as the maintenance of gastric mucosal blood flow and mucous secretion are important for the homeostasis of the gastric mucosa. The present study was performed to ascertain whether or not capsaicin, which can protect the stomach from noxious stimuli, affects gastric motor activity.

Roles of prostaglandin E2-EP1 receptor signaling in regulation of gastric motor activity and emptying

It is widely accepted that the inhibition of gastric motor activity as well as the maintenance of gastric mucosal blood flow and mucous secretion are important for the homeostasis of the gastric mucosa. The present study was performed to ascertain whether or not endogenous PGs, which can protect the stomach from noxious stimuli, affect gastric motor activity and emptying. The myoelectrical activity of rat gastric smooth muscle was increased at intragastric pressures of over 2 cmH(2)O. Replacement of intragastric physiological saline with 1 M NaCl solution significantly increased PGI(2) and PGE(2) in stomach and suppressed the myoelectrical activity under a pressure of 2 cmH(2)O by 70%. Indomethacin inhibited the suppression of myoelectrical activity by 1 M NaCl. The myoelectrical activity under a pressure of 2 cmH(2)O was suppressed by continuous infusion of a selective EP1 agonist (ONO-DI-004, 3-100 nmol·kg(-1)·min(-1)) into the gastric artery in a dose-dependent manner, but not by that of the PGI receptor agonist beraprost sodium (100 nmol·kg(-1)·min(-1)). Suppression of myoelectrical activity with 1 M NaCl was inhibited by continuous infusion of a selective EP1 antagonist (ONO-8711, 100 nmol·kg(-1)·min(-1)) into the gastric artery. Furthermore, gastric emptying was tested in EP1 knockout mice and their wild-type counterparts. Gastric emptying was strongly suppressed with intragastric 1 M NaCl in wild-type mice, but this 1 M NaCl-induced suppression was not seen in EP1 knockout mice. These results suggest that PGE(2)-EP1 signaling has crucial roles in suppression of myoelectrical activity of gastric smooth muscles and inhibition of gastric emptying and that EP1 is an obvious target for drugs that control gastric emptying.

Nitric oxide generated by iNOS reduces deformability of Lewis lung carcinoma cells

Previous studies have indicated that NO plays a crucial role in the metastasis of tumor cells and that tumor cells produce nitric oxide (NO) via inducible nitric oxide synthase (iNOS). Since the deformability of tumor cells is an important factor governing their metastatic potential, in this study we investigated the regulation of tumor cell deformability by NO. Lewis lung tumor cells (3LL cells) were also incubated with a cytokine mixture (IL‐1β, IFNγ, and TNFα). The nitrite/nitrate content of the supernatant was then measured by the Griess method, and iNOS expression was evaluated by RT‐PCR in vitro. Nitrite/nitrate was produced in response to administration of the cytokine mixture, and iNOS mRNA was expressed in the cytokine‐treated cells. The deformability of the 3LL cells was evaluated by measuring the peak pressure generated during their passage through a microfilter at a constant flow rate. Both the cytokine mixture and NO donor (NOC 18) significantly increased the filtration pressure, and the staining of the cells with rhodamine‐phalloidin revealed assembly of F‐actin in the cell membrane. In conclusion, NO plays a role in the decreased deformability of tumor cells, suggesting that NO is one of the factors that regulates metastasis.

Effects of inhaled monoethanolamine on bronchoconstriction

We previously reported a 65‐year‐old man who aspirated an alkaline detergent containing 3.3% w/v (weight of solute per volume of solution) monoethanolamine (MEA) into his lungs, causing asthma‐like symptoms. We presently describe the mechanism of MEA‐induced bronchoconstriction according to findings in guinea pigs. In anesthetized, artificially ventilated animals, changes in airway opening pressure (Pao) were measured as an index of bronchoconstriction. An aerosol of 3.3% MEA solution (0.1 ml kg−1) inhaled through a tracheal cannula induced significantly stronger bronchoconstriction than an aerosol of potassium hydroxide (KOH) solution (0.1 ml kg−1) at the same pH. MEA‐induced bronchoconstriction was significantly suppressed by premedication with intravenously injected atropine sulfate (3 mg kg−1), a muscarinic receptor antagonist, or diphenhydramine hydrochloride (10 mg kg−1), a histamine‐H1 receptor antagonist. MEA‐induced bronchoconstriction was not enhanced by premedication with an intravenous injection of neostigmine (0.1 mg kg−1), an acetylcholinesterase inhibitor. When bronchoconstriction was induced by MEA, histamine concentrations in bronchoalveolar lavage fluid (BALF) were not significantly greater than in BALF after KOH‐induced bronchoconstriction or in BALF after inhalation of physiologic saline. In vitro, contraction of trachea denuded of epithelium during superfusion with MEA (10 mm) was suppressed by premedication with pyrilamine maleate, a histamine‐H1 receptor antagonist, at 10 and 100 µm. Contraction of trachea denuded of epithelium during superfusion with MEA (10 mm) was suppressed by premedication with atropine sulfate at 10 and 100 µm. These results suggest that asthma‐like symptoms may result partly from agonistic MEA effects at histamine‐H1 receptors and muscarinic receptors. Copyright