Atsushi Imaizumi

Inhibitory effect of statins on renal epithelial-to-mesenchymal transition.

Background/Aim: Recent studies have suggested that statins may play a role in the protection against renal failure which is independent of cholesterol reduction. Activation of RhoGTPases is a key step in renal tubular cells’ epithelial-to-mesenchymal transition (EMT) which contributes to renal interstitial fibrosis. We hypothesized that statins could act by inhibiting the synthesis of the isoprenoids, such as geranylgeranyl pyrophosphate, which is essential for membrane attachment and biological activity of RhoGTPases, RhoA and Rac1. Methods: Human proximal tubular epithelial cells (HK2) were used to examine the inhibitory effect of statins on EMT induced with medium conditioned by activated peripheral blood mononuclear cells. Results: Our study demonstrates that the statins lovastatin, simvastatin, and pravastatin inhibit HK2 cells to undergo EMT. Inhibition of EMT in HK2 cells with these statins resulted in a reduction of RhoA and Rac1 activation in both the cytoplasmic and membrane-bound forms, in preservation of the expression of the epithelial cell markers E-cadherin and cytokeratin-19, and in a decrease in Fn-EDA expression, a marker for the myofibroblast phenotype. The decreased levels of activated RhoA and Rac1 in both the cytoplasmic and membrane fractions of the cells were reversed by geranylgeranyl pyrophosphate and mevalonate, and thus attributable to the inhibition of isoprenylation of RhoGTPases by statins. Conclusion: This phenomenon could explain the beneficial effect of statins on EMT and on renal fibrosis prevention.

Rapid, sensitive and specific diagnosis of Bordetella pertussis using the polymerase chain reaction

Use of a repetitive DNA sequence of Bordetella pertussis allowed successful detection of the organism by the polymerase chain reaction (PCR). The method was highly sensitive, being able to detect B. pertussis in specimens containing only a few cells. It was also highly specific, with no amplification of specimens containing other organisms, for example Haemophilus influenzae or Neisseria, being observed. A diagnosis could be made within 1 day. The PCR assay was also evaluated in clinical specimens. Among 47 nasopharyngeal specimens obtained from 24 patients with laboratory‐confirmed pertussis, 27 were positive by PCR and 19 by culture. In particular, all three bronchial aspirates from one patient with pertussis were positive by PCR, but only one showed positive on culture. Eleven specimens from parapertussis patients and 65 specimens from patients without pertussis tested negative. It was concluded that this newly developed PCR method for the diagnosis of pertussis was more rapid and sensitive than the usual culture method. Polymerase chain reaction could have a major impact on the treatment and control of this infection and would be a useful tool for studying the pathogenesis of B. pertussis infection.

Pertussis toxin induces bronchopulmonary hyperresponsiveness in guinea-pigs while antagonizing the effects of formyl-L-methionyl-L-leucyl-L-phenylalanine

Pertussis toxin injected i.v. at 0.8-20 micrograms/kg markedly enhanced bronchoconstriction induced by the i.v. administration of histamine or serotonin (5-HT) (0.5-16 micrograms/kg) to propranolol-treated guinea-pigs, under conditions where propranolol or pertussis toxin alone were poorly effective. In contrast, bronchoconstriction and the accompanying leukopenia induced by the i.v. administration of the secretagogue formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP) were suppressed by pertussis toxin. Bronchoconstriction induced by histamine or 5-HT was not enhanced when perfused lungs from pertussis toxin-treated guinea-pigs were studied in vitro, under conditions where bronchoconstriction and thromboxane A2 release evoked by fMLP were suppressed. Pertussis toxin negatively modifies signal transduction in cells involved in the lung responses to fMLP both in vivo and in vitro, but positively and only in vivo it modifies signal transduction in cells involved in the lung responses to the direct constricting agents histamine and 5-HT. As hyperresponsiveness to histamine and 5-HT were exclusively found in vivo, the target for pertussis toxin is probably not the adrenergic nor the cholinergic systems, since neither hexamethonium, isoprenaline, atropine nor vagotomy were effective. In addition, since dexamethasone and nedocromil sodium were inactive and enrichment of bronchoalveolar lavage with inflammatory cells was not noted, despite lung invasion by neutrophils and lymphocytes, acute inflammation does not account for pertussis toxin-induced hyperresponsiveness.

Guinea-pig treatment with pertussis toxin suppresses macrophage-dependent bronchoconstriction by fMLP and fails to inhibit the effects of PAF

1 Bronchoconstriction and thromboxane B2 (TxB2) release following the intra‐tracheal administration of the secretagogue N‐formyl‐l‐methionyl‐l‐leucyl‐l‐phenylalanine (fMLP) to lungs from pertussis toxin‐treated guinea‐pigs in vivo and in vitro were inhibited as compared to saline‐treated animals, under conditions where the responses to PAF were modified less effectively. 2 The cell target accounting for bronchoconstriction by fMLP and for inhibition by pertussis toxin is located in the airways and is probably the alveolar macrophage. Indeed (a) fMLP‐induced superoxide anions and TxB2 formation by alveolar macrophages were inhibited by pertussis toxin given in vivo; (b) Gi proteins of membranes from alveolar macrophages were ADP‐ribosylated in vivo by pertussis toxin and (c) bronchoconstriction and TxB2 release in response to the intra‐tracheal administration of fMLP to lungs from pertussis toxin‐treated animals were restored when alveolar macrophages from control guinea‐pigs were transferred into the airways of pertussis toxin‐treated animals before lung isolation. 3 Pertussis toxin administered to guinea‐pigs in vivo, reduced the subsequent TxB2 formation and superoxide anion release by alveolar macrophages stimulated with PAF, but failed to inhibit PAF‐induced bronchoconstriction. 4 Formation of TxB2 by alveolar macrophages following the intra‐tracheal administration of fMLP accounts for bronchoconstriction and requires pertussis toxin‐sensitive Gi proteins. PAF operates via a different mechanism, which is independent of Gi‐like protein and involves mediators other than TxB2 and superoxide anions.