Antibody to Endogenous Cardiotonic Steroid Reverses Vascular Fibrosis and Restores Vasorelaxation in Chronic Kidney Disease.

Marinobufagenin (MBG) is implicated in chronic kidney disease, where it removes Fli1-induced inhibition of the collagen-1. We hypothesized that (i) in nephrectomized rats, aortic fibrosis develops due to elevated plasma MBG and inhibited Fli1, and (ii) that the antibody to MBG reduces collagen-1 and improves vasodilatation. A partial nephrectomy was performed in male Sprague-Dawley rats. Sham-operated animals comprised the control group. At 5 weeks following nephrectomy, rats were administered the vehicle (n = 8), or the anti-MBG antibody (n = 8). Isolated aortic rings were tested for their responsiveness to sodium nitroprusside following endothelin-1-induced constriction. In nephrectomized rats, there was an increase in the intensity of collagen staining in the aortic wall vs. the controls. In antibody-treated rats, the structure of bundles of collagen fibers had ordered organization. Western blots of the aorta had lower levels of Fli1 (arbitrary units, 1 ± 0.05 vs. 0.2 ± 0.01; p < 0.001) and greater collagen-1 (arbitrary units, 1 ± 0.01 vs. 9 ± 0.4; p < 0.001) vs. the control group. Administration of the MBG antibody to rats reversed the effect of the nephrectomy on Fli1 and collagen-1 proteins. Aortic rings pretreated with endothelin-1 exhibited 50% relaxation following the addition of sodium nitroprusside (EC50 = 0.28 µmol/L). The responsiveness of the aortic rings obtained from nephrectomized rats was markedly reduced (EC50 = 3.5 mol/L) compared to the control rings. Treatment of rats with the antibody restored vasorelaxation. Thus, the anti-MBG antibody counteracts the Fli1-collagen-1 system and reduces aortic fibrosis.

Fli1 and Tissue Fibrosis in Various Diseases.

Being initially described as a factor of virally-induced leukemias, Fli1 (Friend leukemia integration 1) has attracted considerable interest lately due to its role in both healthy physiology and a variety of pathological conditions. Over the past few years, Fli1 has been found to be one of the crucial regulators of normal hematopoiesis, vasculogenesis, and immune response. However, abnormal expression of Fli1 due to genetic predisposition, epigenetic reprogramming (modifications), or environmental factors is associated with a few diseases of different etiology. Fli1 hyperexpression leads to malignant transformation of cells and progression of cancers such as Ewing's sarcoma. Deficiency in Fli1 is implicated in the development of systemic sclerosis and hypertensive disorders, which are often accompanied by pronounced fibrosis in different organs. This review summarizes the initial findings and the most recent advances in defining the role of Fli1 in diseases of different origin with emphasis on its pro-fibrotic potential.

Type 1 melanocortin receptors in pro-opiomelanocortin-, vasopressin-, and oxytocin-immunopositive neurons in different areas of mouse brain.

Information on the localization of the Type 1 melanocortin receptors (MC1Rs) in different regions of the brain is very scarce. As a result, the role of MC1Rs in the functioning of brain neurons and in the central regulation of physiological functions has not been studied. This work aimed to study the expression and distribution of MС1Rs in different brain areas of female C57Bl/6J mice. Using real-time polymerase chain reaction, we demonstrated the Mс1R gene expression in the cerebral cortex, midbrain, hypothalamus, medulla oblongata, and hippocampus. Using an immunohistochemical approach, we showed the MС1R localization in neurons of the hypothalamic arcuate, paraventricular and supraoptic nuclei, nucleus tractus solitarius (NTS), dorsal hippocampus, substantia nigra, and cerebral cortex. Using double immunolabeling, the MC1Rs were visualized on the surface and in the bodies and outgrowths of pro-opiomelanocortin (POMC)-immunopositive neurons in the hypothalamic arcuate nucleus, NTS, hippocampal CA3 and CA1 regions, and cerebral cortex. Co-localization with POMC indicates that MC1R, like MC3R, is able to function as an autoreceptor. In the paraventricular and supraoptic nuclei, MC1Rs were visualized on the surface and in the cell bodies of vasopressin- and oxytocin-immunopositive neurons, indicating a relationship between hypothalamic MC1R signaling and vasopressin and oxytocin production. The data obtained indicate a wide distribution of MC1Rs in different areas of the mouse brain and their localization in POMC-, vasopressin- and oxytocin-immunopositive neurons, which may indicate the participation of MC1Rs in the control of many physiological processes in the central nervous system.

Canrenone Restores Vasorelaxation Impaired by Marinobufagenin in Human Preeclampsia.

Previous studies implicated cardiotonic steroids, including Na/K-ATPase inhibitor marinobufagenin (MBG), in the pathogenesis of preeclampsia (PE). Recently, we demonstrated that (i) MBG induces fibrosis in rat tissues via a mechanism involving Fli1, a negative regulator of collagen-1 synthesis, and (ii) MBG sensitive Na/K-ATPase inhibition is reversed by mineralocorticoid antagonists. We hypothesized that in human PE elevated MBG level is associated with the development of fibrosis of the umbilical arteries and that this fibrosis can be attenuated by canrenone. Fifteen patients with PE (mean BP = 118 ± 4 mmHg; 34 ± 2 years; 38 ± 0.3 weeks gest. age) and twelve gestational age-matched normal pregnant subjects (mean BP = 92 ± 2 mmHg; 34 ± 1 years; 39 ± 0.2 weeks gest. age) were enrolled in the study. PE was associated with a higher plasma MBG level, with a four-fold decrease in Fli1 level and a three-fold increase in collagen-1 level in the PE umbilical arteries vs. those from the normal subjects (p < 0.01). Isolated rings of umbilical arteries from the subjects with PE exhibited impaired responses to the relaxant effect of sodium nitroprusside vs. control vessels (EC50 = 141 nmol/L vs. EC50 = 0.9 nmol/L; p < 0.001). The effects of PE on Fli1 and collagen-1 were blocked by the in vitro treatment of umbilical arteries by 10 µmol/L canrenone. Similar results were obtained for umbilical arteries pretreated with MBG. These data demonstrate that elevated MBG level is implicated in the development of the fibrosis of umbilical arteries in PE, and that this could be blocked by mineralocorticoid antagonists.

The Leptin, Dopamine and Serotonin Receptors in Hypothalamic POMC-Neurons of Normal and Obese Rodents.

The pro-opiomelanocortin (POMC)-expressing neurons of the hypothalamic arcuate nucleus (ARC) are involved in the control of food intake and metabolic processes. It is assumed that, in addition to leptin, the activity of these neurons is regulated by serotonin and dopamine, but only subtype 2C serotonin receptors (5-HT2CR) was identified earlier on the POMC-neurons. The aim of this work was a comparative study of the localization and number of leptin receptors (LepR), types 1 and 2 dopamine receptors (D1R, D2R), 5-HT1BR and 5-HT2CR on the POMC-neurons and the expression of the genes encoding them in the ARC of the normal and diet-induced obese (DIO) rodents and the agouti mice (A y /a) with the melanocortin obesity. As shown by immunohistochemistry (IHC), all the studied receptors were located on the POMC-immunopositive neurons, and their IHC-content was in agreement with the expression of their genes. In DIO rats the number of D1R and D2R in the POMC-neurons and their expression in the ARC were reduced. In DIO mice the number of D1R and D2R did not change, while the number of LepR and 5-HT2CR was increased, although to a small extent. In the POMC-neurons of agouti mice the number of LepR, D2R, 5-HT1BR and 5-HT2CR was increased, and the D1R number was reduced. Thus, our data demonstrates for the first time the localization of different types of the serotonin and dopamine receptors on the POMC-neurons and a specific pattern of the changes of their number and expression in the DIO and melanocortin obesity.