Abstract-Patients with chronic renal failure develop a "uremic" cardiomyopathy characterized by diastolic dysfunction, cardiac hypertrophy, and systemic oxidant stress. Patients with chronic renal failure are also known to have increases in the circulating concentrations of the cardiotonic steroid marinobufagenin (MBG). On this background, we hypothesized that elevations in circulating MBG may be involved in the cardiomyopathy. First, we observed that administration of MBG (10 g/kg per day) for 4 weeks caused comparable increases in plasma MBG as partial nephrectomy at 4 weeks. MBG infusion caused increases in conscious blood pressure, cardiac weight, and the time constant for left ventricular relaxation similar to partial nephrectomy. Decreases in the expression of the cardiac sarcoplasmic reticulum ATPase, cardiac fibrosis, and systemic oxidant stress were observed with both MBG infusion and partial nephrectomy. Next, rats were actively immunized against a MBG-BSA conjugate or BSA control, and partial nephrectomy was subsequently performed. Immunization against MBG attenuated the cardiac hypertrophy, impairment of diastolic function, cardiac fibrosis, and systemic oxidant stress seen with partial nephrectomy without a significant effect on conscious blood pressure. These data suggest that the increased concentrations of MBG are important in the cardiac disease and oxidant stress state seen with renal failure.

Abstract-We have observed recently that experimental renal failure in the rat is accompanied by increases in circulating concentrations of the cardiotonic steroid, marinobufagenin (MBG), and substantial cardiac fibrosis. We performed the following studies to examine whether MBG might directly stimulate cardiac fibroblast collagen production. In vivo studies were performed using the 5/6th nephrectomy model of experimental renal failure (PNx), MBG infusion (MBG), PNx after immunization against MBG, and concomitant PNx and adrenalectomy. Physiological measurements with a Millar catheter and immunohistochemistry were performed. In vitro studies were then pursued with cultured isolated cardiac fibroblasts. We observed that PNx and MBG increased MBG levels, blood pressure, heart size, impaired diastolic function, and caused cardiac fibrosis. PNx after immunization against MBG and concomitant PNx and adrenalectomy had similar blood pressure as PNx but less cardiac hypertrophy, diastolic dysfunction, and cardiac fibrosis. MBG induced increases in procollagen-1 expression by cultured cardiac fibroblasts at 1 nM concentration. These increases in procollagen expression were accompanied by increases in collagen translation and increases in procollagen-1 mRNA without any demonstrable increase in procollagen-1 protein stability. The stimulation of fibroblasts with MBG could be prevented by administration of inhibitors of tyrosine phosphorylation, Src activation, epidermal growth factor receptor transactivation, and N-acetyl cysteine. Based on these findings, we propose that MBG directly induces increases in collagen expression by fibroblasts, and we suggest that this may be important in the cardiac fibrosis seen with experimental renal failure. (Hypertension. 2007;49:215-224.)

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The cardiotonic steroid marinobufagenin (MBG) has been implicated in the pathogenesis of experimental uremic cardiomyopathy, which is characterized by progressive cardiac fibrosis. We examined whether the transcription factor Friend leukemia integration-1 (Fli-1) might be involved in this process. Fli-1-knockdown mice demonstrated greater cardiac collagen-1 expression and fibrosis compared with wild-type mice; both developed increased cardiac collagen expression and fibrosis after 5/6 nephrectomy. There was a strong inverse relationship between the expressions of Fli-1 and procollagen in primary culture of rat cardiac and human dermal fibroblasts as well as a cell line derived from renal fibroblasts and MBG-induced decreases in nuclear Fli-1 as well as increases in procollagen-1 expression in these cells. Transfection of a Fli-1 expression vector prevented increased procollagen-1 expression from MBG. MBG exposure induced a rapid translocation of the δ-isoform of protein kinase C (PKCδ) to the nucleus. This translocation was prevented by pharmacological inhibition of phospholipase C, and MBG-induced increases in procollagen-1 expression were prevented with a PKCδ- but not a PKCα-specific inhibitor. Finally, immunoprecipitation studies strongly suggest that MBG induced phosphorylation of Fli-1. We feel these data support a causal relationship with MBG-induced translocation of PKCδ, which results in phosphorylation of as well as decreases in nuclear Fli-1 expression, which, in turn, leads to increases in collagen production. Should these findings be confirmed, we speculate that this pathway may represent a therapeutic target for uremic cardiomyopathy as well as other conditions associated with excessive fibrosis.

It has been widely acknowledged that non-coding RNAs are master-regulators of genomic functions. However, the significance of the presence of ncRNA within introns has not received proper attention. ncRNA within introns are commonly produced through the post-splicing process and are specific signals of gene transcription events, impacting many other genes and modulating their expression. This study, along with the following discussion, details the association of thousands of ncRNAs—snoRNA, miRNA, siRNA, piRNA and long ncRNA—within human introns. We propose that such an association between human introns and ncRNAs has a pronounced synergistic effect with important implications for fine-tuning gene expression patterns across the entire genome.

Objective Obese leptin deficient (ob/ob) mice are a model of adiposity that displays increased levels of fat, glucose and liver lipids. Our hypothesis is that HO-1 overexpression ameliorates fatty liver development. Design and Methods Obese mice were administered cobalt protoporphyrin (CoPP) and stannic mesoporphyrin (SnMP) for 6 weeks. Heme, HO-1, HO activity, PGC1α, FGF21, glycogen content and lipogenesis were assessed. Results CoPP administration increased hepatic HO-1 protein levels and HO activity, decreased hepatic heme, body weight gain, glucose levels and resulted in decreased steatosis. Increased levels of HO-1 produced a decrease in lipid droplet size, FAS levels involving recruitment of FGF21, PPARα and Glut 1. These beneficial effects were reversed by inhibition of HO activity. Conclusion Increased levels of HO-1 and HO activity reduced the levels of obesity by reducing hepatic heme and lipid accumulation. These changes were manifested by decreases in cellular heme, increases in FGF21, glycogen content and fatty liver. The beneficial effect of HO-1 induction results from an increase in PPARα and FGF21 levels and a decrease in PGC1α, levels they were reversed by SnMP. Low levels of HO-1 and HO activity are responsible for fatty liver.

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Among thousands of non-protein-coding RNAs which have been found in humans, a significant group represents snoRNA molecules that guide other types of RNAs to specific chemical modifications, cleavages, or proper folding. Yet, hundreds of mammalian snoRNAs have unknown function and are referred to as "orphan" molecules. In 2006, for the first time, it was shown that a particular orphan snoRNA (HBII-52) plays an important role in the regulation of alternative splicing of the serotonin receptor gene in humans and other mammals. In order to facilitate the investigation of possible involvement of snoRNAs in the regulation of pre-mRNA processing, we developed a new computational web resource, snoTARGET, which searches for possible guiding sites for snoRNAs among the entire set of human and rodent exonic and intronic sequences. Application of snoTARGET for finding possible guiding sites for a number of human and rodent orphan C/D-box snoRNAs showed that another subgroup of these molecules (HBII-85) have statistically elevated guiding preferences toward exons compared to introns. Moreover, these energetically favorable putative targets of HBII-85 snoRNAs are non-randomly associated with genes producing alternatively spliced mRNA isoforms. The snoTARGET resource is freely available at: (http://hsc.utoledo.edu/depts/bioinfo/snotarget.html).