The relationship between miRNA-26b and connective tissue growth factor in rat models of aortic banding and debanding

Background/Aims@#Connective tissue growth factor (CTGF) is a profibrotic factor implicated in pressure overload-mediated myocardial fibrosis. In this study, we determined the role of predicted CTGF-targeting microRNAs (miRNAs) in rat models of aortic stenosis and reverse cardiac remodeling. @*Methods@#Minimally invasive ascending aortic banding was performed in 24 7-week-old male Sprague-Dawley rats, which were divided into three groups. The banding group consisted of eight rats that were sacrificed immediately after 6 weeks of aortic constriction. The debanding group underwent aortic constriction for 4 weeks and was sacrificed 2 weeks after band removal. The third group underwent sham surgery. We investigated the expression of CTGF, transforming growth factor-β1 (TGFβ1), and matrix metalloproteinase-2 using ELISA and examined miRNA-26b, miRNA-133a, and miRNA-19b as predicted CTGF-targeting miRNAs based on miRNA databases in 24-hour TGFβ-stimulated and TGFβ- washed fibroblasts and myocardial tissues from all subjects. @*Results@#CTGF was elevated in 24-hour TGFβ-stimulated fibroblasts and decreased in 24-hour TGFβ-washed fibroblasts. miRNA-26b was significantly increased in TGFβ-washed fibroblasts compared with control and TGFβ-stimulated fibroblasts (p < 0.05). CTGF expression was significantly higher in the banding group than that in the sham and debanding groups. The relative expression levels of miRNA-26b were higher in the debanding group than in the banding group. @*Conclusions@#The results of our study using models of aortic banding and debanding suggested that miRNA-26b was significantly increased after aortic debanding. The in vitro model yielded the same results: miRNA-26b was upregulated after removal of TGFβ from fibroblasts.

The relationship between miRNA-26b and connective tissue growth factor in rat models of aortic banding and debanding

Background/Aims@#Connective tissue growth factor (CTGF) is a profibrotic factor implicated in pressure overload-mediated myocardial fibrosis. In this study, we determined the role of predicted CTGF-targeting microRNAs (miRNAs) in rat models of aortic stenosis and reverse cardiac remodeling. @*Methods@#Minimally invasive ascending aortic banding was performed in 24 7-week-old male Sprague-Dawley rats, which were divided into three groups. The banding group consisted of eight rats that were sacrificed immediately after 6 weeks of aortic constriction. The debanding group underwent aortic constriction for 4 weeks and was sacrificed 2 weeks after band removal. The third group underwent sham surgery. We investigated the expression of CTGF, transforming growth factor-β1 (TGFβ1), and matrix metalloproteinase-2 using ELISA and examined miRNA-26b, miRNA-133a, and miRNA-19b as predicted CTGF-targeting miRNAs based on miRNA databases in 24-hour TGFβ-stimulated and TGFβ- washed fibroblasts and myocardial tissues from all subjects. @*Results@#CTGF was elevated in 24-hour TGFβ-stimulated fibroblasts and decreased in 24-hour TGFβ-washed fibroblasts. miRNA-26b was significantly increased in TGFβ-washed fibroblasts compared with control and TGFβ-stimulated fibroblasts (p < 0.05). CTGF expression was significantly higher in the banding group than that in the sham and debanding groups. The relative expression levels of miRNA-26b were higher in the debanding group than in the banding group. @*Conclusions@#The results of our study using models of aortic banding and debanding suggested that miRNA-26b was significantly increased after aortic debanding. The in vitro model yielded the same results: miRNA-26b was upregulated after removal of TGFβ from fibroblasts.

Increased Immunoreactivity of LGR4 in Histologically Aggressive Basal Cell Carcinoma

No abstract available.

Paricalcitol attenuates lipopolysaccharide-induced inflammation and apoptosis in proximal tubular cells through the prostaglandin E₂ receptor EP4

BACKGROUND: Vitamin D is considered to exert a protective effect on various renal diseases but its underlying molecular mechanism remains poorly understood. This study aimed to determine whether paricalcitol attenuates inflammation and apoptosis during lipopolysaccharide (LPS)-induced renal proximal tubular cell injury through the prostaglandin E₂ (PGE₂) receptor EP4. METHODS: Human renal tubular epithelial (HK-2) cells were pretreated with paricalcitol (2 ng/mL) for 1 hour and exposed to LPS (1 μg/mL). The effects of paricalcitol pretreatment in relation to an EP4 blockade using AH-23848 or EP4 small interfering RNA (siRNA) were investigated. RESULTS: The expression of cyclooxygenase-2, PGE₂, and EP4 were significantly increased in LPS-exposed HK-2 cells treated with paricalcitol compared with cells exposed to LPS only. Paricalcitol prevented cell death induced by LPS exposure, and the cotreatment of AH-23848 or EP4 siRNA offset these cell-protective effects. The phosphorylation and nuclear translocation of p65 nuclear factor-kappaB (NF-κB) were decreased and the phosphorylation of Akt was increased in LPS-exposed cells with paricalcitol treatment. AH-23848 or EP4 siRNA inhibited the suppressive effects of paricalcitol on p65 NF-κB nuclear translocation and the activation of Akt. The production of proinflammatory cytokines and the number of terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive cells were attenuated by paricalcitol in LPS exposed HK-2 cells. The cotreatment with an EP4 antagonist abolished these anti-inflammatory and antiapoptotic effects. CONCLUSION: EP4 plays a pivotal role in anti-inflammatory and antiapoptotic effects through Akt and NF-κB signaling after paricalcitol pretreatment in LPS-induced renal proximal tubule cell injury.

Myocardial Mechanics in a Rat Model with Banding and Debanding of the Ascending Aorta

BACKGROUND: Aortic banding and debanding models have provided useful information on the development and regression of left ventricular hypertrophy (LVH). In this animal study, we aimed to evaluate left ventricular (LV) deformation related to the development and regression of LVH. METHODS: Minimally invasive ascending aorta banding was performed in rats (10 Sprague Dawley rats, 7 weeks). Ten rats underwent a sham operation. Thirty-five days later, the band was removed. Echocardiographic and histopathologic analysis was assessed at pre-banding, 35 days of banding and 14 days of debanding. RESULTS: Banding of the ascending aorta created an expected increase in the aortic velocity and gradient, which normalized with the debanding procedure. Pressure overload resulted in a robust hypertrophic response as assessed by gross and microscopic histology, transthoracic echocardiography [heart weight/tibia length (g/m); 21.0 +/- 0.8 vs. 33.2 +/- 2.0 vs. 26.6 +/- 2.8, p < 0.001]. The circumferential (CS) and radial strains were not different between the groups. However, there were significant differences in the degree of fibrosis according to the banding status (fibrosis; 0.10 +/- 0.20% vs. 5.26 +/- 3.12% vs. 4.03 +/- 3.93%, p = 0.003), and global CS showed a significant correlation with the degree of myocardial fibrosis in this animal model (r = 0.688, p = 0.028). CONCLUSION: In this animal study, simulating a severe LV pressure overload state, a significant increase in the LV mass index did not result in a significant reduction in the LV mechanical parameters. The degree of LV fibrosis, which developed with pressure overload, was significantly related to the magnitude of left ventricular mechanics.

Desalinated underground seawater of Jeju Island (Korea) improves lipid metabolism in mice fed diets containing high fat and increases antioxidant potential in t-BHP treated HepG2 cells

This study was performed to investigate the effect of desalinated underground seawater (named as 'magma seawater', MSW) of Jeju Island in Korea on lipid metabolism and antioxidant activity. MSW was collected from underground of Han-Dong in Jeju Island, and freely given to high fat diet (HFD)-fed C57BL/6 mice for 10 weeks. Although there were no significant differences in the body weight changes and plasma lipid levels, hepatic triglyceride levels were significantly lower in the MSW group than in the normal tap water (TW)-drunken control group. Furthermore, the activity of fatty acid synthase (FAS) was significantly decreased and carnitine palmitoyltransferase (CPT) activity was increased in MSW group compared to TW group. Similarly, real-time PCR analysis revealed that mRNA expressions of lipogenic genes were lowered in MSW groups compared to the control group. In a morphometric observation on the liver tissue, accumulation of fats was remarkably reduced in MSW group. Meanwhile, in vitro assay, free radical scavenging activity measured by using diphenylpicrylhydrazyl (DPPH) was increased in MSW group. The 2'-7'-dichlorofluorescein diacetate (DCF-DA) staining followed with fluorescent microscopy showed a low intensity of fluorescence in MSW-treated HepG2 cells, compared to TW-treated HepG2 cells, which indicated that the production of reactive oxygen species by tert-butyl hydroperoxide (t-BHP) in HepG2 cells was decreased by MSW treatment. The antioxidant effect of MSW on t-BHP-induced oxidative stress in HepG2 cells was supported by the increased activities of intracellular antioxidant enzymes such as catalase and glutathione reductase. From these results, we speculate that MSW has an inhibitory effect on lipogenesis in liver and might play a protective role against cell damage by t-BHP-induced oxidative stress.

Tissue-specific expression and subcellular localization of ALADIN, the absence of which causes human triple A syndrome

Triple A syndrome is a rare genetic disorder caused by mutations in the achalasia-addisonianism-alacrima syndrome (AAAS) gene which encodes a tryptophan aspartic acid (WD) repeat-containing protein named alacrima-achalasia-adrenal insufficiency neurologic disorder (ALADIN). Northern blot analysis shows that the 2.1 kb AAAS mRNA is expressed in various tissues with stronger expression in testis and pancreas. We show that human ALADIN is a protein with an apparent molecular weight of 60 kDa, and expressed in the adrenal gland, pituitary gland and pancreas. Furthermore, biochemical analysis using anti-ALADIN antibody supports the previous finding of the localization of ALADIN in the nuclear membrane. The mutations S544G and S544X show that alteration of S544 residue affects correct targeting of ALADIN to the nuclear membrane.