Paricalcitol Attenuates Cardiac Fibrosis and Expression of Endothelial Cell Transition Markers in Isoproterenol-Induced Cardiomyopathic Rats.

OBJECTIVES: Acute cardiomyopathy is a health problem worldwide. Few studies have shown an association between acute cardiomyopathy and low vitamin D status. Paricalcitol, a vitamin D receptor activator, clinically benefits patients with advanced kidney disease. The effect of paricalcitol supplement on cardiac remodeling in cardiomyopathic rats is unknown. This experimental study investigated the effect of paricalcitol in rats with cardiomyopathy induced by isoproterenol. DESIGN: Prospective, randomized, controlled experimental study. SETTING: Hospital-affiliated animal research institution. SUBJECTS: Eight-week-old male Wistar-Kyoto rats. INTERVENTIONS: Male Wistar-Kyoto rats were first injected intraperitoneally with isoproterenol to create a rat model of acute cardiomyopathy. Then paricalcitol was administered intraperitoneally to isoproterenol-injected rats at a dosage of 200 ng three times a week for 3 weeks. Relevant cardiomyopathy-related variables were measured regularly in three groups of rats, controls, isoproterenol, and isoproterenol plus paricalcitol. Rat hearts were obtained for evaluation of cardiac fibrosis using Masson trichrome staining and commercially available software, and evaluation of cell transition using immunofluorescence staining analysis. MEASUREMENTS AND MAIN RESULTS: Isoproterenol infusions generated significant cardiac fibrosis (p < 0.001). Subsequent paricalcitol treatment attenuated the isoproterenol-induced cardiac fibrosis (p = 0.006). Fluorescence showed colocalization of endothelial and fibroblast cell markers (cluster differentiation 31 and α-smooth muscle actin, respectively) in the isoproterenol-treated hearts. Paricalcitol injections attenuated the isoproterenol-induced fluorescence intensity of two cell markers (p < 0.01). CONCLUSIONS: Paricalcitol injections may ameliorate isoproterenol-induced cardiac fibrosis possibly through regulating cell transition.

Loss of Egr-1 sensitizes pancreatic ß-cells to palmitate-induced ER stress and apoptosis.

UNLABELLED: Pancreatic ß-cells are particularly susceptible to fatty-acid-induced endoplasmic reticulum (ER) stress and apoptosis. To understand how ß-cells sense fatty acid stimuli and translate into a long-term adaptive response, we investigated whether palmitic acid (PA) regulates early growth response-1 (Egr-1), an immediate-early transcription factor, which is induced by many environmental stimuli and implicated in cell proliferation, differentiation, and apoptosis. We found that Egr-1 was rapidly and transiently induced by PA in MIN6 insulinoma cells, which was accompanied by calcium influx and ERK1/2 phosphorylation. Calcium chelation and MEK1/2 inhibition blocked PA-induced Egr-1 upregulation, suggesting that PA induces Egr-1 expression through a calcium influx-MEK1/2-ERK1/2 cascade. Knockdown of Egr-1 increased PA-induced caspase-3 activation and ER stress markers and decreased PA-induced Akt phosphorylation and insulin secretion and signaling. Akt replenishment and insulin supplementation rescued PA-induced apoptosis in Egr-1 knockdown cells. These results suggest that the absence of Egr-1 loses its ability to couple the short-term insulin/Akt pathway to long-term survival adaptation. Finally, Egr-1-deficient mouse islets are more susceptible to ex vivo stimuli of apoptosis. In human pancreatic tissues, EGR1 expression correlated with expression of ER stress markers and anti-apoptotic gene. In conclusion, Egr-1 is induced by PA and further attempts to rescue ß-cells from ER stress and apoptosis through improving insulin/Akt signaling. Our study underscores Egr-1 as a critical early sensor in pancreatic ß-cells to translate fatty acid stimuli into a cellular adaptation mechanism. KEY MESSAGE: PA stimulates Egr-1 expression via a calcium influx-MEK1/2-ERK1/2-Elk-1 cascade. Egr-1 attenuates PA-induced ER stress and apoptosis. Egr-1 maintains Akt survival pathway to protect ß-cells from PA-induced apoptosis. Egr-1-deficient islets are prone to ex vivo stimuli of apoptosis. Human EGR1 expression correlates with genes for ER stress and anti-apoptosis.

Gypenosides inhibits migration and invasion of human oral cancer SAS cells through the inhibition of matrix metalloproteinase-2 -9 and urokinase-plasminogen by ERK1/2 and NF-kappa B signaling pathways.

Gypenosides (Gyp), found in Gynostemma pentaphyllum Makino, has been used as a folk medicine in the Chinese population for centuries and is known to have diverse pharmacologic effects, including anti-proliferative and anti-cancer actions. However, the effects of Gyp on prevention from invasion and migration of oral cancer cells are still unsatisfactory. The purpose of this study was to investigate effects of Gyp treatment on migration and invasion of SAS human oral cancer cells. SAS cells were cultured in the presence of 90 and 180 µg/mL Gyp for 24 and 48 hours. Gyp induced cytotoxic effects and inhibited SAS cells migration and invasion in dose- and time-dependent response. Wound-healing assay and boyden chamber assay were carried out to investigate Gyp-inhibited migration and invasion of SAS cells. Gyp decreased the abundance of several proteins, including nuclear factor-kappa B (NF-κB), cyclooxygenase-2 (COX-2), extracellular signal-regulated kinase 1/2 (ERK1/ 2), matrix metalloproteinase-9, -2 (MMP-9, -2), sevenless homolog (SOS), Ras, urokinase-type plasminogen activator (uPA), focal adhesion kinase (FAK) and RAC-alpha serine/threonine-protein kinase (Akt), in a time-dependent manner. In addition, Gyp decreased mRNA levels of MMP-2, MMP-7, MMP-9 but did not affect FAK and Rho A mRNA levels in SAS cells. These results provide evidences for the role of Gyp as a potent anti-metastatic agent, which can markedly inhibit the metastatic and invasive capacity of oral cancer cells. The inhibition of NF-κB and MMP-2, -7 and -9 signaling may be one of the mechanisms that is present in Gyp-inhibited cancer cell invasion and migration.

Gypenosides causes DNA damage and inhibits expression of DNA repair genes of human oral cancer SAS cells.

Gypenosides (Gyp) are the major components of Gynostemma pentaphyllum Makino, a Chinese medical plant. Recently, Gyp has been shown to induce cell cycle arrest and apoptosis in many human cancer cell lines. However, there is no available information to address the effects of Gyp on DNA damage and DNA repair-associated gene expression in human oral cancer cells. Therefore, we investigated whether Gyp induced DNA damage and DNA repair gene expression in human oral cancer SAS cells. The results from flow cytometric assay indicated that Gyp-induced cytotoxic effects led to a decrease in the percentage of viable SAS cells. The results from comet assay revealed that the incubation of SAS cells with Gyp led to a longer DNA migration smear (comet tail) when compared with control and this effect was dose-dependent. The results from real-time PCR analysis indicated that treatment of SAS cells with 180 mug/ml of Gyp for 24 h led to a decrease in 14-3-3sigma, DNA-dependent serine/threonine protein kinase (DNAPK), p53, ataxia telangiectasia mutated (ATM), ataxia-telangiectasia and Rad3-related (ATR) and breast cancer gene 1 (BRCA1) mRNA expression. These observations may explain the cell death caused by Gyp in SAS cells. Taken together, Gyp induced DNA damage and inhibited DNA repair-associated gene expressions in human oral cancer SAS cells in vitro.