leptin and pyrazolanthrone

Leptin, an adipokine secreted by adipose tissue, induces cell invasion and metastasis. MMP7 is a member of the matrix metalloproteinase family that plays an important role in cell invasion. Here we evaluate the possible role and underlying mechanism of MMP7 in the leptin-mediated cell invasion in ovarian cancer cell lines. All experiments were carried out in cultured SKOV3, OVCAR3, and CaoV-3 ovarian cell lines. MMP7 expression was determined using the Western blot following treatment to various concentrations of leptin for defined time intervals. The activation of ERK, JNK, and P38 MAP kinases were determined using Western blotting. Wound healing and BD matrigel invasion assays were used to measure cell migration and invasion. The siRNA approach and pharmacological inhibitors of ERK and JNK pathway were used to confirm the receptor-dependent effect of leptin and a role for ERK and JNK pathway. Zymography assay was employed to determine MMP2 and MMP9 activation. Results show that leptin induces ERK1/2 and JNK1/2 activation and subsequently promotes MMP7 expression in SKOV3 (4.8 ± 0.14 fold of control, P < 0.01) and OVCAR3 (3.1 ± 0.19 fold of control, P < 0.01) ovarian cancer cell lines. These effects was reversed by knockdown of OB-Rb and/or pre-incubation with PD98059 (ERK1/2 inhibitor), SP600125 (JNK1/2 inhibitor). Gelatin zymography showed that MMP7 gene silencing attenuated leptin-induced MMP9 activation in SKOV3 cell line. Taken together, our results suggest new evidences for a modulatory effect of leptin in regulation of ovarian cancer cell invasion by stimulating MMP7 expression via ERK and JNK pathways.

Topics: Anthracenes; Cell Line, Tumor; Cell Movement; Female; Flavonoids; Gene Knockdown Techniques; Humans; Leptin; MAP Kinase Signaling System; Matrix Metalloproteinase 7; Neoplasm Invasiveness; Ovarian Neoplasms

Angiotensin-(1-7) (Ang-[1-7]), recognized as a new bioactive peptide in the renin-angiotensin system, shows biological and pharmacological properties in diabetic cardiovascular diseases. The leptin-induced p38 mitogen-activated protein kinase (MAPK) pathway has been reported to contribute to high glucose (HG)-induced injury. In the present study, we showed the mechanism of how Ang-(1-7) can protect against HG-stimulated injuries in H9c2 cells.. H9c2 cells were treated with 35 mmol/L glucose (HG) for 24 h to establish a model of HG-induced damage. Apoptotic cells were observed by Hoechst 33258 staining. Cell viability was analyzed by cell counter kit-8. The expression of protein was detected by western blot. Reactive oxygen species was tested by 2',7'-dichlorodihydrofluorescein diacetate staining. Mitochondrial membrane potential was measured by 5,5',6,6'-Tetrachloro-1,1',3,3'-tetraethyl-imidacarbocyanine iodide staining.. The present results showed that treating H9c2 cells with HG obviously enhanced the expressions of both the leptin and phosphorylated (p)-MAPK pathway. However, the overexpression levels of leptin and p-p38 MAPK/p-extracellular signal-regulated protein kinase 1/2 (ERK1/2), but not p-c-Jun N-terminal kinase, were significantly suppressed by treatment of the cells with Ang-(1-7). Additionally, leptin antagonist also markedly suppressed the overexpressions of p38 and ERK1/2 induced by HG, whereas leptin antagonist had no influence on the overexpression of c-Jun N-terminal kinase. More remarkable, Ang-(1-7), leptin antagonist, SB203580 or SP600125, respectively, significantly inhibited the injuries induced by HG, such as the increased cell viability, decreased apoptotic rate, reduction of ROS production and increased mitochondrial membrane potential. Furthermore, the overexpressions of p38 MAPK, ERK1/2 and leptin were suppressed by N-actyl-L-cystine.. The present findings show that Ang-(1-7) protects from HG-stimulated damage as an inhibitor of the reactive oxygen species-leptin-p38 MAPK/ERK1/2 pathways, but not the leptin-c-Jun N-terminal kinase pathway in vitro.

Topics: Acetylcysteine; Angiotensin I; Animals; Anthracenes; Apoptosis; Cell Line; Glucose; Imidazoles; Leptin; MAP Kinase Signaling System; Membrane Potential, Mitochondrial; Myocytes, Cardiac; Peptide Fragments; Pyridines; Rats; Reactive Oxygen Species

Intracellular signaling pathways regulated by leptin have largely been studied in metabolically important organs such as adipose tissue and peripheral blood mononuclear cells, suggesting that leptin plays a key role in pathophysiology of insulin resistance. However, whether synthetic analog of leptin, metreleptin, has similar effects on cardiac myocytes (CM) and uterine smooth muscle cells (USMC) has not yet been studied. Hence, in order to address these questions, we extended previous observations and investigated in vitro signaling study whether metreleptin may activate key signaling pathways. We observed that metreleptin activates Jak2 and STAT3 signaling pathways in dose- and time-dependent manner in CM and USMC. Also, we found that metreleptin increases ERK1/2, JNK and/or p38 phosphorylation in CM. In vitro metreleptin administration also increased ERK1/2 and/or p38 phosphorylation in USMC. By contrast, JNK was not regulated by in vitro metreleptin administration in USMC. Moreover, metreleptin-activated all signaling pathways were blocked by pre-treatment of PD98095 (ERK inhibitor), SB203580 (p38 inhibitor) and/or SP600125 (JNK inhibitor), respectively. Finally, metreleptin increased cell size (hypertrophy) in both CM and USMC. Our data provide novel insights into the role of Jak2, STAT3, ERK1/2, JNK and/or p38 as probable mediators of the action of leptin in regulating hypertrophy in CM and USMC.

Topics: Anthracenes; Cell Line; Enzyme Activation; Extracellular Signal-Regulated MAP Kinases; Female; Humans; Imidazoles; Janus Kinase 2; JNK Mitogen-Activated Protein Kinases; Leptin; Muscle, Smooth; Myocytes, Cardiac; Myocytes, Smooth Muscle; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Pyridines; Signal Transduction; STAT3 Transcription Factor; Uterus

Obesity is a risk factor for thoracic ossification of ligament flavum (TOLF) that is characterized by ectopic bone formation in the spinal ligaments. Hyperleptinemia is a common feature of obese people, and leptin, an adipocyte-derived cytokine with proliferative and osteogenic effects in several cell types, is believed to be an important factor in the pathogenesis of TOLF. However, how leptin might stimulate cell osteogenic differentiation in TOLF is not totally understood. We reported here that leptin-induced osteogenic effect in TOLF cells is associated with activation of signaling molecules STAT3, JNK, and ERK1/2 but not p38. Blocking STAT3 phosphorylation with a selective inhibitor, AG490, significantly abolished leptin-induced osteogenic differentiation of TOLF cells, whereas blocking ERK1/2 and JNK phosphorylation with their selective inhibitors PD98059 and SP600125, respectively, had only marginal effects. In addition, we showed that STAT3 interacted with Runt-related transcription factor 2 (Runx2) in the nucleus, and STAT3, Runx2, and steroid receptor coactivator steroid receptor coactivator-1 were components of the transcription complex recruited on Runx2 target gene promoters in response to leptin treatment. Our experiments identified STAT3, Runx2, and steroid receptor coactivator-1 as critical molecules in mediating leptin-stimulated cell osteogenesis in TOLF.

Topics: Aged; Anthracenes; Bone Diseases; Cell Differentiation; Core Binding Factor Alpha 1 Subunit; Female; Flavonoids; Histone Acetyltransferases; Humans; Leptin; Ligaments, Articular; Male; Middle Aged; Mitogen-Activated Protein Kinase 3; Models, Biological; Nuclear Receptor Coactivator 1; STAT3 Transcription Factor; Thoracic Diseases; Transcription Factors

The hypertrophy of vascular smooth muscle cells (VSMCs) is critical in vascular remodeling associated with hypertension, atherosclerosis, and restenosis. Recently, leptin has appeared to play a pivotal role in vascular remodeling. However, the mechanism by which leptin induces hypertrophy in vascular smooth muscle cells is still unknown. We studied the role of leptin as a potential hypertrophic factor in rat VSMCs. In the present study, leptin significantly increased [(3)H]leucine incorporation and the total protein/DNA ratio in VSMCs. The maximal hypertrophic effect was at 100ng/ml of leptin. Leptin induced phosphorylation and activation of p38 mitogen-activated protein (p38 MAP) kinase and of signal transducers and activators of transcription 3 in a concentration- and time-dependent manner. A p38 MAP kinase inhibitor SB203580 significantly inhibited leptin-induced hypertrophy, AG490 (a JAK2 inhibitor) partially inhibited it, and other MAP kinase inhibitors, PD98059 (an ERK inhibitor) and SP600125 (a JNK inhibitor), had no effect. These results indicate that leptin directly stimulates cellular hypertrophy via p38 MAP kinase in rat VSMCs.

Topics: Animals; Anthracenes; Aorta; Arteriosclerosis; Blotting, Western; Cells, Cultured; DNA; DNA-Binding Proteins; Dose-Response Relationship, Drug; Electrophoresis, Polyacrylamide Gel; Enzyme Activation; Enzyme Inhibitors; Flavonoids; Hypertrophy; Leptin; Leucine; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Rats; Rats, Sprague-Dawley; STAT3 Transcription Factor; Time Factors; Trans-Activators; Tyrphostins