Leptin Induces Apoptosis via ERK/cPLA2/Cytochrome c Pathway in Human Bone Marrow Stromal Cells

Abstract

Leptin, the Ob gene product, has emerged recently as a key regulator of bone mass. However, the mechanism mediating leptin effect remains controversial. Because the action of leptin is dependent on its receptors, we analyzed their expression in osteoblast-lineage primary human bone marrow stromal cells (hBMSC). Both the short and long forms of leptin receptors were detected in hBMSC. Leptin significantly decreased the viability of hBMSC. This cytotoxic effect was prevented by ZVal-Ala-Asp-fluoromethylketone, a pan-caspase inhibitor, implicating that leptin-induced hBMSC death was caspase-dependent. Further investigation demonstrated that leptin activated caspase-3 and caspase-9, but not caspase-8, and increased the cleavage of poly-(ADP-ribose) polymerase and cytochrome c release into cytosol. Leptin activated ERK, but not p38 and JNK, and up-regulated cPLA2 activity; the latter was abolished by pre-treatment of cells with the MEK inhibitor (PD98059 or U0126) or cPLA2 inhibitor (AACOCF3). PD98059, U0126, and AACOCF3 also diminished the leptin-induced cytochrome c release into cytosol, cell death, and caspase-3 activation. These data indicated that leptin induced hBMSC apoptosis via ERK/cPLA2/cytochrome c pathway with activation of caspase-9 and caspase-3, and cleavage of poly(ADP-ribose) polymerase. To our knowledge, this is the first study demonstrating the direct detrimental effect of leptin on bone cells.Leptin, the protein product encoded by obese gene, is a circulating hormone produced primarily by the adipose tissue and is a multifunctional hormone that plays important roles in body weight homeostasis, neuroendocrine function, fertility, immune function, and angiogenesis (1-3). The biological actions of leptin on target tissues are carried out through interaction with its specific receptor, Ob-R (4). A hallmark of Ob-R expression is the presence of several receptor variants (Ob-Ra through Ob-Rf) that are generated by alternative splicing; they share the same extracellular domain but differ in the length of the transmembrane/cytoplasmic coding regions (2). The long Ob-Rb subtype (Ob-R L ) appears as the functional, signal-transducing isoform, responsible for the action of leptin. The roles of the shorter Ob-R isoforms (Ob-R S ) remain to be characterized. Although leptin's precise sites of action are not known, its effect is thought largely mediated via hypothalamus. However, the wide expression of Ob-R L throughout the body suggests that leptin may also operate directly in peripheral tissues (5). There is now a significant amount of evidence implicating that leptin is active in the periphery (6).Recently, leptin has emerged as a key element in the regulation of bone mass. However, the mechanism by which leptin acts upon the bone remains unclear. Thomas et al. (7) reported that leptin enhanced the differentiation of hBMSC, 1 and Steppan et al. (8) reported that the femurs of leptindeficient ob/ob mice were shorter than those in the normal mice, and intraperitoneal injection of leptin significantly increas...

Functional intrinsic and extrinsic apoptotic pathways in human fetal mesenchymal stem cells

“…Consistent with the absence of caspase-8 cleavage following exposure to SSP (data not shown), SSP-induced cell death could be attenuated by incubation with both caspase-9 and pancaspase inhibitors in a dose-dependent manner (Figure 1f) while a selective caspase-8 inhibitor (IETD-fmk) had no protective effect (not shown). These results show that fetal MSC are very sensitive to apoptotic cell death via the mitochondrial pathway and confirm the importance of mitochondria in stem cell apoptosis, 11 also demonstrated following leptin treatment of adult bone marrow stromal cells 9 and the exposure of neural stem cells to oxidative stress. 10,12 Having established that the intrinsic (mitochondrial) pathway for apoptotic death was functional in fetal MSC, we extended the study to explore the role of the extrinsic (death receptor) pathway.…”

“…6 Preliminary data also suggest that fetal MSC can be differentiated along myogenic and neural lineages, and so represent a ready source of human multipotent cells. Although it might be expected that highly proliferative stem cells would be equipped with efficient death machinery, there is little published work on apoptotic signalling pathways in MSC or bone marrow-derived stromal cells, [7][8][9] and even fewer studies of apoptosis in other stem cells. 10 Since apoptosis is a major cause of graft failure, we hypothesised that there would be robust and functional intrinsic and extrinsic apoptotic signalling pathways in naïve human MSC.…”

Leptin Counteracts Sodium Butyrate-induced Apoptosis in Human Colon Cancer HT-29 Cells via NF-κB Signaling

“…However, we cannot formally exclude the involvement of ERK pathways in this anti-apoptotic effect. On the other hand, leptin has been shown to promote apoptosis in human bone marrow stromal cells through activation of ERK pathways (42) and in adipose tissue (43). We have no clear explanation for these cell type discrepancies but it might be dependent on the type of cell signaling targeted.…”

In vitro Combination Treatment with Perifosine and UCN-01 Demonstrates Synergism against Prostate (PC-3) and Lung (A549) Epithelial Adenocarcinoma Cell Lines

“…To assess nuclear morphology, A549 cells were grown on a single well-chambered slide (Labtek) and incubated with UCN-01 (250 nm) or perifosine (10 m) or a combination of the two for 48 h. Protocol was followed as stated in (28). Combination treatment included both drugs given at the same time, one given for 24 h and then the second added and vice versa.…”