Original articleSHP2-mediated signaling cascade through gp130 is essential for LIF-dependent ICaL, [Ca2+]i transient, and APD increase in cardiomyocytes
Introduction
Leukemia inhibitory factor (LIF) is a member of the IL-6 family of cytokines that induces a wide range of responses in a variety of cells [1]. LIF is known to have various effects on cell growth, differentiation, and function [2], [3], [4]. The receptors of the IL-6 family of cytokines have common subunits, gp130 [5], [6]. The binding of the IL-6 family cytokines to their receptors activates Janus kinases (JAK1, JAK2, and TYK2) [7], [8], leading to the recruitment of signal transducing molecules such as protein tyrosine phosphatase 2 (SHP2) and signal transducers and activators of transcription3 (STAT3) [9], [10], [11]. It is considered that there are two major cascades downstream of gp130, i.e. JAK/STAT3 and SHP2/ERK [12], [13].
In cardiomyocytes, we have reported that LIF induces cardiac hypertrophy [14]. The JAK/STAT3 pathway plays an important role in mediating this cardiac hypertrophy.
On the other hand, we have reported that LIF increases L-type Ca2+ current (ICaL) and intracellular Ca2+ concentration ([Ca2+]i transient) in cardiomyocytes, and this ICaL augmentation is independent of PKA but dependent on mitogen-activated protein kinase (MEK) [15]. We have also reported that LIF phosphorylates the serine residue at the position 1829 of the Cav1.2 subunit via the actions of extracellular signal-regulated kinase (ERK) and that this phosphorylation increases ICaL in cardiomyocytes [16]. Therefore, the MEK/ERK pathway might be involved in the LIF-mediated increase of ICaL. There may be, however, a possible crosstalk between the JAK/STAT3 pathway and the MEK/ERK pathway downstream of gp130, and therefore the role of the JAK/STAT3 pathway in the LIF-mediated increase of ICaL remains to be elucidated.
In this study, using the knock-in mouse line, we attempted to clarify that it is not the JAK/STAT3 pathway, but rather the SHP2/ERK pathway below gp130 that plays an important role in this ICaL augmentation.
Section snippets
Materials
The knock-in mouse line [12], in which the SHP2 signal was disrupted by replacing the mouse gp130 gene with the human gp130 mutant cDNA (gp130F759/F759 group), and wild-type mice (WT group) were used for this experiment. These mice were 6–10 weeks of age and weighed 21–28 g. In the gp130F759/F759 group, only the JAK/STAT3 cascade can be activated, and both the JAK/STAT3 and SHP2/ERK cascades can be activated in the WT group.
Cell preparation for the patch clamp
After 1000 units of heparin and 50 mg of sodium pentobarbital were
L-type Ca2+ current
The cardiomyocytes isolated from the WT group and the gp130F759/F759 group appeared similar. There was no difference in the cell capacitance between the WT group (138.5 ± 1.9 pF) and the gp130F759/F759 group (142.0 ± 2.5 pF) and there was no difference in the ICaL density of the control (WT group: 7.54 ± 0.20 pA/pF, n = 76, gp130F759/F759 group: 7.80 ± 0.25 pA/pF, n = 54, NS).
Figs. 1A and B show representative traces of the control and the administration of LIF or IL-6 + sIL-6r. Figs. 1C and D show the
Discussion
These experiments, using the knock-in mice, clarified the distinct signal transduction pathway of ICaL augmentation by LIF.
We have reported that LIF increases ICaL by the phosphorylation of the Cav1.2 subunit via the actions of ERK and that this LIF-mediated increase of ICaL might be independent of PKA and involve the MEK/ERK pathway. However, the existence of crosstalk between the JAK/STAT3 and the MEK/ERK pathways could not be ruled out. In the knock-in mice, gp130F759/F759, used in this
Acknowledgments
This work was supported by the Suntory Fund for Advanced Cardiac Therapeutics, Keio University School of Medicine. The transgenic mouse was kindly provided from Dr. Takuya Ohtani, Osaka University School of Medicine.
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