Korean J Physiol Pharmacol.  2008 Dec;12(6):337-342. 10.4196/kjpp.2008.12.6.337.

Diversity of Ion Channels in Human Bone Marrow Mesenchymal Stem Cells from Amyotrophic Lateral Sclerosis Patients

Affiliations
  • 1Division of Molecular and Life Sciences, Hanyang University, Ansan, 426-791, Korea.
  • 2Department of Neurology, Hanyang University Hospital, Seoul 133-792, Korea.
  • 3Bioengineering Institute, CoreStem Inc., Seoul 133-822, Korea.
  • 4Department of Biomedical Engineering, College of Medicine, Chungbuk National University, Cheongju 361-763, Korea.
  • 5Department of Physiology, College of Medicine, Chungbuk National University, Cheongju 361-763, Korea. yangmik@chungbuk.ac.kr

Abstract

Human bone marrow mesenchymal stem cells (hBM-MSCs) represent a potentially valuable cell type for clinical therapeutic applications. The present study was designed to evaluate the effect of long-term culturing (up to 10th passages) of hBM-MSCs from eight individual amyotrophic lateral sclerosis (ALS) patients, focusing on functional ion channels. All hBM-MSCs contain several MSCs markers with no significant differences, whereas the distribution of functional ion channels was shown to be different between cells. Four types of K+ currents, including noise-like Ca+2-activated K+ current (IKCa), a transient outward K+ current (Ito), a delayed rectifier K+ current (IKDR), and an inward-rectifier K+ current (Kir) were heterogeneously present in these cells, and a TTX-sensitive Na+ current (INa,TTX) was also recorded. In the RT-PCR analysis, Kv1.1, heag1, Kv4.2, Kir2.1, MaxiK, and hNE-Na were detected. In particular, INa,TTX showed a significant passage-dependent increase. This is the first report showing that functional ion channel profiling depend on the cellular passage of hBM-MSCs

Keyword

Bone marrow; Stem cells; Functional ion channels; Tetrodotoxin-sensitive Na+ current; Passage-dependency

MeSH Terms

Amyotrophic Lateral Sclerosis
Bone Marrow
Humans
Ion Channels
Mesenchymal Stromal Cells
Stem Cells
Ion Channels

Figure

  • Fig. 1. Characterization of hBM-MSCs. (A) Flow cytometry analysis showing that hBM-MSCswere positive for CD29, CD44, CD105, and CD73 and were negative for CD34, CD45, and HLA-DR. The table shows mean values (%). (B) hBM-MSCs expressed markers for OPN, LIFR, ABCG2, CXCR4, CD44, collagen X, collagen1 and alpha1. The hBM-MSCs RNAs were obtained from different donors (I and II, n=4) (C) Differentiation capacity of hBM-MSCs to adipocytes (upper) and osteoblasts (lower). 200× magnification.

  • Fig. 2. Different patterns of membrane currents recorded in hBM-MSCs. Current traces elicited by the voltage step (inset) in hBM-MSCs. (A) a slowly activating current similar to IKDR at potentials from +20 to +100 mV that coexisted with IKCa. (B) Ito (arrow) with IKCa. (C) INa,TTX (arrow) with IKDR and IKCa. (D) IKir (arrow) with IKDR and IKCa. (E) Current-voltage relationships were plotted from A, B, C and D.

  • Fig. 3. Pharmacological effects of functional ion channels in hBM-MSCs. (A) Membrane currents were recorded in the presence of 10 mM TEA or co-application of TEA and 300 μM 4-AP. (B) INa,TTX was continuously recorded under control conditions and in the presence of TTX. INa,TTX was blocked by TTX (left panel) and also blocked by verapamil (right panel). (C) The I-V relationships of Kir were obtained by ramp voltage induced currents against membrane potentials in bath solutions containing 5, 15, 30, 75, and 150 mM K+ as indicated (left panel). Reversal potentials from six patches were observed and plotted as a function of external [K+] concentrations (right panel). The dotted line represents the slope from the Nernst equation (slope, 58 mV/decade). Experimental values (O) were fitted by linear regression (slope, 66 mV/decade).

  • Fig. 4. Comparison of mRNA expression of ion channels between passages. (A) Kv1.1, heag1 (for IKDR), Kv4.2 (for Ito), Kir2.1 (for Kir), MaxiK (for IKCa) and hNE-Na (for INa.TTX) were detected in hBM-MSCs, but not SCN5A (for TTX-resistant INa). β -actin was used as the control. (B) The “recording rate (%)” of INa,TTX in various passage of hBM-MSCs. (C) Relative passage-dependent mRNA quantities of the hNE-Na gene by quantitative RT-PCR (n=3). The inset in Fig. 4C shows a representative result for the hNE-Na gene from two different samples. (D) Relative passage-dependent MaxiK gene expression levels in hBM-MSCs. Inset in Fig. 4D displays representative passage-dependent MaxiK channel gene expression patterns from two different samples. ∗p<0.05, ∗∗p<0.005 when compared with each 3rd passage group.


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