From hair to heart: nestin-expressing hair-follicle-associated pluripotent (HAP) stem cells differentiate to beating cardiac muscle cells

<p>We have previously demonstrated that the neural stem-cell marker nestin is expressed in hair follicle stem cells located in the bulge area which are termed hair-follicle-associated pluripotent (HAP) stem cells. HAP stem cells from mouse and human could form spheres in culture, termed hair spheres, which are keratin 15-negative and CD34-positive and could differentiate to neurons, glia, keratinocytes, smooth muscle cells, and melanocytes in vitro. Subsequently, we demonstrated that nestin-expressing stem cells could effect nerve and spinal cord regeneration in mouse models. In the present study, we demonstrated that HAP stem cells differentiated to beating cardiac muscle cells. We separated the mouse vibrissa hair follicle into 3 parts (upper, middle, and lower), and suspended each part separately in DMEM containing 10% FBS. All three parts of hair follicle differentiated to beating cardiac muscle cells as well as neurons, glial cells, keratinocytes and smooth muscle cells. The differentiation potential to cardiac muscle is greatest in the upper part of the follicle. The beat rate of the cardiac muscle cells was stimulated by isoproterenol and inhibited by propanolol. HAP stem cells have potential for regenerative medicine for heart disease as well as nerve and spinal cord repair.</p

Early-age-dependent selective decrease of differentiation potential of hair-follicle-associated pluripotent (HAP) stem cells to beating cardiac-muscle cells

<p>We have previously discovered nestin-expressing hair-follicle-associated pluripotent (HAP) stem cells and have shown that they can differentiate to neurons, glia, and many other cell types. HAP stem cells can be used for nerve and spinal cord repair. We have recently shown the HAP stem cells can differentiate to beating heart-muscle cells and tissue sheets of beating heart-muscle cells. In the present study, we determined the efficiency of HAP stem cells from mouse vibrissa hair follicles of various ages to differentiate to beating heart-muscle cells. We observed that the whiskers located near the ear were more efficient to differentiate to cardiac-muscle cells compared to whiskers located near the nose. Differentiation to cardiac-muscle cells from HAP stem cells in cultured whiskers in 4-week-old mice was significantly greater than in 10-, 20-, and 40-week-old mice. There was a strong decrease in differentiation potential of HAP stem cells to cardiac-muscle cells by 10 weeks of age. In contrast, the differentiation potential of HAP stem cells to other cell types did not decrease with age. The possibility of rejuvenation of HAP stem cells to differentiate at high efficiency to cardiac-muscle cells is discussed.</p

Isoproterenol directs hair follicle-associated pluripotent (HAP) stem cells to differentiate <i>in vitro</i> to cardiac muscle cells which can be induced to form beating heart-muscle tissue sheets

<p>Nestin-expressing hair-follicle-associated pluripotent (HAP) stem cells are located in the bulge area of the follicle. Previous studies have shown that HAP stem cells can differentiate to neurons, glia, keratinocytes, smooth muscle cells, and melanocytes in vitro. HAP stem cells effected nerve and spinal cord regeneration in mouse models. Recently, we demonstrated that HAP stem cells differentiated to beating cardiac muscle cells. The differentiation potential to cardiac muscle cells was greatest in the upper part of the follicle. The beat rate of the cardiac muscle cells was stimulated by isoproterenol. In the present study, we observed that isoproterenol directs HAP stem cells to differentiate to cardiac muscle cells in large numbers in culture compared to HAP stem cells not supplemented with isoproterenol. The addition of activin A, bone morphogenetic protein 4, and basic fibroblast growth factor, along with isoproternal, induced the cardiac muscle cells to form tissue sheets of beating heart muscle cells. These results demonstrate that HAP stem cells have great potential to form beating cardiac muscle cells in tissue sheets.</p

Time course of HCO₃⁻ transport in lMAL and sMAL.

<p>Vehicle was added to the bath after the collection durng the control period. HCO₃⁻ transport was stable with time both in lMALs and sMALs, although the PD was decreased only in sMALs.</p

Effects of AVP on HCO₃⁻ transport in lMAL and sMAL.

<p>The addition of 10⁻¹⁰ M AVP to the bath decreased HCO₃⁻ absorption both in lMALs and sMALs, but the PD increased only in lMALs.</p

Effects of AVP, ANP and cGMP on HCO₃⁻ transport in sMALs and lMALs.

<p>Values are mean ± SE. Abbreviations: L, tubular length; C, control period; E, experimental period; C′, recovery period; [T_CO2]_p, total CO₂ concentration in perfusate and bath; [T_CO2]_c, total CO₂ concentration in the collected solution; JTCO2, net bicarbonate absorption, NCR, normalized collection rate; PD, transepitherial potential difference;</p>*<p>p<0.05 vs. control period.</p

Expression levels of V1aR and V2R mRNA in sMALs and lMALs.

<p>The expression levels of V1aR in sMALs and lMALs were examined using real time PCR. sMALs and lMALs were dissected out of control rats after 30°C in a 0.1% collagenase solution in the presence of VRC. * p<0.05 vs. sMAL, n = 3–5.</p

Effect of ANP on HCO₃⁻ transport in the presence of AVP in sMALs.

<p>ANP at the concentration of 10⁻⁸ M stimulated HCO₃⁻ reabsorption in the presence of 10⁻¹⁰ M AVP in sMALs.</p

Effects of ANP and cGMP on HCO₃⁻ transport in the presence of AVP in lMALs.

<p>ANP at the concentration of 10⁻¹⁰ M and 10⁻⁸ M significantly increased HCO₃⁻ reabsorption in the presence of 10⁻¹⁰ M AVP in lMALs (A and B, respectively). cGMP at the concentration of 10⁻⁴ M mimicked the effect of ANP on HCO₃⁻ transport in the presence of 10⁻¹⁰ M AVP in lMALs (C).</p

Effect of ANP on HCO₃⁻ transport in lMALs in the absence of AVP.

<p>ANP at the concentration of 10⁻⁸ M did not cause changes in HCO₃⁻ absorption in lMALs in the absence of AVP.</p