Wrinkles are an outward sign of cutaneous aging appearing preferentially on ultraviolet B (UVB)-exposed areas. The anti-wrinkle effects of herbal extracts were investigated in an animal model. Female albino hairless mice (HR/ICR) were randomly allocated to the control group (non-irradiated vehicle), positive control group (UVB irradiated-vehicle), and two herbal extract mixture groups (HE-1 and HE-2). HE-1 included Glycyrrhizae radix, Rhei Rhizoma, Cornus officinalis, and Sesami semeni, and HE-2 included Swertia pseudo-chinensis, Sophora flavescens, Scutellaria baicalensis, and Salvia miltiorrhiza. The herbal extract mixtures were pre-treated dorsally with 0.2 ml per individual five times per week for four weeks prior to the start of UVB irradiation. At the fifth week, the animals were exposed to UVB irradiation for a subsequent eight weeks, three times per week. The intensity of irradiation showed a gradual increase, from 30 mJ/cm 2 to 240 mJ/cm2 (1 MED: 60 mJ/cm2 ). Dorsal skin samples were stained with H&E in order to examine the epidermal thickness. In addition, Masson-Trichrome staining was performed for determination of the amount of collagen fiber. Treatments with HE-1&2 resulted in an increase in the amount of collagen fiber, a better appearance, and fewer wrinkles, compared with the positive control. As determined by hydroxyproline assay, treatments with HE-1&2 led to a significant increase in the amount of collagen, compared with the positive control group (p<0.05). Chronic UVB irradiation to skin of hairless mice resulted in an increase in expression of matrix metalloproteinase-1 (MMP-1), however, treatments with HE-1&2 tended to decrease the expression of MMP-1. These results indicate that the herbal extracts used in this study have a preventive effect on UVB-induced wrinkle formation in a hairless mouse model, due in part to inhibition of MMP-1 expression and increment of collagen amount.

Human embryonic stem cells (hESCs) are promising cell source because of their unique self-renewal and pluripotency. Although hESC-derived cardiac cells are currently generated worldwide, cryopreservation of these cells is still limited due to low rate of post-thaw survival. Cryopreservation of hESC-derived cardiac cells is critical in that their long-term storage can accelerate their use in regenerative medicine. However, to date, there are few reports on efficient cryopreservation and post-thaw survival of hESC-derived cardiac cells. In this study, we evaluated the effects of ginsenoside, which is known to improve survival of rat embryonic cardiomyocytes against myocardial ischemia injury in diabetic rats (Wu et al., 2011), on the survival of hESC-derived cardiac cells after thawing. We induced differentiation into cardiac cells using our previously reported method (Kim et al., 2011). Differentiated, pre-beating stage cardiac cells were cryopreserved using either mass cryopreservation or vitrification. To evaluate the effects of ginsenoside (Re, Rb), we compared three sets: pre- and post-thaw treatment, pre- or post-thaw treatment only. The survival of post-thaw cardiac cells were evaluated using Trypan-blue and Annexin V staining. In addition, the three groups were treated with ROCK inhibitor Y-27632, and compared with non-treatment groups. The effect of ginsenoside was significant in post-thaw treatment group, i.e, thawed cells expressed cardiac specific genes and showed specific functionality such as spontaneous beating. Taken together, we demonstrated favorable effects of ginsenoside on the survival of hESC-derived cardiac cells after cryopreservation and thawing. These results suggest a possible application of well-known cardioprotectant ginsenoside in cell-based tissue engineering using hESC-derived cardiac cells.