SKIN CARE
microscope. As shown in Figure 1, treatment with the
peptide increased MBNL1 protein amount in fibroblasts in a dose response manner and in a similar way to microcurrent stimulation. To further confirm the microcurrent-like
effect of the novel peptide, we evaluated the ability of the peptide to induce collagen contraction. HDFa cells were treated with 0.01 mg/mL of the peptide, and after 24 hours they were added to a collagen I hydrogel that included 0.01 mg/mL of the peptide. The collagen I hydrogel was left for 24 hours
to allow for contraction. Non-treated cells exposed to microcurrents stimulation were used as a positive control. Hydrogel contraction was quantified by the calculation of the surface area variation. The novel peptide induced collagen
contraction by 37.5% with respect to the control condition. Further, it showed similar efficacy when compared to microcurrent stimulation (Figure 2). Finally, we confirmed the appearance of
the myofibroblast phenotype modulation after treatment with the novel peptide in HDFa cells by checking EDA-fibronectin, a cell marker implicated in cell contraction and adhesion functions. Cells coming from the assay of collagen gel contraction in vitro were used to detect EDA-fibronectin. Briefly, 3D collagen gels were digested
by a collagenase treatment and then EDA- fibronectin levels were detected and evaluated in the cells by means of immunofluorescence and quantified by confocal microscopy. As shown in Figure 3, the novel peptide
induced the accumulation of EDA-fibronectin marker in fibroblasts by 72.3%, compared to control, thus confirming the modulation towards the myofibroblast phenotype. Further, the increase of EDA-fibronectin was greater than after microcurrent stimulation.
Mimicking the skin benefits of microcurrents in the muscle layer Microcurrent stimulation is known to improve energy production by mitochondria, which is believed to contribute to enhanced muscle tone, providing better skin support, and ultimately, a firmer facial appearance. In muscle cells, MBNL1
160 140 120 100 80 60 40 20 0
** 28.5 A
180 160 140 120 100 80 60 40 20 0
*** *** 37.5 B Control Electrical stimulation
0.01mg/mL Uplevity e-Lift peptide
21
Control
Electrical stimulation
Uplevity e-Lift peptide
0.001 mg/mL
Figure 2: A) Percentage of collagen gel contraction (***p<0.001, calculated using an unpaired Student´s t-test). B) Representative images showing collagen gel contraction after Uplevity e-Lift peptide treatment and after microcurrent stimulation (<1000mA, 70mv/mm, 1h)
A
200 180 160 140 120 100 80 60 40 20 0
Control **** ** 72.3 B Control Electrical stimulation
0.01mg/mL Uplevity e-Lift peptide
Electrical stimulation
Uplevity e-Lift peptide
0.001 mg/mL
Figure 3: A) Percentage of EDA-fibronectin levels (**p<0.01, ****p<0.0001, calculated using an unpaired Student´s t-test). B) Representative images of EDA-fibronectin marker (green) and nuclei (blue) after Uplevity e-Lift peptide treatment and after microcurrent stimulation (<1000mA, 70mv/mm, 1h)
protein has been described to be involved in mitochondria energization. Therefore, first we checked whether the novel
peptide could induce MBNL1 protein in muscle cells. To verify this, human skeletal muscle cells (hSkMC) were first allowed to differentiate into myocytes and myotubes. Then, they were treated with 0.5 mg/mL
of the peptide for 48 hours and total protein content was extracted. The levels of MBNL1 protein were measured by HTRF (homogeneous time-resolved Fluorescence). Our results demonstrated that the novel peptide treatment induced the accumulation of MBNL1 protein in
A
160 140 120 100 80 60 40 20 0
B ****
muscle cells by 28.5%, with respect to the control condition (Figure 4). With this, we then moved on to determine
whether the novel peptide was able to induce muscle energization. To do this, we checked the mitochondrial membrane potential, a direct indicator of ATP production by cells. For that, hSkMC cells were first allowed to differentiate into myocytes and myotubes. Then, they were treated with 0.1 mg/
mL or 0.5 mg/mL of the peptide for 24 h or stimulated with microcurrents (C-Dish device, IonOptix). Finally, the JC-1 fluorescent probe was added. Mitochondrial membrane potential
**** **** 27.5 49.3
Control
Electrical stimulation
0.1 mg/mL Uplevity e-Lift peptide
0.5 mg/mL Uplevity e-Lift peptide
Control
Uplevity e-Lift peptide
0.5 mg/mL
Figure 4: Percentage of MBNL1 levels in muscle cells (**p<0.01, calculated using an unpaired Student´s t-test)
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Control
Electrical stimulation
0.1 mg/mL Uplevity e-Lift peptide
0.5 mg/mL Uplevity e-Lift peptide
Figure 5: A) Percentage of mitochondrial membrane potential improvement (****p<0.0001, calculated using an unpaired Student´s t-test). B) Representative images of JC-1 signal (red) and nuclei (blue) after Uplevity e-Lift peptide treatment and after microcurrent stimulation (<1000mA, 70mV/mm, 15 min)
February 2024 PERSONAL CARE
MBNL1 levels (%)
EDA-Fibronectin levels (%)
Variation in MBNL1 levels (%)
Mitochondrial
membrane potential improvement (%)
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