Mature and Immature Corneocyte Detection Force Distance Curves vs. Microfluorometry

May 1, 2013 | Contact Author | By: Anthony J. Ribaudo
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Title: Mature and Immature Corneocyte Detection Force Distance Curves vs. Microfluorometry
atomic force microscopy (AFM)x epifluorescencex force-distance curves (F-D)x hysteresis loopx microfluorometryx scanning probe microscopy (SPM)x
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Keywords: atomic force microscopy (AFM) | epifluorescence | force-distance curves (F-D) | hysteresis loop | microfluorometry | scanning probe microscopy (SPM)

Abstract: Here, the author compares two methods to determine the maturity of corneocytes based on their cross-linking that could be used to evaluate the anti-aging effects of molecular agents. The first utilizes microfluorometry, while the second involves F-D curves generated via contact mode AFM. Both methods successfully detected differences in mature or immature corneocytes with 95% confidence.

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AJ Ribaudo, Mature and immature corneocyte detection: Force distance curves vs. microfluorometry, Cosm & Toil 128(5) 350-358 (May 2013)

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Corneocyte maturity plays a crucial role in dermatology and cosmetic science because immature corneocytes in the stratum corneum (SC) can cause impaired barrier properties. This poses a threat to overall health, since it is the outermost layers of the SC that protect the body from invasion by pathogens and viruses. Interestingly, studying corneocyte maturity has enabled dermatologists to determine there are no differences in corneocyte maturity across different racial groups. Further, a causal relationship has been found between excessive amounts of immature corneocytes and dry skin in all ethnic groups.

Since corneocytes act as indicators, they can be used to correlate the age and health of skin. Therefore, the techniques discussed here could be used to determine the efficacy of skin care products. For example, cosmetic scientists have demonstrated that the use of cosmetic moisturizers containing niacinamide and hexamidine help promote the development of mature corneocytes, in turn improving the barrier properties of skin. This enhancement of barrier properties is attributed to increased levels of tortuosity and better covalent bonding of the SC to intercellular lipids that protect skin and prevent water loss.

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This content is adapted from an article in GCI Magazine. The original version can be found here.

 

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Table 1. Instrument parameters for microfluorometry experiments

Table 1. Instrument parameters for microfluorometry experiments

Microfluorometry was performed using an instrument that operates in the incident mode in the visible range at a specific wavelength, as determined by the filter selected. The related instrument settings used for the study are shown here.

Table 2. Microfluorometry data in counts per second (cps)

Table 2. Microfluorometry data in counts per second (cps)

The microfluorometry results summarized here indicate that the mature corneocytes had a higher average fluorescence intensity (54) than the immature corneocytes (43).

Table 3. F-D data generated by contact mode AFM of mature vs. immature corneocytes

Table 3. F-D data generated by contact mode AFM of mature vs. immature corneocytes

To demonstrate the utility of the method, F-D curves were generated from five mature and five immature corneocytes. The data here shows that the immature corneocytes had a higher adhesive energy (21,612 Joules x10-18) than the mature corneocytes (3,331 Joules x10-18).

Figure 1. Schematic of the epifluorescence microscope

Figure 1. Schematic of the epifluorescence microscope

Schematic of the epifluorescence microscope showing the photomultiplier tube (PMT) detector

Figure 2. The tip of a typical F-D curve

Figure 2. The tip of a typical F-D curve

The tip approaches the surface (dotted black line) and makes contact at point A from the right side of the figure. At point D (red line) the tip experiences adhesive forces and finally breaks free at point B, creating a hysteresis loop. The slope of the deflection curve along the line segment BC gives the Young’s modulus of the sample.

Figure 3. Epifluorescent image of corneocytes stained with Nile red as seen under the epifluorescence microscope

Figure 3. Epifluorescent image of corneocytes stained with Nile red as seen under the epifluorescence microscope

Fluorescence intensity was determined utilizing a circuit composed of a voltmeter integrated with various electronic modules. The digital readout was obtained using a computer program developed in-house that displays the count rate continuously. A typical epifluorescent image of corneocytes stained with the Nile red is shown here.

Figure 4. A typical F-D hysteresis loop

Figure 4. A typical F-D hysteresis loop

A typical F-D hysteresis loop obtained from a mature corneocyte (top) compared to one obtained from an immature corneocyte (bottom)

Figure 5. Topographic (left) and deflection image (right) of a typical mature corneocyte

Figure 5. Topographic (left) and deflection image (right) of a typical mature corneocyte

The cross-linking in a typical mature corneocyte appears ropelike.

Figure 6. Topographic (left) and deflection image (right) of a typical immature corneocyte

Figure 6. Topographic (left) and deflection image (right) of a typical immature corneocyte

The smoother surface topography exhibited by an immature corneocyte

Footnotes [Ribaudo 128(5)]

a The Leitz MPV 1.1 with a Vertical Ploem Illuminator coupled to a fluorescence microscope was used for this study.

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