Name: precision implementation of minimal erythema dose (med) testing to assess individual variation in human inflammatory response
Uploaded: 2019-10-03T13:30:00
Description: Watch this Scientific Journal Video about Precision Implementation of Minimal Erythema Dose MED Testing to Assess Individual Variation in Human Inflammatory Response at JoVE.com

This work was supported by a grant from the Virginia Tech College of Science Discovery Fund.

All methods described below including the use of human volunteers have been reviewed and approved by the local Institutional Review Board (IRB), and are in accordance with the Declaration of Helsinki and Belmont Report. All participants (N=72) signed informed consent as proscribed by the IRB protocol. Inclusion/exclusion criteria and discontinuation procedures were designed to maximize participant safety, and any deviation from these procedures should be considered in light of their impact on risk and tolerability to hum...

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Precision implementation of MED testing as described here could offer several advantages over other extant lab-based inflammatory challenges that have achieved popular use. For example, suction blister protocols17,18,19 raise a fluid-filled blister on the skin that is subsequently aspirated with a syringe to gain direct access to the cytokine microenvironment. Although skin blistering is a well-known tool for studying skin immun...

Minimal erythema dose (MED) testing is an FDA-approved procedure to evaluate cutaneous sensitivity to radiation typically in the UVB range, although the MED can be determined at other wavelengths in the UV and visible spectrum1. Erythema is defined as superficial reddening on the surface of the skin caused by engorgement of capillaries (later stages of erythema are more commonly known as sunburn). MED testing has been used extensively in the dermatology literature and clinical phototherapy settings to identify the minimal amount of ultraviolet (UV) radiation that will produce the smallest unit of measurable change in the redness of the skin. MED testing can be accomplished with a commercially available UV lamp, equivalent to what is used in most commercial tanning facilities.
MED testing involves continuous dispersal of UV radiation or light from the visible spectrum onto the surface of the skin for a predetermined length of time, with dosage schedules depending primarily on pigmentation of the skin and the intensity and type of radiation. This procedure is commonly used in clinical settings to determine dosage schedules for patients receiving UV radiation therapy for skin conditions such as psoriasis and eczema2,3. Basic procedures for determining the MED in clinical settings have been described elsewhere4, and can be used to adjust the total dosage of UV radiation upward or downward, depending on individual variation in skin sensitivity.
Skin pigmentation is perhaps the most important subject-specific variable in conducting and measuring results from the MED procedure6. This is because the duration of UV exposure required to evoke the minimal erythema response is principally determined by the lightness or darkness of the participant&#8217;s skin, as defined by the participant&#8217;s Fitzpatrick skin type (FST). FST7 is a numerical scheme for classifying human skin color. The Fitzpatrick scale is a recognized tool for dermatological research into human skin pigmentation8,9, and classifies human skin into one of six categories from lightest (FST I) to darkest (FST VI).
Darker FST typologies require longer UV duration, therefore accurate classification of FST is important. There is an extensive literature on methods for accurate assessment of FST, using a wide variety of approaches including self-report, dermatologist interview and instrumentation-based assessment. Observer ratings of FST have been shown to be correlated with current, but not natural skin color10, however FST can be determined subjectively11 using self-report via questionnaire12 and/or objective assessment via spectrophotometry. Fitzpatrick typing by spectrophotometry has been shown to correlate closely to participant self-report in a number of studies10,13,14,15.
Despite the utility and widespread use of MED testing in clinical services, this procedure has not been widely adopted in laboratory settings for measurement of individual variation in response to pro-inflammatory stimulation. The purpose of the methodology outlined here is to provide techniques and step-by-step procedures that increase the precision and reproducibility of the MED testing procedure, in order to facilitate future work in laboratory settings focused on fine-grained quantification of intra-individual variability in inflammatory response. We further provide representative results that illustrate the capability of this standardized protocol to accurately capture person-to-person variation in inflammation.

<table>
	<tbody>
		<tr>
			<td>6-aperture dose testing patch (&#8220;Cuff&#8221;)</td>
			<td>Daavlin</td>
			<td>&#160;</td>
			<td>&#160;</td>
		</tr>
		<tr>
			<td>Medical grade adhesive solvent</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Non-reflective UV proof cloth</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Radiometer</td>
			<td>SolarLight</td>
			<td>Model 6.2 UVB Meter</td>
			<td></td>
		</tr>
		<tr>
			<td>Single use aloe or burn gel</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Spectrophotometer</td>
			<td>Konika-Minolta</td>
			<td>CM-2600D</td>
			<td></td>
		</tr>
		<tr>
			<td>Stopwatch</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>UV lamp &#8211; Fiji Sun</td>
			<td>Sperti</td>
			<td></td>
			<td>Emission spectrum 280nm-400nm, approximately 25% UVB</td>
		</tr>
		<tr>
			<td>UV-proof safety glasses (2 pair)</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>UV-proof sleeve</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>White cotton gloves (2 pair)</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

precision implementation of minimal erythema dose (med) testing to assess individual variation in human inflammatory response

Minimal erythema dose (MED) testing is frequently used in clinical settings for determining the smallest amount of ultraviolet (UV) irradiation necessary to produce erythema (inflammatory reddening) on the surface of the skin. In this context, the MED is regarded as a key factor in determining starting doses for UV phototherapy for common skin conditions such as psoriasis and eczema. In research settings, MED testing also has potential to be a powerful tool for assessing within- and between-persons variation in inflammatory responses. However, MED testing has not been widely adopted for use in research settings, likely owing to a lack of published guidelines, which is a barrier to obtaining reproducible results from this assay. Also, protocols and equipment for establishing MED vary widely, making it difficult to compare results across laboratories. Here, we describe a precise and reproducible method to induce and measure superficial erythema using newly designed protocols and methods that can easily be adapted to other equipment and laboratory environments. The method described here includes detail on procedures that will allow extrapolation of a standardized dosage schedule to other equipment so that this protocol can be adapted to any UV radiation source.

The timing schedule presented in Table 1 is a novel dosage schedule that was calculated to capture the MED, on average, at the mid-point of the exposure event (i.e., aperture 3 or 4) for each FST. The basis for the calculated schedule is as follows.
Prior work has established that for individuals with FST 2, the median MED for radiation in the UVB range is 66.9 milliwatts (mW) per cm2, 77.429 mW/cm2 for FST 3 and 85.0 for FST 416. ...

Watch this Scientific Journal Video about Precision Implementation of Minimal Erythema Dose MED Testing to Assess Individual Variation in Human Inflammatory Response at JoVE.com

Precision Implementation of Minimal Erythema Dose MED Testing to Assess Individual Variation in Human Inflammatory Response

Minimal erythema dose (MED) testing is used to establish dosage schedules for ultraviolet radiation phototherapy. It can assess individual variation in inflammatory response but lacks methodology for achieving reproducible results. Here, we present a precision implementation of MED and demonstrate its ability to capture individual variation in inflammatory response.

Precision Implementation of Minimal Erythema Dose (MED) Testing to Assess Individual Variation in Human Inflammatory Response

Minimal erythema dose (MED) testing is frequently used in clinical settings for determining the smallest amount of ultraviolet (UV) irradiation necessary ...

Minimal erythema dose testing, or MED testing, is a technique with a long history in dermatological and clinical phototherapy settings. And it's primarily used to determine the smallest amount of radiation required to produce erythema, or visible reddening on the surface of the skin. In this particular study, what we've done is develop several new techniques and procedures that allow MED testing to be adapted for somewhat different purpose, which is to precisely quantify variation in human inflammatory activation.The main advantage of precision dosing and measurement in MED testing pertains to the reproducibility of results, which then opens up new possibilities for systematically examining other factors that could be related to inflammation. Demonstrating the procedure will be two PhD students from my lab, Holly Sullivan-Toole and Corinne Carlton. Begin by escorting the participant to the testing room.Explain the MED testing procedure to the participant and confirm understanding. Place cuff one on the non-dominant forearm. Remove only the protective wax paper backing from the lateral, not central, portions of the cuff.After situating cuff one at the intended exposure site, place landmarks using a permanent marker to ensure that cuff two will be situated at precisely the same location. Then, mark the skin at four points outside of the creases of each of the side flaps of cuff one, the upper right, upper left, lower right, and lower left points. Ensure the marks are dark enough to last for 24 hours, so they may be used to place cuff three in precisely the same location at the follow-up appointment.Next, use a calibrated spectrophotometer to record readings at each of the six open apertures in sequence. Ensure that the spectrophotometer is placed in the center of the cuff apertures, while avoiding moles, scars, or other blemishes to the extent possible. Finally, permanently record all SCI values.To ensure consistent readings with the same calibration point, keep the spectrophotometer in the on position for the duration of the MED procedure and do not turn off until the post-exposure readings have been completed. After removal of cuff one, situate cuff two at the same location using the landmarks drawn on the skin for cuff one. Put on the necessary safety equipment, including UV-proof glasses, sleeves, and gloves, and make sure the participant does the same.Prior to activating the lamp, arrange the participant's arm such that the UV rays from the lamp will be perpendicular to the angle of cuff two on the participant's arm. Briefly activate the lamp to adjust the distance to cuff two until the radiometer sensor reads 270 microwatts per centimeter squared. Next, place the radiometer sensor facing the UV lamp, parallel to the surface of the skin, and as close as possible to the location of cuff two.Then, remove the first aperture covering before activating the UV source. Remove each window in the dose testing patch according to the schedule displayed here, which illustrates separate dosage schedules depending on the participant's Fitzpatrick Skin Type. Activate the UV source and the stopwatch simultaneously, and remove each aperture covering on cuff two according to the correct dosage schedule, which is based on Fitzpatrick Skin Type.Next, turn off the lamp at the precise time specified by the dosage schedule. Do not deactivate the stopwatch, as an additional series of spectrophotometer readings should be gathered exactly seven minutes subsequent to deactivation of the lamp, as described below. Lastly, after exactly seven minutes have passed since the deactivation of the lamp, record the final spectrophotometer readings from each aperture in cuff two.Begin by calibrating the spectrophotometer in preparation for the follow-up appointment, 24 hours later. Prepare cuff three by removing all of the aperture coverings and leaving the white wax paper backing on the central portion of the patch. When placing cuff three on the participant's arm, remove the white wax paper backing from the two side flaps of the patch.Use the landmarks on the participant's forearm to place cuff three in the same location as the previous two patches. Finally, take a reading at each of the six open apertures in sequence. These results show an accurate skin response to MED testing.Here are complete data for a single representative subject, broken down by assessment period. Pre-exposure, seven minutes post exposure, and 24 hours follow-up. Each aperture, two through six, receives 25%more energy than the one before it.Further, all 72 subjects are shown, indicating the variation that are likely to be encountered, across different FSTs. Perhaps the most important aspects of precision MED testing relate to the adherence to the proper dosage schedule for the identified Fitzpatrick Skin Type and also consistently maintaining the correct distance between the surface of the skin and the UV light source. Part of our motivation in developing these techniques is to facilitate future research examining how inflammatory responses could relate to any number of potentially interesting variables, such as psychological, pharmacological, or health-related factors.

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Minimal Erythema Dose (MED) Testing

Minimal Erythema Dose MED Testing

Ultraviolet radiation (UV) therapy is  sometimes used as a treatment for various common skin conditions, including  psoriasis, acne, and eczema. The ...

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