A New Technique for Quantitative Analysis of Hair Loss in Mice Using Grayscale Analysis

Summary

Alopecia is a common form of hair loss which can occur in many different conditions, including as a side-effect of chemotherapy. We have developed a method to quantify hair loss in mice, utilizing a standard gel imager to perform a grayscale analysis, to facilitate study of promising new alopecia therapies.

Abstract

Alopecia is a common form of hair loss which can occur in many different conditions, including male-pattern hair loss, polycystic ovarian syndrome, and alopecia areata. Alopecia can also occur as a side effect of chemotherapy in cancer patients. In this study, our goal was to develop a consistent and reliable method to quantify hair loss in mice, which will allow investigators to accurately assess and compare new therapeutic approaches for these various forms of alopecia. The method utilizes a standard gel imager to obtain and process images of mice, measuring the light absorption, which occurs in rough proportion to the amount of black (or gray) hair on the mouse. Data that has been quantified in this fashion can then be analyzed using standard statistical techniques (i.e., ANOVA, T-test). This methodology was tested in mouse models of chemotherapy-induced alopecia, alopecia areata and alopecia from waxing. In this report, the detailed protocol is presented for performing these measurements, including validation data from C57BL/6 and C3H/HeJ strains of mice. This new technique offers a number of advantages, including relative simplicity of application, reliance on equipment which is readily available in most research laboratories, and applying an objective, quantitative assessment which is more robust than subjective evaluations. Improvements in quantification of hair growth in mice will improve study of alopecia models and facilitate evaluation of promising new therapies in preclinical studies.

Introduction

Alopecia (hair loss) can be a psychologically and emotionally distressing event with multiple causes. Male-pattern baldness is the most common cause of alopecia, affecting approximately two thirds of males by age 35¹. A similar pattern of hair loss can be observed in females with polycystic ovarian syndrome. In both of these disorders, the hair loss is androgen mediated. Alopecia can also occur as an autoimmune disease, alopecia areata, which affects 1.7% of the population².  Alopecia can occur as a side-effect of some medical treatments, such as chemotherapy³.  A high percentage (65-85%) of chemotherapy patients experience s....

Protocol

Ethics Statement: All studies involving animals must be approved by the IACUC of the institution (for the data which follows, protocols were approved by the Montefiore IACUC, protocol #11-6-240 and #13-7-100). Animals are provided light anesthesia as indicated for the sole purpose of keeping them still during photography, there are no painful procedures required for this protocol.

1. Acquisition of Photographs

1. Set focus and field of view for gel imager using paper with printed text. Verify uniformity of light source across photographed region. To help ensure uniformity, use a gel imager with a built in light source for reflectiv....

Results

This technique was validated using C3H/HeJ engrafted mice, the mouse model of alopecia areata¹⁶. These animals develop global hair loss which progresses gradually over time. However, the hair loss occurs in patches, and may vary from one mouse to the next, introducing considerable variability and hampering qualitative assessments. This model provided an opportunity to test correlations between measurements of optical density at different locations on the mouse as an assessment of validity.

Discussion

In this report a detailed description is provided of a new technique for quantifying hair loss in rodents. This technique utilizes a gel imager for image acquisition and analysis, equipment which is readily available in most laboratories. The measurements have been shown to be robust to minor variations in technique (Figures 4-5), and are well-correlated to the degree of visual hair loss in treatment models (Figure 6). These techniques have been successfully employed in .......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
KODAK Gel Logic 100 Imaging System	Eastman Kodak Company, Rochester, NY, USA		Gel imager for obtaining and analyzing photographs, must have built in light source for reflective photography.
The Kodak/Tiffen Q-13 Gray Scale	Imatest		Greyscale standard
C57BL/6J Mice	Jackson Laboratories, Bar Harbor, Maine		Mice for representative study
C3H/HeJ engrafted mouse	Jackson Laboratories, Bar Harbor, Maine		Mice for representative study