Methods for the isolation and 3D culture of dermal papilla cells from human hair follicles

Abstract

The dermal papilla is a cluster of mesenchymal cells located at the base of the hair follicle which have a number of important roles in the regulation of hair growth. As a consequence, in vitro models of these cells are widely used to study the molecular mechanisms which underlie hair follicle induction, growth and maintenance. While dermal papilla from rodent hair follicles can be digested prior to cell isolation, the unique extracellular matrix composition found in human dermal papilla renders enzymes such as trypsin and collagenase insufficient for digestion of the dermal papilla into a single cell suspension. As such, to grow human dermal papilla cells in vitro, the papilla has to first be isolated via a micro-dissection approach from the follicle. In this article we describe the micro-dissection and culture methods, which we use within our laboratory, for the study of human dermal papilla cells.

Keywords: dermal papilla; hair follicle; inversion; micro-dissection; spheroid culture.

Figure 1

Human dermal papilla micro‐dissection method. (A) Skin biopsy prior to papilla isolation. (B) Petri dish lid with drops of DMEM for dissection. (C) Transection of the hair follicle with scissors. (D) A transected hair follicle end bulb. (E) Inversion of the end bulb. (F) Inverted end bulb and exposed dermal papilla. (G) Transection of the dermal papilla. (H) Dermal papilla adhered to a 35mm dish with a scratch. (I) Growth of cells in an starburst formation around a single dermal papilla.

Figure 2

Hanging drop method for spheroid formation. (A) A 20μl pipette placing 10 μl drops, each containing 3000 cells, on the inverted lid of a petri dish. (B) Approximately 60 drops placed on the inverted lid of a 10cm petri dish. (C) Petri dishes containing spheres stacked in an incubator. (D) Bright field images showing sphere formation in a single hanging drop.