Decellularization Strategies for Regenerative Medicine: From Processing Techniques to Applications

Table 1

Summary of decellularization agents and techniques.

Category	Agents and techniques	Mechanism/description	Significant effects	References
Chemical and enzymatic	(1) Surfactants	Lyse cells by disarranging the phospholipid membrane
	Sodium dodecyl sulfate (SDS)	(i) Ionic	(i) Cytotoxic: requires extensive wash process	[45]
			(ii) Alters microstructure (i.e., collagen fibers)	[46, 47]
	Triton X-100	(i) Nonionic	(i) Less damaging to structure of tissue than ionic surfactants	[34, 48]
		(ii) Commonly used with ammonium hydroxide
	Sodium deoxycholate (SD)	(i) Ionic	(i) Causes agglutination of DNA when used without DNase	[49]
		(ii) Commonly used with DNase	(ii) Remnant DNA fragments	[45, 49–51]
	CHAPS	(i) Zwitterionic	(i) Maintains structural ECM proteins	[40]
			(ii) Remnant cytoplasmic proteins	[40]
			(iii) Maintains ultrastructure	[40, 41]
	(2) Acids and bases	Solubilize cell membrane by utilizing charged properties
	Peracetic acid	(i) Highly corrosive	(i) Insufficient cell removal	[45]
		(ii) Oftentimes used for sterilization	(ii) Increases stiffness of ECM	[45, 52]
	Ethylenediaminetetraacetic acid (EDTA)	(i) Commonly used with trypsin	(i) Decreases salt- and acid-soluble ECM proteins	[53]
	Reversible alkaline swelling	(i) Induces negative charge on collagen to cause swelling	(i) Alters mechanical properties	[48]
		(ii) Used with tridecyl alcohol ethoxylate
	(3) Enzymes	Typically used to supplement other chemical & mechanical treatments
	Trypsin	(i) Breaks cell-matrix adhesions
		(ii) Commonly used with EDTA
	Deoxyribonuclease (DNase)	(i) Breaks down DNA fragments
		(ii) Commonly used with SD
	Ribonuclease (RNase)	(i) Breaks down RNA fragments
Mechanical	High hydrostatic pressure (HHP)	(i) Pressures greater than 600 MPa applied to lyse cells	(i) Remnant DNA fragments	[23, 28]
			(ii) Denatures ECM proteins	[23]
	Supercritical carbon dioxide	(i) Applies CO2 at pressures above 7.40 MPa and temperatures above 31.1°C	(i) Requires entrainer to remove polar phospholipid membrane	[54]
			(ii) Maintains ECM proteins & mechanical properties	[54]
	Freeze-thaw	(i) Alternate between freezing temperatures (−80°C) and biological temperatures (~37°C)	(i) Maintains ECM proteins & mechanical properties	[55, 56]
			(ii) Remnant DNA	[55]