Matrix metalloproteinases (MMPs): MMPs are a group of calcium-dependent, zinc-containing enzymes. In different tissues, they have been identified 24 different MMPs, varying substrate specifications, and multiple functions [
58]. Besides, during microbial infection, the MMPs play an essential role by degrading the extracellular matrix products from different organs that exhibit antimicrobial activity against wound pathogens [
59]. Based on MMPs domain organization and substrate preference, these can be classified into the four most relevant groups: (1) collagenases, (2) gelatinases, (3) stromelysins, and (4) matrilysins. Collagenase: This group is comprised of enzymes MMP-1, MMP-8, and MMP-13, which are the enzymes in mammals with the capability to break down the triple helix of collagen [
60]. Such MMPs can also downgrade several other non-ECM and ECM molecules. Interstitial collagenase (MMP-1) breaks type II collagen and appears to have activity, especially with type III collagen. Polymorphonuclear collagenase (MMP-8) has the most significant activity against type I. MMP-13 has a unique intensive capability to break all three types of collagen (I, II, and III) [
61]. Gelatinases: MMP-9 (gelatinase B) and MMP-2 (gelatinase A) are the main enzymes that are upregulated in chronic wounds [
62]. Fibroblasts secrete MMP-2, while the larger molecule MMP-9 is mainly produced by leukocytes and, possibly, by keratinocytes. These enzymes play an essential role in the remodeling because they have additional fibronectin located inside the catalytic domain. One of the main functions of gelatinase A is to accelerate migration, while gelatinase B promotes cell migration and re-epithelialization [
63]. Stromelysins: This group is composed of three members: Stromelysin-1 (MMP-3), stromelysin-2 (MMP-10), and stromelysin-3 (MMP-11), which play a varied role in the degradation of the extracellular matrix [
64]. Stromelysins are expressed by epithelial and fibroblast cells and are secreted to the extracellular space, where they play essential roles in biological processes such as mammary gland development, immunity, and wound healing [
65]. Matrilysins: During the process of tissue remodeling, MMP-7, also known as matrilysins-1, is believed to degrade components of the extracellular matrix (ECM) such as laminin, entactin, and type IV collagen [
66]. Additionally, in humans, MMP7 expression is observed in IPF lung tissue but not healthy control samples. It is also detectable in BAL fluid, where levels are increased in patients with IPF and inversely correlated with FVC [
67]. Serine proteases: Serine proteases are proteins with abundant sources distributed among all living cells and are important enzymes because some of them hydrolyze peptide bonds [
68]. These proteins contain serine residues in their active catalytic center, which has a molecular mechanism similar to esterase. Serine protease derives its name from the presence of residual nucleophilic serine in the active site that attacks the carbonyl components of the substrate [
69].
The enzymatic activities of serine proteases are tightly regulated within translation transcription, zymogen activation, autolysis, and interaction with natural inhibitors. Thrombin, is one of the most noticeable members of the serine protease family, is a 36
-kDa protein comprised of two chains, A and B, linked by a disulfide bond [
70]. Grouping peptidases classify proteolytic enzymes based on sequencing similarities and structure into families and clans based on catalytic mechanism, PA Clan proteases and the E*form, and homology [
71].
Dead tissues present in a wound site serve as reservoirs for the development of bacteria and contain high levels of inflammatory mediators, which promote a continuous case of inflammation and reduce cellular migration that is necessary for wound regeneration. Proper wound cleaning and debridement are elemental for granulation, followed by re-epithelization. Among the known debridement methods, enzymatic debridement is a highly efficient method that uses proteolytic enzymes naturally present in the body.
While proteases are not recommended for use in delivery routes such as oral, due to their susceptibility to inhibitors, ease of denaturing, and need to remain long enough in the site of action to achieve positive pharmacokinetics, they have been successfully studied and applied via the topical route. The most frequently used proteases for topical wound healing applications include collagenases, cysteine proteases, and serine proteases, although animal secretions such as snake venom,
Lucilia sericata secretions, and fish epithelial mucus, all of which include several enzymatic and non-enzymatic proteins (proteases being among these) have been demonstrated to have good results [
76,
77,
78].