Granzyme B is elevated in autoimmune blistering diseases and cleaves key anchoring proteins of the dermal-epidermal junction

Abstract

In healthy skin, epidermis and dermis are anchored together at the dermal-epidermal junction (DEJ), a specialized basement membrane pivotal for skin integrity and function. However, increased inflammation in the DEJ is associated with the disruption and separation of this junction and sub-epidermal blistering. Granzyme B (GzmB) is a serine protease secreted by immune cells. Dysregulated inflammation may lead to increased GzmB accumulation and proteolysis in the extracellular milieu. Although elevated GzmB is observed at the level of the DEJ in inflammatory and blistering skin conditions, the present study is the first to explore GzmB in the context of DEJ degradation in autoimmune sub-epidermal blistering. In the present study, GzmB induced separation of the DEJ in healthy human skin. Subsequently, α6/β4 integrin, collagen VII, and collagen XVII were identified as extracellular substrates for GzmB through western blot, and specific cleavage sites were identified by mass spectrometry. In human bullous pemphigoid, dermatitis herpetiformis, and epidermolysis bullosa acquisita, GzmB was elevated at the DEJ when compared to healthy samples, while α6/β4 integrin, collagen VII, and collagen XVII were reduced or absent in the area of blistering. In summary, our results suggest that regardless of the initial causation of sub-epidermal blistering, GzmB activity is a common final pathway that could be amenable to a single targeted treatment approach.

Retinal Distribution and Extracellular Activity of Granzyme B: A Serine Protease That Degrades Retinal Pigment Epithelial Tight Junctions and Extracellular Matrix Proteins

“…Densitometric analysis revealed that the cleaved bands for FN and LAM-5 reached significance (p < 0.05), while that of COL-IV did not (p > 0.05) (Figures 8D-F). There were no cleaved bands observed for COL-I ( Supplementary Figure 2), consistent with the earlier finding that COL-1 is not a substrate of GzmB (15,16).…”

“…BM is an important outer retinal ECM that regulates the exchange between the (1) metabolically active combination of photoreceptor and RPE and (2) the choriocapillaris blood supply. Several of the ECM proteins within BM are known substrates for extracellular GzmB activity, including fibronectin (FN), vitronectin (VN), and laminin (LAM) and a small subset of collagens (COL) (1,(15)(16)(17)(18) (Figure 1). The remodeling of BM during aging and AMD is known to also affect RPE cell adhesion and function, which in turn, compromises oBRB function (18,19).…”

Collagen XVII Processing and Blistering Skin Diseases

“…Acta Dermato-Venereologica 105 Collagen XVII processing and blistering skin diseases Theme issue: Blistering skin disorders in infiltrated cells in BP lesional skin and that not only does this enzyme cleave COL17, but it also cleaves other molecules present at the DEJ, including α6/β4 integrins and collagen VII (29).…”

Proteomics Reveals Extracellular Matrix Injury in the Glomeruli and Tubulointerstitium of Kidney Allografts with Early Antibody-Mediated Rejection

“…Cathepsins and caspases can cleave ECM and the cytoskeleton and thus relate to our proteomics findings in AMR, [41][42][43] whereas granzymes are mainly expressed by immune cells. 44,45 We next used MEROPS 46 to identify proteases predicted to cleave the ECM proteins in each AMR compartment. CTSL, CTSS, and CTSV were the top 3 proteases expressed in the kidney and predicted to cleave the highest number of ECM proteins dysregulated in the AMR glomeruli ( Fig.6C).…”