Potential of Systemic Allogeneic Mesenchymal Stromal Cell Therapy for Children with Recessive Dystrophic Epidermolysis Bullosa

Hereditary epidermolysis bullosa (EB) is associated with skin blistering and the development of chronic non-healing wounds. Although clinical studies have shown that cell-based therapies improve wound healing, recruitment of therapeutic cells to blistering skin and to more advanced skin lesions remains a challenge. Here, we analyzed cytokines and chemokines in blister fluids (BF) of patients affected by dystrophic, junctional and simplex EB. Our analysis revealed high levels of CXCR1, CXCR2, CCR2 and CCR4 ligands, particularly dominant in dystrophic and junctional EB. In vitro migration assays demonstrated preferential recruitment of CCR4+ lymphocytes and CXCR1+, CXCR2+ and CCR2+ myeloid cells toward EB-derived BF. Immuno-phenotyping of skin-infiltrating leukocytes confirmed substantial infiltration of the EB-affected skin with resting (CD45RA+) and activated (CD45RO+) T cells and CXCR2+ CD11b+ cells, many of which were identified as CD16b+ neutrophils. Our studies also showed that abundance of CXCR2 ligand in BF also creates a favorable milieu for the recruitment of the CXCR2+ stem cells, as validated by in vitro and in-matrix migration assays. Collectively, this study identified several chemotactic pathways that control the recruitment of leukocytes to the EB-associated skin lesions. These chemotactic axes could be explored for the refinement of the cutaneous homing of the therapeutic stem cells.

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Pro-Inflammatory Chemokines and Cytokines Dominate the Blister Fluid Molecular Signature in Patients with Epidermolysis Bullosa and Affect Leukocyte and Stem Cell Migration

Alexeev

Salas‐Alanís

Palisson

et al. 2017

“…In addition, it has become evident that insufficient knowledge about collagen VII biochemistry and pathology delays development of therapies. Specifically, better understanding of the physiological assembly and turnover of collagen VII and of molecular disease mechanisms in DEB is needed Petrof et al, 2015;Watt et al, 2015). In the process of developing therapeutic approaches to replace a defective or missing protein using a vehicle and a therapeutic agent with limited life, be it nonintegrative gene therapy, cell therapy, or protein therapy, it is essential to know the natural turnover of the protein-that is, collagen VII in case of DEB.…”

Section: Introductionmentioning

confidence: 99%

Collagen VII Half-Life at the Dermal-Epidermal Junction Zone: Implications for Mechanisms and Therapy of Genodermatoses

Kühl

Mezger

Haußer

et al. 2016

The tissue half-life of proteins largely determines treatment frequency of non-gene-editing-based therapies targeting the cause of genodermatoses. Surprisingly, such knowledge is missing for a vast number of proteins involved in pathologies. The dermal-epidermal junction zone is believed to be a rather static structure, but to our knowledge no detailed analysis of the stability of proteins within this zone has been performed. Here, we addressed the in vivo half-life of collagen type VII using genetic ablation of its expression and therapeutic introduction of exogenous collagen VII in a preclinical model. A similar in vivo stability of collagen VII was observed in the skin, tongue, and esophagus, with a half-life of about 1 month. Collagen VII expressed by intradermally injected mesenchymal stromal cells also exhibited a similar half-life. Our study provides key information needed for the development of protein replacement or cell-based therapies for dystrophic epidermolysis bullosa caused by genetic deficiency of collagen VII. Moreover, by showing what we define as an intermediate half-life of collagen VII, our study challenges the view of the dermal-epidermal junction zone as a static structure with very slow turnover.

“…Panel a reprinted with permission from Elsevier (Petrof et al, 2015). ( b ) Confocal image of the epidermal-dermal junction in noncorrected RDEB cells, in which collagen VIIA is not detected.…”

Section: Figurementioning

confidence: 99%

Research Techniques Made Simple: The Application of CRISPR-Cas9 and Genome Editing in Investigative Dermatology

Guitart

Johnson

2016

Designer nucleases have gained widespread attention for their ability to precisely modify genomic DNA in a programmable manner. These genome-editing nucleases make double-stranded breaks at specified loci, and desired changes can be made to modify, ablate, or excise target genes. This technology has been used widely to develop human disease models in laboratory animals and to study gene functions by silencing, activating, or modifying them. Furthermore, the recent discovery of a bacterially derived programmable nuclease termed clustered regularly interspaced palindromic repeats (CRISPR)-associated protein 9 (Cas9) has revolutionized the field because of its versatility and wide applicability. In this article, we discuss various modalities used to achieve genome editing with an emphasis on CRISPR-Cas9. We discuss genome-editing strategies to either repair or ablate target genes, with emphasis on their applications for investigating dermatological diseases. Additionally, we highlight preclinical studies showing the potential of genome editing as a therapy for congenital blistering diseases and as an antimicrobial agent, and we discuss limitations and future directions of this technology.