Cystatin <scp>M/E</scp> knockdown by lentiviral delivery of sh<scp>RNA</scp> impairs epidermal morphogenesis of human skin equivalents

Jansen, Patrick A.; Bogaard, Ellen H. van den; Kersten, Ferry F.J.; Oostendorp, Corien; Vlijmen-Willems, Ivonne M.J.J. van; Oji, Vinzenz; Traupe, Heiko; Hennies, Hans Christian; Schalkwijk, Joost; Zeeuwen, Patrick L.J.M.

doi:10.1111/exd.12022

Cited by 9 publications

(11 citation statements)

References 21 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We have previously described the use of in vitro human skin equivalents to study skin biology and inflammation [20,21,[24][25][26][27] . We here took a transgenic approach to selectively overexpress hBD2 or hBD3, using lentiviral gene delivery in primary keratinocytes as we have described previously [25] .…”

Section: Effect Of Lentiviral Delivery Of Hbd2 and Hbd3 On 3-dimensiomentioning

confidence: 99%

See 1 more Smart Citation

The Effects of Human Beta-Defensins on Skin Cells in vitro

Kilsdonk

Jansen²,

Bogaard³

et al. 2017

Dermatology

Self Cite

View full text Add to dashboard Cite

Background: Defensins are antimicrobial peptides that exert immunomodulatory and chemotactic functions. Based on these properties and their high expression levels in the skin, they are likely to affect skin inflammation, infection, and wound healing. This may lead to therapeutic applications in (burn) wound healing. Objective: We aimed to investigate the effects of human β-defensins (hBDs) on keratinocytes and fibroblasts, 2 major skin cell types involved in skin regeneration. Methods: Monolayer keratinocyte and fibroblast cultures were exposed to recombinant hBDs, and we overexpressed hBD2 and hBD3 in keratinocytes of reconstructed epidermal equivalents by lentiviral transduction. The effects were measured by immunohistochemistry, quantitative real-time PCR, and migration assays. Kinome analyses were performed on cultured keratinocytes to investigate the signal transduction events elicited by hBD stimulation. Results: We found that hBD3 induced the expression of cytokines and chemokines in keratinocytes, which was not observed in fibroblasts. hBD2, however, stimulated cell migration only in fibroblasts, which was not found for hBD3. Both defensins are likely to exert receptor-mediated effects in keratinocytes, as witnessed by changes in protein kinase activation following stimulation by hBD2 and hBD3. Kinome analysis suggested that protein kinase C activation was a common event for both defensins. We observed, however, considerable differences in keratinocyte responses between stimulation by exogenous recombinant defensins and endogenous defensins expressed following lentiviral transduction. Conclusion: Defensins exert modest biological effects on skin cells that are potentially beneficial in wound healing, but many questions regarding the biological mechanisms of action and relevance for the in vivo situation are still remaining.

show abstract

Section: Effect Of Lentiviral Delivery Of Hbd2 and Hbd3 On 3-dimensiomentioning

confidence: 99%

“…We here took a transgenic approach to selectively overexpress hBD2 or hBD3, using lentiviral gene delivery in primary keratinocytes as we have described previously [25] . We used the involucrin promoter to drive hBD expression only in the stratum granulosum of the epidermis.…”

Section: Effect Of Lentiviral Delivery Of Hbd2 and Hbd3 On 3-dimensiomentioning

confidence: 99%

The Effects of Human Beta-Defensins on Skin Cells in vitro

Kilsdonk

Jansen²,

Bogaard³

et al. 2017

Dermatology

Self Cite

View full text Add to dashboard Cite

show abstract

“…Immunohistochemical analysis was performed as described earlier . Tissues were fixed in 4% paraformaldehyde and embedded in paraffin.…”

Section: Methodsmentioning

confidence: 99%

Characterization of pigmented dermo‐epidermal skin substitutes in a long‐term in vivo assay

Böttcher‐Haberzeth

Biedermann

Klar

et al. 2015

Experimental Dermatology

View full text Add to dashboard Cite

In our laboratory, we have been using human pigmented dermo-epidermal skin substitutes for short-term experiments since several years. Little is known, however, about the long-term biology of such constructs after transplantation. We constructed human, melanocyte-containing dermo-epidermal skin substitutes of different (light and dark) pigmentation types and studied them in a long-term animal experiment. Developmental and maturational stages of the epidermal and dermal compartment as well as signs of homoeostasis were analysed 15 weeks after transplantation. Keratinocytes, melanocytes and fibroblasts from human skin biopsies were isolated and assembled into dermo-epidermal skin substitutes. These were transplanted onto immuno-incompetent rats and investigated 15 weeks after transplantation. Chromameter evaluation showed a consistent skin colour between 3 and 4 months after transplantation. Melanocytes resided in the epidermal basal layer in physiological numbers and melanin accumulated in keratinocytes in a supranuclear position. Skin substitutes showed a mature epidermis in a homoeostatic state and the presence of dermal components such as Fibrillin and Tropoelastin suggested advanced maturation. Overall, pigmented dermo-epidermal skin substitutes show a promising development towards achieving near-normal skin characteristics and epidermal and dermal tissue homoeostasis. In particular, melanocytes function correctly over several months whilst remaining in a physiological, epidermal position and yield a pigmentation resembling original donor skin colour.

show abstract

“…The critical cystatin E/M target seems to be cathepsin L/V, as simultaneous ablation of cathepsin L rescued cystatin E/M deficient mice lethality (Zeeuwen et al, 2010). Based on the deletion of cystatin E/M in a reconstructed human skin model, which showed an impaired epidermis development instead of ichthyosis-like features, it was suggested that the inhibitor might be critical also for humans (Jansen et al, 2012). Moreover, the transglutaminase activity is also dependent on release of cathepsins from lysosomes and Ca 2+ from endoplasmic reticulum and thereby on cystatin E/M cathepsin L/V axis.…”

Section: Cornificationmentioning

confidence: 99%

“…However, calpain-1-dependent LMP, which is triggered after tight envelope formation to prevent uncontrolled degradation, occurs from unknown reason Eckhart et al, 2013;Costanzo et al, 2015). Anyhow, dysregulation of proteases and their inhibitors in the skin might be related to development of various diseases, such as Netherton syndrome, Papillon-Lefevre syndrome as well as impaired skin barrier function (Zeeuwen, 2004;Cheng et al, 2009;Zeeuwen et al, 2009;Jansen et al, 2012).…”

Section: Cornificationmentioning

confidence: 99%

Lysosomes in programmed cell death pathways: from initiators to amplifiers

Kavčič

Pegan

Turk

2016

Biological Chemistry

View full text Add to dashboard Cite

Lysosome is the central organelle for intracellular degradation of biological macromolecules and organelles. The material destined for degradation enters the lysosomes primarily via endocytosis, autophagy and phagocytosis, and is degraded through the concerted action of more than 50 lysosomal hydrolases. However, lysosomes are also linked with numerous other processes, including cell death, inflammasome activation and immune response, as well as with lysosomal secretion and cholesterol recycling. Among them programmed cell death pathways including apoptosis have received major attention. In most of these pathways, cell death was accompanied by lysosomal membrane permeabilization and release of lysosomal constituents with an involvement of lysosomal hydrolases, including the cathepsins. However, it is less clear, whether lysosomal membrane permeabilization is really critical for the initiation of cell death programme(s). Therefore, the role of lysosomal membrane permeabilization in various programmed cell death pathways is reviewed, as well as the mechanisms leading to it.

show abstract

Cystatin M/E knockdown by lentiviral delivery of shRNA impairs epidermal morphogenesis of human skin equivalents

Cited by 9 publications

References 21 publications

The Effects of Human Beta-Defensins on Skin Cells in vitro

The Effects of Human Beta-Defensins on Skin Cells in vitro

Characterization of pigmented dermo‐epidermal skin substitutes in a long‐term in vivo assay

Lysosomes in programmed cell death pathways: from initiators to amplifiers

Contact Info

Product

Resources

About