scholarly journals Novel Insights Into Cellular Changes in HPV8-E7 Positive Keratinocytes: A Transcriptomic and Proteomic Analysis

2021 ◽  
Vol 12 ◽  
Author(s):  
Matthias Kirschberg ◽  
Adnan Shahzad Syed ◽  
Hanife Güler Dönmez ◽  
Sandra Heuser ◽  
Astrid Wilbrand-Hennes ◽  
...  
Keyword(s):  
Human Keratinocytes ◽  
Signaling Cascades ◽  
Therapeutic Approaches ◽  
Primary Human Keratinocytes ◽  
Cellular Targets ◽  
Phosphorylated Proteins ◽  
Non Melanoma Skin Cancer ◽  
Damage Repair ◽  
Downstream Effects ◽  
Experimental Approaches

Human papillomavirus type 8 (HPV8) is associated with the development of non-melanoma skin cancer. In the past we already delved into the mechanisms involved in keratinocyte invasion, showing that the viral E7 oncoprotein is a key player that drives invasion of basal keratinocytes controlled by the extracellular protein fibronectin. To unravel further downstream effects in E7 expressing keratinocytes we now aimed at characterizing gene and protein/phosphoprotein alterations to narrow down on key cellular targets of HPV8-E7. We now show that gene expression of GADD34 and GDF15 are strongly activated in the presence of E7 in primary human keratinocytes. Further analyses of fibronectin-associated factors led to the identification of the Src kinase family members Fyn and Lyn being aberrantly activated in the presence of HPV8-E7. Phospho-proteomics further revealed that E7 not only targets cell polarity and cytoskeletal organization, but also deregulates the phosphorylation status of nuclear proteins involved in DNA damage repair and replication. Many of these differentially phosphorylated proteins turned out to be targets of Fyn and Lyn. Taken together, by using unbiased experimental approaches we have now arrived at a deeper understanding on how fibronectin may affect the signaling cascades in HPV8 positive keratinocytes, which may be key for skin tumorigenesis and that may also aid in the development of novel therapeutic approaches for betaHPV-mediated cancers.

scholarly journals The Effect of a Polyester Nanofibrous Membrane with a Fibrin-Platelet Lysate Coating on Keratinocytes and Endothelial Cells in a Co-Culture System

Nanomaterials ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 457
Author(s):  
Andreu Blanquer ◽  
Jana Musilkova ◽  
Elena Filova ◽  
Johanka Taborska ◽  
Eduard Brynda ◽  
...  
Keyword(s):  
Wound Healing ◽  
Endothelial Cells ◽  
Chronic Wounds ◽  
Human Keratinocytes ◽  
Skin Wound ◽  
Platelet Lysate ◽  
Therapeutic Approaches ◽  
Skin Wound Healing ◽  
New Therapeutic Approaches ◽  
Proliferation And Differentiation

Chronic wounds affect millions of patients worldwide, and it is estimated that this number will increase steadily in the future due to population ageing. The research of new therapeutic approaches to wound healing includes the development of nanofibrous meshes and the use of platelet lysate (PL) to stimulate skin regeneration. This study considers a combination of a degradable electrospun nanofibrous blend of poly(L-lactide-co-ε-caprolactone) and poly(ε-caprolactone) (PLCL/PCL) membranes (NF) and fibrin loaded with various concentrations of PL aimed at the development of bioactive skin wound healing dressings. The cytocompatibility of the NF membranes, as well as the effect of PL, was evaluated in both monocultures and co-cultures of human keratinocytes and human endothelial cells. We determined that the keratinocytes were able to adhere on all the membranes, and their increased proliferation and differentiation was observed on the membranes that contained fibrin with at least 50% of PL (Fbg + PL) after 14 days. With respect to the co-culture experiments, the membranes with fibrin with 20% of PL were observed to enhance the metabolic activity of endothelial cells and their migration, and the proliferation and differentiation of keratinocytes. The results suggest that the newly developed NF combined with fibrin and PL, described in the study, provides a promising dressing for chronic wound healing purposes.

Towards gene therapy for haemophilia B using primary human keratinocytes

1993 ◽  
Vol 3 (2) ◽  
pp. 180-183 ◽  
Author(s):  
Ann J. Gerrard ◽  
David L. Hudson ◽  
George G. Brownlee ◽  
Fiona M. Watt
Keyword(s):  
Gene Therapy ◽  
Human Keratinocytes ◽  
Primary Human Keratinocytes ◽  
Haemophilia B

Expression profiling of vitamin D treated primary human keratinocytes

2007 ◽  
Vol 100 (3) ◽  
pp. 574-592 ◽  
Author(s):  
Pamela Renate Moll ◽  
Veronika Sander ◽  
Anna-Maria Frischauf ◽  
Klaus Richter
Keyword(s):  
Vitamin D ◽  
Expression Profiling ◽  
Human Keratinocytes ◽  
Primary Human Keratinocytes

scholarly journals HaCaT Cells as a Reliable In Vitro Differentiation Model to Dissect the Inflammatory/Repair Response of Human Keratinocytes

2017 ◽  
Vol 2017 ◽  
pp. 1-12 ◽  
Author(s):  
Irma Colombo ◽  
Enrico Sangiovanni ◽  
Roberta Maggio ◽  
Carlo Mattozzi ◽  
Stefania Zava ◽  
...  
Keyword(s):  
Cell Density ◽  
Skin Diseases ◽  
Inflammatory Responses ◽  
Human Keratinocytes ◽  
Suitable Model ◽  
Hacat Cells ◽  
Primary Human Keratinocytes ◽  
Proinflammatory Mediators ◽  
Cytokines And Chemokines

Cultured primary human keratinocytes are frequently employed for studies of immunological and inflammatory responses; however, interpretation of experimental data may be complicated by donor to donor variability, the relatively short culture lifetime, and variations between passages. To standardize the in vitro studies on keratinocytes, we investigated the use of HaCaT cells, a long-lived, spontaneously immortalized human keratinocyte line which is able to differentiate in vitro, as a suitable model to follow the release of inflammatory and repair mediators in response to TNFα or IL-1β. Different treatment conditions (presence or absence of serum) and differentiation stimuli (increase in cell density as a function of time in culture and elevation of extracellular calcium) were considered. ELISA and Multiplex measurement technologies were used to monitor the production of cytokines and chemokines. Taken together, the results highlight that Ca2+ concentration in the medium, cell density, and presence of serum influences at different levels the release of proinflammatory mediators by HaCaT cells. Moreover, HaCaT cells maintained in low Ca2+ medium and 80% confluent are similar to normal keratinocytes in terms of cytokine production suggesting that HaCaT cells may be a useful model to investigate anti-inflammatory interventions/therapies on skin diseases.

Human keratinocytes migrate to the negative pole in direct current electric fields comparable to those measured in mammalian wounds

1996 ◽  
Vol 109 (1) ◽  
pp. 199-207 ◽  
Author(s):  
K.Y. Nishimura ◽  
R.R. Isseroff ◽  
R. Nuccitelli
Keyword(s):  
Electric Fields ◽  
Human Keratinocytes ◽  
Voltage Gradient ◽  
Primary Human Keratinocytes ◽  
Directed Movement ◽  
Average Cell ◽  
Negative Pole ◽  
Mammalian Skin ◽  
Dc Electric Fields

Previous measurements of the lateral electric fields near skin wounds in guinea pigs have detected DC fields between 100–200 mV/mm near the edge of the wound. We have studied the translocation response of motile primary human keratinocytes migrating on a collagen substrate while exposed to similar physiological DC electric fields. We find that keratinocytes migrate randomly on collagen in fields of 5 mV/mm or less, but in larger fields they migrate towards the negative pole of the field, exhibiting galvanotaxis. Since these cells have an average cell length of 50 microns, this implies that they are able to detect a voltage gradient as low as 0.5 mV along their length. This cath-odally-directed movement exhibits increased directedness with increasing field strengths between 10 and 100 mV/mm. We observe a maximally directed response at 100 mV/mm with half of the cells responding to the field within 14 minutes. The average speed of migration tended to be greater in fields above 50 mV/mm than in smaller fields. We conclude that human keratinocytes migrate towards the negative pole in DC electric fields that are of the same magnitude as measured in vivo near wounds in mammalian skin.

The hSkn-1a POU transcription factor enhances epidermal stratification by promoting keratinocyte proliferation

2001 ◽  
Vol 114 (10) ◽  
pp. 1913-1923 ◽  
Author(s):  
J. Hildesheim ◽  
U. Kuhn ◽  
C.L. Yee ◽  
R.A. Foster ◽  
K.B. Yancey ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Human Keratinocytes ◽  
Dominant Negative ◽  
Keratinocyte Differentiation ◽  
Transactivation Domain ◽  
Hacat Keratinocytes ◽  
Primary Human Keratinocytes ◽  
Keratinocyte Proliferation ◽  
Proliferation And Differentiation ◽  
Epidermal Homeostasis

Skn-1a is a POU transcription factor that is primarily expressed in the epidermis and is known to modulate the expression of several genes associated with keratinocyte differentiation. However, the formation of a stratified epidermis requires a carefully controlled balance between keratinocyte proliferation and differentiation, and a role for Skn-1a in this process has not been previously demonstrated. Here, our results show, surprisingly, that human Skn-1a contributes to epidermal stratification by primarily promoting keratinocyte proliferation and secondarily by enhancing the subsequent keratinocyte differentiation. In organotypic raft cultures of both primary human keratinocytes and immortalized HaCaT keratinocytes, human Skn-1a expression is associated with increased keratinocyte proliferation and re-epithelialization of the dermal substrates, resulting in increased numbers of keratinocytes available for the differentiation process. In these same raft cultures, human Skn-1a expression enhances the phenotypic changes of keratinocyte differentiation and the upregulated expression of keratinocyte differentiation genes. Conversely, expression of a dominant negative human Skn-1a transcription factor lacking the C-terminal transactivation domain blocks keratinocytes from proliferating and stratifying. Keratinocyte stratification is dependent on a precise balance between keratinocyte proliferation and differentiation, and our results suggest that human Skn-1a has an important role in maintaining epidermal homeostasis by promoting keratinocyte proliferation.

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