Orf virus (ORFV) infection in a three-dimensional human skin model: Characteristic cellular alterations and interference with keratinocyte differentiation

Abstract

ORF virus (ORFV) is the causative agent of contagious ecthyma, a pustular dermatitis of small ruminants and humans. Even though the development of lesions caused by ORFV was extensively studied in animals, only limited knowledge exists about the lesion development in human skin. The aim of the present study was to evaluate a three-dimensional (3D) organotypic culture (OTC) as a human skin model for ORFV infection considering lesion development, replication of the virus, viral gene transcription and modulation of differentiation of human keratinocytes by ORFV. ORFV infection of OTC was performed using the ORFV isolate B029 derived from a human patient. The OTC sections showed a similar structure of stratified epidermal keratinocytes as human foreskin and a similar expression profile of the differentiation markers keratin 1 (K1), K10, and loricrin. Upon ORFV infection, OTCs exhibited histological cytopathic changes including hyperkeratosis and ballooning degeneration of the keratinocytes. ORFV persisted for 10 days and was located in keratinocytes of the outer epidermal layers. ORFV-specific early, intermediate and late genes were transcribed, but limited viral spread and restricted cell infection were noticed. ORFV infection resulted in downregulation of K1, K10, and loricrin at the transcriptional level without affecting proliferation as shown by PCNA or Ki-67 expression. In conclusion, OTC provides a suitable model to study the interaction of virus with human keratinocytes in a similar structural setting as human skin and reveals that ORFV infection downregulates several differentiation markers in the epidermis of the human skin, a hitherto unknown feature of dermal ORFV infection in man.

A Review on Human Orf: A Neglected Viral Zoonosis

“…31,75 Although both cell-mediated and humoral immune responses have been confirmed in sheep and human ORFV infections, cell-mediated immunity plays the major role against the virus. 75,76 Antibodies do not appear to confer protective immunity to ORFV although IgG2 isotype might be important in defense against the virus. IgG2 is not transported in the milk of ruminants which might explain why colostral antibodies are not protective in lambs and kids.…”

Requisite instruments for the establishment of three‐dimensional epidermal human skin equivalents—A methods review

“…91,92 Alternatively, juvenile foreskin can be used. 45,[93][94][95] The use of younger skin is preferred because cells from younger donors are more robust, appear to have a higher proliferative capacity and have greater longevity than samples from older donors. 68,94 The time between surgery and isolation of cells must be kept as short as possible (approx.…”

“…First approach: 43,45,95 In brief -To separate skin into epidermis and dermis dispase II solution is used. Afterwards, the epidermis is digested using trypsin/ ethylenediaminetetraacetic acid (EDTA).…”

“…Skin research covers a variety of areas including: skin disease, 42-44 infection, 45 pharmaceuticals, [46][47][48][49] ageing, 50 toxicology, [51][52][53] melanoma models, 54 wound healing 39,46,[55][56][57][58][59] and impacts of climate change on the skin. 60 Studying the skin requires robust and reliable experimental models that mimic in vivo conditions closely enough for safe translation to human use.…”

Investigación de parámetros bioquímicos y perfil de citocinas en ovejas con ectima contagioso

“…Varios estudios han establecido la condición de células inmunes en la piel de las ovejas reinfectadas o infectadas por primera vez. Las principales células de los queratinocitos infectados son los neutrófilos, las células B, las células dendríticas y las células T (10,11).…”