In vitro evaluation of a synthetic (Biobrane®) and a biopolymer (Epicite) wound dressing with primary human juvenile and adult fibroblasts after different colonization strategies.

BACKGROUND: The three-dimensional [3D] wound dressings Biobrane® and Epicite are used in the wound management. Fibroblasts are important for successful deep wound healing. The direct effect of Biobrane® and Epicite on human fibroblasts, particularly of juvenile individuals, remains unclear. Therefore, this study compared the survival and growth characteristics of juvenile and adult dermal fibroblasts on Biobrane® and Epicite using different culture models. METHOD: Murine (L929), primary juvenile and adult human fibroblasts were seeded on both materials using two dimensional (2D, slide culture) or 3D culture at the medium-air interface and dynamical rotatory culture. Cell adherence, viability, morphology, actin cytoskeleton architecture and DNA content were monitored. Scanning electron microscopy (SEM) analyses could be only performed from Biobrane®. Permeability of both materials were tested. RESULTS: The majority of all tested fibroblasts species survived on both dressings with no significant differences between 1 and 14 days. Juvenile and adult fibroblasts exerted typical fibroblast morphology with spindle-shaped cell bodies on the materials. SEM visualized morphological differences between murine and human fibroblasts on Biobrane®. Juvenile and adult fibroblasts colonized Biobrane® in rotatory culture after 7 days the most. The Biobrane® rotatory culture of L929 and juvenile fibroblasts showed after 7 days the significantly highest DNA amount. No major gender differences could be observed. Biobrane® had a higher permeability than Epicite. CONCLUSION: Both wound dressing can be colonized by fibroblasts suggesting their high cytocompatibility. Fibroblast survival and morphology on Biobrane® and Epicite depended on the culture system and the fibroblast source.

Complement activation induces excessive T cell cytotoxicity in severe COVID-19.

Severe COVID-19 is linked to both dysfunctional immune response and unrestrained immunopathology, and it remains unclear whether T cells contribute to disease pathology. Here, we combined single-cell transcriptomics and single-cell proteomics with mechanistic studies to assess pathogenic T cell functions and inducing signals. We identified highly activated CD16+ T cells with increased cytotoxic functions in severe COVID-19. CD16 expression enabled immune-complex-mediated, T cell receptor-independent degranulation and cytotoxicity not found in other diseases. CD16+ T cells from COVID-19 patients promoted microvascular endothelial cell injury and release of neutrophil and monocyte chemoattractants. CD16+ T cell clones persisted beyond acute disease maintaining their cytotoxic phenotype. Increased generation of C3a in severe COVID-19 induced activated CD16+ cytotoxic T cells. Proportions of activated CD16+ T cells and plasma levels of complement proteins upstream of C3a were associated with fatal outcome of COVID-19, supporting a pathological role of exacerbated cytotoxicity and complement activation in COVID-19.

Growth characteristics of human juvenile, adult and murine fibroblasts: a comparison of polymer wound dressings.

OBJECTIVE: Fibroblasts are important for the successful healing of deep wounds. However, the influence exerted by Cuticell, a natural polymer on fibroblasts and by the synthetic polymer, Suprathel, made of poly-L-lactic acid, is not sufficiently characterised. This study compared the survival and growth characteristics of human juvenile and adult dermal fibroblasts as well as murine fibroblast cell line L929, on a natural polymer with those of a synthetic polymer using different culture models. METHOD: Murine, juvenile and adult human fibroblasts were seeded on both the natural and synthetic polymers using statical slide culture or the medium air interface and dynamical rotatory culture. Cell adherence, viability, morphology and actin cytoskeleton architecture were monitored for 1-7 days. Biomaterial permeability was checked with a previously established diffusion chamber. RESULTS: The majority of the murine and adult human fibroblasts survived in slide and rotatory cultures on both wound dressings. The fibroblasts seeded on the synthetic polymer exhibited phenotypically a typical spread shape with multiple cell adhesion sites earlier than those on the natural polymer. The highest survival rates in all tested fibroblast species over the entire observation time were detected in rotatory culture (mean: >70%). Nevertheless, it led to cell-cluster formation on both materials. In the medium air interface culture, few adult fibroblasts adhered and survived until the seventh day of culture on both the natural and synthetic polymers, and no viable juvenile and L929 fibroblasts could be found by day seven. Apart from a significant higher survival rate of L929 in slide culture on the natural polymer compared with the synthetic polymer at the end of the culturing period (p<0.0001), and a higher cell survival of L929 on the natural polymer in medium air interface culture, only minor differences between both materials were evident. This suggested a comparable cytocompatibility of both materials. Permeability testing revealed slightly higher permeance of the natural polymer compared with the synthetic polymer. CONCLUSION: Cell survival rates depended on the culture system and the fibroblast source. Nevertheless, the juvenile skin fibroblasts were the most sensitive. This observation suggests that wound dressings used in treating children should be tested beforehand with juvenile fibroblasts to ensure the dressing does not compromise wound healing. Future experiments should also include the response of compromised fibroblasts, for example, from burn patients.