The treatment of chronic wounds still challenges modern medicine because of these wounds’ heterogenic pathophysiology. Processes such as inflammation, ischemia and bacterial infection play major roles in the progression of a chronic wound. In recent years, preclinical wound models have been used to understand the underlying processes of chronic wound formation. However, the wound models used to investigate chronic wounds often lack translatability from preclinical models to patients, and often do not take exaggerated inflammation into consideration. Therefore, we aimed to investigate prolonged inflammation in a porcine wound model by using resiquimod, a TLR7 and TLR8 agonist. Pigs received full thickness excisional wounds, where resiquimod was applied daily for 6 days, and untreated wounds served as controls. Dressing change, visual documentation and wound scoring were performed daily. Biopsies were collected for histological as well as gene expression analysis. Resiquimod application on full thickness wounds induced a visible inflammation of wounds, resulting in delayed wound healing compared to non-treated control wounds. Gene expression analysis revealed high levels of IL6, MMP1 and CD68 expression after resiquimod application, and histological analysis showed increased immune cell infiltration. By using resiquimod, we were able to show that prolonged inflammation delayed wound healing, which is often observed in chronic wounds in patients. The model we used shows the importance of inflammation in wound healing and gives an insight into the progression of chronic wounds.

Extracellular vesicles (EVs) and their miRNA cargo are intercellular communicators transmitting their pleiotropic messages between different cell types, tissues, and body fluids. Recently, they have been reported to contribute to skin homeostasis and were identified as members of the senescence-associated secretory phenotype of human dermal fibroblasts. However, the role of EV-miRNAs in paracrine signaling during skin aging is yet unclear. Here we provide evidence for the existence of small EVs in the human skin and dermal interstitial fluid using dermal open flow microperfusion and show that EVs and miRNAs are transferred from dermal fibroblasts to epidermal keratinocytes in 2D cell culture and in human skin equivalents. We further show that the transient presence of senescent fibroblast derived small EVs accelerates scratch closure of epidermal keratinocytes, whereas long-term incubation impairs keratinocyte differentiation in vitro. Finally, we identify vesicular miR-23a-3p, highly secreted by senescent fibroblasts, as one contributor of the EV-mediated effect on keratinocytes in in vitro wound healing assays. To summarize, our findings support the current view that EVs and their miRNA cargo are members of the senescence-associated secretory phenotype and, thus, regulators of human skin homeostasis during aging.

The inability of leptin to suppress food intake in diet-induced obesity, sometimes referred to as leptin resistance, is associated with several distinct pathological hallmarks. One prevailing theory is that impaired transport of leptin across the blood-brain barrier (BBB) represents a molecular mechanism that triggers this phenomenon. Recent evidence, however, has challenged this notion, suggesting that leptin BBB transport is acquired during leptin resistance. To resolve this debate, we utilized a novel cerebral Open Flow Microperfusion (cOFM) method to examine leptin BBB transport in male C57BL/6J mice, fed a chow diet or high fat diet (HFD) for 20 days. Basal plasma leptin levels were 3.8-fold higher in HFD-fed mice (p < 0.05). Leptin administration (2.5 mg/kg) elicited similar pharmacokinetic profiles of circulating leptin. However, while leptin reduced food intake by 20% over 22 h in chow-fed mice, it did not affect food intake in HFD-fed mice. In spite of this striking functional difference, hypothalamic leptin levels, as measured by cOFM, did not differ between chow-fed mice and HFD-fed mice following leptin administration. These data suggest that leptin transport across the BBB is not impaired in non-obese leptin resistant mice and thus unlikely to play a direct role in the progression of pharmacological leptin resistance.

In burn injuries, risk factors and limitations to treatment success are difficult to assess clinically. However, local cellular responses are characterized by specific gene-expression patterns. MicroRNAs (miRNAs) are single-stranded, non-coding RNAs that regulate mRNA expression on a posttranscriptional level. Secreted through exosome-like vesicles (ELV), miRNAs are intracellular signalers and epigenetic regulators. To date, their role in the regulation of the early burn response remains unclear. Here, we identified 43 miRNAs as potential regulators of the early burn response through the bioinformatics analysis of an existing dataset. We used an established human ex vivo skin model of a deep partial-thickness burn to characterize ELVs and miRNAs in dermal interstitial fluid (dISF). Moreover, we identified miR-497-5p as stably downregulated in tissue and dISF in the early phase after a burn injury. MiR-218-5p and miR-212-3p were downregulated in dISF, but not in tissue. Target genes of the miRNAs were mainly upregulated in tissue post-burn. The altered levels of miRNAs in dISF of thermally injured skin mark them as new biomarker candidates for burn injuries. To our knowledge, this is the first study to report miRNAs altered in the dISF in the early phase of deep partial-thickness burns.

Cornelia Pipper, Natalie Bordag, Bernadette Reiter, Kyriakos Economides, Peter Florian, Thomas Birngruber, Frank Sinner, Manfred Bodenlenz, Anita Eberl