Effect of Health Status and Heat-Induced Inactivation on the Proteomic Profile of Plasma Rich in Growth Factors Obtained from Donors with Chronic Inflammatory Skin Conditions

Abstract

Atopic dermatitis, psoriasis and lichen sclerosus are among the most challenging conditions treated by dermatologists worldwide, with potentially significant physical, social and psychological impacts. Emerging evidence suggests that autologous-platelet-rich plasma could be used to manage skin inflammation. However, the presence of soluble autoimmune components could hinder their therapeutic potential. The aim of this study was to analyze the proteomic profile of plasma rich in growth factors (PRGFs) obtained from donors with inflammatory skin conditions to evaluate the impact of skin health status on the composition and bioactivity of PRGF-based treatments. Venous blood from healthy volunteers and patients with psoriasis, lichen sclerosus and atopic dermatitis was processed to produce PRGF supernatant. Half of the samples were subjected to an additional thermal treatment (56 °C) to inactivate inflammatory and immune molecules. Proteomic analysis was performed to assess the protein profile of PRGFs from healthy and non-healthy patients and the effect of Immunosafe treatment. Differential abundance patterns of several proteins related to key biological processes have been identified, including complement activation, blood coagulation, and glycolysis- and gluconeogenesis-related genes. These results also demonstrate that the thermal treatment (Immunosafe) contributes to the inactivation of the complement system and, as a consequence, reduction in the immunogenic potential of PRGF products.

Keywords: atopic dermatitis; lichen sclerosus; platelet-rich plasma; proteome; psoriasis; skin inflammation; thermal inactivation.

Figure 1

Proteomic heatmap with hierarchical clustering of MS-based label-free quantified proteins (LFQ) across the PRGF-supernatant-derived samples (SPs and INSPs) obtained from donors with different underlying health conditions: health, dermatitis, psoriasis, and lichen sclerosus. The protein list has been filtered to contain exclusively differentially abundant proteins identified in at least one pairwise comparison (p ≤ 0.05).

Figure 2

Volcano plots representing differentially abundant proteins in PRGF supernatants (SPs) and Immunosafe-treated PRGF supernatants (INSPs) between healthy (H) and pathological donors: dermatitis (D), psoriasis (P) and lichen sclerosus (L). The plots were constructed using log2 fold-change (log2FC) and p values. The colored dots represent the following cases: proteins upregulated in PRGF samples from pathological donors (blue), not differentially abundant proteins (green), and proteins downregulated in PRGF samples from pathological donors (red), at p ≤ 0.05.

Figure 3

Schematic summary of differential protein expression between healthy and pathological donors as revealed by quantitative proteomic profiling.

Figure 4

Volcano plots represent differentially abundant proteins in PRGF supernatants (SPs) and Immunosafe-treated PRGF supernatants (INSPs) between pathological donors: dermatitis (D), psoriasis (P) and lichen sclerosus (L). The plots were constructed using log2 fold-change (log2FC) and p values. The colored dots represent the following cases: proteins upregulated in PRGF samples from the following pathological donors: P, L and L, respectively (blue), not differentially abundant proteins (green), and proteins downregulated in PRGF samples from the following pathological donors: P, L, and L, in each case (red).

Figure 5

Functional analyses of differentially expressed genes (DEGs) between PRGF supernatants (SPs) and Immunosafe-treated PRGF supernatants (INSPs) obtained from donors including those with dermatitis (D), psoriasis (P) and lichen sclerosus (L) disorders and healthy donors (H). (A) Sankey dots diagrams representing Gene Ontology (GO) enrichment analyses, and (B) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment maps. Differentially expressed gene products are colored in red (upregulated in pathological supernatants) or green (downregulated in pathological supernatants). GO and KEGG enrichment analyses of DEGs were performed using DAVID. Only significantly enriched KEGG pathways and GO terms in biological process (BP), cellular component (CC), and molecular function (MF) branches are presented (p ≤ 0.05). All the statistically significant p values of the terms were negative 10-base log transformed.

Figure 6

Functional analyses of differentially expressed genes (DEGs) in PRGF supernatants (SPs) and Immunosafe-treated PRGF supernatants (INSPs) between pathological donors: dermatitis (D), psoriasis (P) and lichen sclerosus (L). (A) Sankey dot diagrams representing Gene Ontology (GO) enrichment analyses, and (B) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment maps. Differentially expressed gene products are colored in red (upregulated in P, L, and L, respectively) or green (downregulated in P, L, and L, in each case). GO and KEGG enrichment analyses of DEGs were retrieved using DAVID. Only significantly enriched KEGG pathways and GO terms in biological process (BP), cellular component (CC), and molecular function (MF) branches are presented (p ≤ 0.05). All the statistically significant p values of the terms were negative 10-base log transformed.

Figure 7

Volcano plots representing differentially abundant proteins between PRGF supernatants (SPs) and Immunosafe-treated PRGF supernatants (INSPs) within health condition: healthy (H), dermatitis (D), psoriasis (P) and lichen sclerosus (L). The plots were constructed using log2 fold-change (log2FC) and p values. The colored dots represent the following cases: proteins upregulated in SP samples (blue), not differentially abundant proteins (green), and proteins downregulated in SP samples (red), at p ≤ 0.05.

Figure 8

Schematic summary of the effect of Immunosafe treatment on differential protein abundance of PRGF supernatants as revealed by quantitative proteomic profiling.

Figure 9

Functional analyses of differentially abundant gene products between PRGF supernatants (SPs) and Immunosafe-treated PRGF supernatants (INSPs) within health condition: healthy (H), dermatitis (D), psoriasis (P) and lichen sclerosus (L). (A) Sankey dot diagrams representing Gene Ontology (GO) enrichment analyses, and (B) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment maps. Differentially expressed gene products are colored in red (upregulated in SP) or green (downregulated in SP). GO and KEGG enrichment analyses of differentially abundant gene products were retrieved using DAVID. Only significantly enriched KEGG pathways and GO terms in biological process (BP), cellular component (CC), and molecular function (MF) branches are presented (p ≤ 0.05). All the statistically significant p values of the terms were negative 10-base log transformed.