Neutrophils provide cellular communication between ileum and mesenteric lymph nodes at graft-versus-host disease onset.

Conditioning-induced damage of the intestinal tract plays a critical role during the onset of acute graft-versus-host disease (GVHD). Therapeutic interference with these early events of GVHD is difficult, and currently used immunosuppressive drugs mainly target donor T cells. However, not donor T cells but neutrophils reach the sites of tissue injury first, and therefore could be a potential target for GVHD prevention. A detailed analysis of neutrophil fate during acute GVHD and the effect on T cells is difficult because of the short lifespan of this cell type. By using a novel photoconverter reporter system, we show that neutrophils that had been photoconverted in the ileum postconditioning later migrated to mesenteric lymph nodes (mLN). This neutrophil migration was dependent on the intestinal microflora. In the mLN, neutrophils colocalized with T cells and presented antigen on major histocompatibility complex (MHC)-II, thereby affecting T cell expansion. Pharmacological JAK1/JAK2 inhibition reduced neutrophil influx into the mLN and MHC-II expression, thereby interfering with an early event in acute GVHD pathogenesis. In agreement with this finding, neutrophil depletion reduced acute GVHD. We conclude that neutrophils are attracted to the ileum, where the intestinal barrier is disrupted, and then migrate to the mLN, where they participate in alloantigen presentation. JAK1/JAK2-inhibition can interfere with this process, which provides a potential therapeutic strategy to prevent early events of tissue damage-related innate immune cell activation and, ultimately, GVHD.

Production of BMP4 by endothelial cells is crucial for endogenous thymic regeneration.

The thymus is not only extremely sensitive to damage but also has a remarkable ability to repair itself. However, the mechanisms underlying this endogenous regeneration remain poorly understood, and this capacity diminishes considerably with age. We show that thymic endothelial cells (ECs) comprise a critical pathway of regeneration via their production of bone morphogenetic protein 4 (BMP4) ECs increased their production of BMP4 after thymic damage, and abrogating BMP4 signaling or production by either pharmacologic or genetic inhibition impaired thymic repair. EC-derived BMP4 acted on thymic epithelial cells (TECs) to increase their expression of Foxn1, a key transcription factor involved in TEC development, maintenance, and regeneration, and its downstream targets such as Dll4, a key mediator of thymocyte development and regeneration. These studies demonstrate the importance of the BMP4 pathway in endogenous tissue regeneration and offer a potential clinical approach to enhance T cell immunity.

Exogenous TNFR2 activation protects from acute GvHD via host T reg cell expansion.

Donor CD4(+)Foxp3(+) regulatory T cells (T reg cells) suppress graft-versus-host disease (GvHD) after allogeneic hematopoietic stem cell transplantation (HCT [allo-HCT]). Current clinical study protocols rely on the ex vivo expansion of donor T reg cells and their infusion in high numbers. In this study, we present a novel strategy for inhibiting GvHD that is based on the in vivo expansion of recipient T reg cells before allo-HCT, exploiting the crucial role of tumor necrosis factor receptor 2 (TNFR2) in T reg cell biology. Expanding radiation-resistant host T reg cells in recipient mice using a mouse TNFR2-selective agonist before allo-HCT significantly prolonged survival and reduced GvHD severity in a TNFR2- and T reg cell-dependent manner. The beneficial effects of transplanted T cells against leukemia cells and infectious pathogens remained unaffected. A corresponding human TNFR2-specific agonist expanded human T reg cells in vitro. These observations indicate the potential of our strategy to protect allo-HCT patients from acute GvHD by expanding T reg cells via selective TNFR2 activation in vivo.

Impact of non-thermal plasma treatment on MAPK signaling pathways of human immune cell lines.

In the field of wound healing research non-thermal plasma (NTP) increasingly draws attention. Next to its intensely studied antibacterial effects, some studies already showed stimulating effects on eukaryotic cells. This promises a unique potential in healing of chronic wounds, where effective therapies are urgently needed. Immune cells do play an important part in the process of wound healing and their reaction to NTP treatment has yet been rarely examined. Here, we studied the impact of NTP treatment using the kinpen on apoptotic and proliferative cell signaling pathways of two human immune cell lines, the CD4(+)T helper cell line Jurkat and the monocyte cell line THP-1. Depending on NTP treatment time the number of apoptotic cells increased in both investigated cell types according to a caspase 3 assay. Western blot analysis pointed out that plasma treatment activated pro-apoptotic signaling proteins like p38 mitogen-activated protein kinase (p38 MAPK) and c-Jun N-terminal kinase 1 and 2 (JNK 1/2) in both cell types. Stronger signals were detected in Jurkat cells at comparable plasma treatment times. Intriguingly, exposure of Jurkat and THP-1 cells to plasma also activated the pro-proliferative signaling molecules extracellular signal-regulated kinase 1/2 (ERK 1/2) and MAPK/ERK kinase 1 and 2 (MEK 1/2). In contrast to Jurkat cells, the anti-apoptotic heat shock protein 27 (HSP27) was activated in THP-1 cells after plasma treatment, indicating a possible mechanism how THP-1 cells may reduce programmed cell death. In conclusion, several signaling cascades were activated in the examined immune cell lines after NTP treatment and in THP-1 monocytes a possible defense mechanism against plasma impacts could be revealed. Therefore, plasma might be a treatment option for wound healing.

Non-thermal plasma treatment is associated with changes in transcriptome of human epithelial skin cells.

Non-thermal atmospheric pressure plasma has recently gained attention in the field of biomedical and clinical applications. In the area of plasma medicine research, one promising approach is to promote wound healing by stimulation of cells involved. To understand basic molecular and cellular mechanisms triggered by plasma treatment, we investigated biological effects of an argon plasma jet kinpen on human epithelial skin cells. For assessment of transcriptome changes cell culture medium was plasma treated and applied to the HaCaT keratinocyte cell culture (indirect treatment). Consequently, whole-genome microarrays were used to analyze this interaction in detail and identified a statistically significant modification of 3,274 genes including 1,828 up- and 1,446 downregulated genes. Particularly, cells after indirect plasma treatment are characterized by differential expression of a considerable number of genes involved in the response to stress. In this regard, we found a plasma-dependent regulation of oxidative stress answer and increased expression of enzymes of the antioxidative defense system (e.g. 91 oxidoreductases). Our results demonstrate that plasma not only induces cell reactions of stress-sensing but also of proliferative nature. Consistent with gene expression changes as well as Ingenuity Pathway Analysis prediction, we propose that stimulating doses of plasma may protect epithelial skin cells in wound healing by promoting proliferation and differentiation. In conclusion, gene expression profiling may become an important tool in identifying plasma-related changes of gene expression. Our results underline the enormous clinical potential of plasma as a biomedical tool for stimulation of epithelial skin cells.