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Objectives Approximately 20% of people with psoriasis develop PsA. Although genetic, clinical and environmental risk factors have been identified, it is not known why some people with psoriasis develop PsA. The skin disease is traditionally considered the same in both. This study compares transcriptional changes in psoriasis and PsA skin for the first time. Methods Skin biopsies were collected from healthy controls (HC), and uninvolved and lesional skin from patients with PsA. Bulk tissue sequencing was performed and analysed using the pipeline Searchlight 2.0. Transcriptional changes in PsA skin were compared with existing sequencing data from participants with psoriasis without PsA (GSE121212). Psoriasis and PsA datasets could not be directly compared as different analysis methods were used. Data from participants with PsA in the GSE121212 dataset were used for validation. Results Skin samples from 9 participants with PsA and 9 HC were sequenced, analysed and compared with available transcriptomic data for 16 participants with psoriasis compared with 16 HC. Uninvolved skin in psoriasis shared transcriptional changes with lesional skin in psoriasis, but uninvolved skin in PsA did not. Most transcriptional changes in psoriasis and PsA lesional skin were shared, but immunoglobulin genes were upregulated in PsA lesional skin specifically. The transcription factor POU2F1, which regulates immunoglobulin gene expression, was enriched in PsA lesional skin. This was confirmed in the validation cohort. Conclusions Immunoglobulin genes are upregulated in PsA but not in psoriasis skin lesions. This may have implications for the spread from the cutaneous compartment to other tissues.

The migration of dendritic cells from tissues to draining lymph nodes is a critical step in the induction of peripheral tolerance and the initiation of adaptive immune responses. This is dependent on CCR7 expression by dendritic cells. In response to the chemokine CCL21, CCR7 directs dendritic cells into lymphatic vessels in the tissue, and permits their transit from the subcapsular sinus into the lymph node parenchyma. The CCR7 ligands CCL19 and CCL21 also bind ACKR4, an atypical chemokine receptor expressed by keratinocytes in the skin and lymphatic endothelial cells lining the subcapsular sinus. In mice, ACKR4 controls interfollicular CCL21 gradients, and enhances dendritic cell entry into the lymph node parenchyma from the subcapsular sinus. Here we report that Ackr4 deficiency disrupts CCR7-dependent dendritic cell arrival at skin-draining lymph nodes during cutaneous inflammation. We show that this, at least in part, is due to the defective departure of dendritic cells from inflamed skin, and is accompanied by dysregulation of bioavailable CCL19 and CCL21 in the skin. Strikingly, genetic deletion of CCL19 completely rescues the defective inflammation-driven trafficking of dendritic cells caused by Ackr4 deficiency. Thus, by regulating CCL19, ACKR4 helps maintain CCR7-dependent dendritic cell departure from inflamed tissues.

We have assessed expression of MIP-1alpha binding sites on the surface of CD34+ cells from normal bone marrow (NBM) and chronic myeloid leukemia (CML) peripheral blood. This study has highlighted a small subpopulation of CD34+ (15.7 +/- 6.2% in NBM and 9 +/- 4% in CML), which has specific macrophage-inflammatory protein-1alpha (MIP-1alpha) cell surface binding sites. Further phenotypic characterization of the receptor-bearing cells has shown that they do not express the Thy-1 Ag, suggesting that they are committed progenitor cells rather than CD34+ Thy+ stem cells. However, more than 80% of methanol-fixed CD34+ cells do bind MIP-1alpha, suggesting that these cells may possess a pool of internal receptors, although we were unable to induce cell surface expression by cytokine stimulation. The percentage of these CD34+, MIP-1alpha-R+ cells present in the CD34 compartment of NBM is significantly higher than in CML, implicating lack of binding sites as part of the mechanism for the loss of response to this chemokine seen in CML. Specific Ab to the MIP-1alpha receptor implicated in HIV infection, CCR5, revealed that very few CD34+ cells expressed these receptors and that expression was confined to the CD34+ Thy- progenitor population. Data presented in this work suggest that active binding sites for the stem cell growth inhibitor MIP-1alpha are not constitutively expressed on the surface of most resting primitive multipotent cells, and that these cells are not potential targets for HIV-1 infection through CCR5.

Macrophage inhibitory protein (MIP)-1 alpha is a potent inhibitor of hemopoietic stem cell proliferation and is a member of a family of pro-inflammatory mediators, the chemokine family. This molecule along with other members of the chemokine family exists as a peptide of 8 kDa but has a strong tendency for noncovalent extensive self-aggregation. As this aggregation may interfere with biological activity, we have produced nonaggregating variants of MIP-1 alpha which display a range of molecular sizes. The mutants, produced by sequential neutralization of carboxyl-terminal acidic residues, display native molecular masses representative of tetramers, dimers, and monomers. Intriguingly when these mutants are assessed in comparison with native MIP-1 alpha for bioactivity in vitro, they are seen to be equipotent in both stem cell assays and in monocyte shape-change assays, suggesting that there is no requirement for aggregation in either of these biological contexts. This indicates that the aggregated MIP-1 alpha and the aggregated mutants spontaneously disaggregate under assay conditions and ultimately function as monomers. We have further demonstrated the ability of MIP-1 alpha to disaggregate spontaneously in dilute solution by enzyme-linked immunosorbent assay analysis of fractions obtained from gel filtration of varying concentrations of MIP-1 alpha. The aggregation of MIP-1 alpha is therefore a dynamic and reversible phenomenon which has little, if any, impact on bioactivity in vitro.

A macrophage-derived inhibitor of early hematopoietic progenitors (colony-forming unit-spleen, CFU-A) called stem cell inhibitor was found to be identical to macrophage inflammatory protein-1 alpha (MIP-1 alpha). We investigated the effect of MIP-1 alpha on the earliest stem cells that sustain long-term hematopoiesis in vivo in a competitive bone marrow repopulation assay. Because long-term reconstituting (LTR) stem cells are normally quiescent, an in vivo model was first developed in which they are triggered to cycle. A first 5-fluorouracil (5-FU) injection was used to eliminate later progenitors, causing the LTR stem cells, which are normally resistant to 5-FU, to enter the cell cycle and become sensitive to a second 5-FU injection administered 5 days later. Human MIP-1 alpha administered from day 0 to 7 was unable to prevent the depletion of the LTR stem cells by the second 5-FU treatment, as observed on day 7 in this model, suggesting that the LTR stem cells were not prevented from being triggered into cycle despite the MIP-1 alpha treatment. However, the MIP-1 alpha protocol used here did substantially decrease the number of more mature hematopoietic progenitors (granulocyte-macrophage colony-forming cells [CFC], burst- forming unit-erythroid, CFCmulti, and preCFCmulti) recovered in the bone marrow shortly after a single 5-FU injection. In vitro, MIP-1 alpha had no inhibitory effect on the ability of these progenitors to form colonies. This study confirms the in vivo inhibitory effect of MIP- 1 alpha on subpopulations of hematopoietic progenitors that are activated in myelodepressed animals. However, MIP-1 alpha had no effect on the long-term reconstituting stem cells in vivo under conditions in which it effectively reduced all later progenitors.

The characterization of many cytokines involved in the control of hematopoiesis has led to intense investigation into their potential use in ex vivo culture to expand progenitor numbers. We have established the optimum ex vivo culture conditions that allow substantial amplification of transient engrafting murine stem cells and which, simultaneously, augment the ability to sustain serial bone marrow transplantation (BMT). Short-term incubation of unfractionated BM cells in liquid culture with stem cell factor (SCF) and interleukin-11 (IL-11) produced a 50-fold amplification of clonogenic multipotential progenitors (CFU-A). Following such ex vivo expansion, substantially fewer cells were required to rescue lethally irradiated mice. When transplanted in cell doses above threshold for engraftment, BM cells expanded ex vivo resulted in significantly more rapid hematopoietic recovery. In a serial transplantation model, unmanipulated BM was only able to consistently sustain secondary BMT recipients, but BM expanded ex vivo has sustained quaternary BMT recipients that remain alive and well more than 140 days after 4th degree BMT. These results show augmentation of both short-term recovery posttransplant and the ability to serially transplant marrow by preincubation in culture with SCF and IL-11.

Macrophage inflammatory protein 1 alpha (MIP-1 alpha) is a potent stem cell inhibitor and a member of a large and expanding family of related cytokines. In an effort to understand the molecular basis of the activities of MIP-1 alpha, we have sought to characterize the cellular receptors for this molecule. Our results demonstrate the presence of abundant MIP-1 alpha receptors on both human and murine cells. The receptor on K562 cells can bind a range of members of the MIP-1 alpha family and may thus be a general MIP-1 alpha family receptor. Murine FDCPmix cells also bind a range of members of this peptide family, although the receptor(s) that they express appear somewhat more selective for peptides capable of displaying stem cell inhibitory properties. The human and murine receptors do not bind members of the related interleukin 8 family of peptides and are thus distinct from the recently cloned interleukin 8 receptor. We suggest that the receptor on the murine cell is a candidate for the receptor responsible for articulating stem cell inhibitory signals following MIP-1 alpha binding.

TGF-beta and macrophage inflammatory protein-1 alpha (MIP-1 alpha) appear to share a number of biologic properties. We have been attempting to examine the interactions between these two peptides in the hope of gaining an insight into the basis for the apparent functional redundancy. Our earlier observations have indicated that TGF-beta is a potent down-regulator of MIP-1 alpha and MIP-1 beta expression in bone marrow macrophages and also of MIP-1 alpha receptor numbers on FDCPmix cells. We now demonstrate that the interplay between TGF-beta and MIP-1 alpha beta is relatively specific, in that only MIP-1 alpha and MIP-1 beta appear to be potently suppressed by TGF-beta, and that this suppressive activity is restricted to the direct TGF-beta isoforms. Activin and the bone morphogenetic proteins (BMPs) appear to be inactive in this regard. We also demonstrate the existence of an endogenous TGF-beta-mediated block that acts to minimize MIP-1 alpha expression in TGF-beta-expressing macrophages. This coupled with the observations that MIP-1 alpha can induce expression of TGF-beta suggests to us that the complex interactions between MIP-1 alpha and MIP-1 beta and the direct TGF-beta isoforms (beta 1, beta 2, and beta 3) act to ensure minimized MIP-1 alpha beta expression and maximized TGF-beta expression. However, such interplay may also be dependent on the local cytokine or inflammatory profile to which the cells are exposed.