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Mastocytosis is a KIT-related myeloproliferative disease characterised by abnormal expansion of neoplastic mast cells (MC) in the skin or virtually any other organ system. The cutaneous form of adult-onset mastocytosis is almost invariably combined with indolent systemic involvement for which curative therapy is yet not available. Here we evaluated a concept of depleting cutaneous MCs in mastocytosis lesions ex vivo by targeting their secretory granules. Skin biopsies from mastocytosis patients were incubated with or without mefloquine, an antimalarial drug known to penetrate into acidic organelles such as MC secretory granules. Mefloquine reduced the number of dermal MCs without affecting keratinocyte proliferation or epidermal gross morphology at drug concentrations up to 40 mu M. Flow cytometric analysis of purified dermal MCs showed that mefloquine-induced cell death was mainly due to apoptosis and accompanied by caspase-3 activation. However, caspase inhibition provided only partial protection against mefloquine-induced cell death, indicating predominantly caspase-independent apoptosis. Further assessments revealed that mefloquine caused an elevation of granule pH and a corresponding decrease in cytosolic pH, suggesting drug-induced granule permeabilisation. Extensive damage to the MC secretory granules was confirmed by transmission electron microscopy analysis. Further, blockade of granule acidification or serine protease activity prior to mefloquine treatment protected MCs from apoptosis, indicating that granule acidity and granule-localised serine proteases play major roles in the execution of mefloquine-induced cell death. Altogether, these findings reveal that mefloquine induces selective apoptosis of MCs by targeting their secretory granules and suggest that the drug may potentially extend its range of medical applications.

Chronic local inflammation of adipose tissue is an important feature of obesity. Serglycin is a proteoglycan highly expressed by various immune cell types known to infiltrate adipose tissue under obese conditions. To investigate if serglycin expression has an impact on diet-induced adipose tissue inflammation, we subjected Srgn+/+ and Srgn−/− mice (C57BL/6J genetic background) to an 8-wk high-fat and high-sucrose diet. The total body weight was the same in Srgn+/+ and Srgn−/− mice after diet treatment. Expression of white adipose tissue genes linked to inflammatory pathways were lower in Srgn−/− mice. We also noted reduced total macrophage abundance, a reduced proportion of proinflammatory M1 macrophages, and reduced formation of crown-like structures in adipose tissue of Srgn−/− compared with Srgn+/+ mice. Further, Srgn−/− mice had more medium-sized adipocytes and fewer large adipocytes. Differentiation of preadipocytes into adipocytes (3T3-L1) was accompanied by reduced Srgn mRNA expression. In line with this, analysis of single-cell RNA sequencing data from mouse and human adipose tissue supports that Srgn mRNA is predominantly expressed by various immune cells, with low expression in adipocytes. Srgn mRNA expression was higher in obese compared with lean humans and mice, accompanied by an increased expression of immune cell gene markers. SRGN and inflammatory marker mRNA expression was reduced upon substantial weight loss in patients after bariatric surgery. Taken together, this study introduces a role for serglycin in the regulation of obesity-induced adipose inflammation.

Background/Aim: Mast cells are abundant in melanoma tumors, and studies suggest that they can be either detrimental or protective for melanoma growth. However, the underlying mechanisms are not fully understood. Materials and Methods: Here, we adopted an established hanging-drop spheroid system to investigate how mast cells influence melanoma growth and phenotype in a 3-D context. To address the underlying mechanism, we conducted transcriptomic and pathway analyses. Results: In the presence of mast cells or mast cell-conditioned medium, growth of melanoma spheroids was profoundly reduced. Transcriptomic analysis revealed that mast cell-conditioned medium had extensive effects on the gene-expression patterns of melanoma. Pathway analyses revealed profound effects on the expression of genes related to amino acid and protein metabolism. The conditioned medium also induced up-regulation of cancer-related genes, including adhesion molecules implicated in metastatic spreading. In line with this, after transfer to a Matrigel extracellular matrix milieu, spheroids that had been developed in the presence of mast cell-conditioned medium displayed enhanced growth and adhesive properties. However, when assessing the possible impact of nutrient starvation, i.e., reduced nutrient content in mast cell-conditioned medium, we found that the observed effects on growth of melanoma spheroids could potentially be explained by such a scenario. Conclusion: Our findings suggest that the phenotypic alterations of melanoma spheroids grown in the presence of mast cells or mast cell-conditioned media are, at least partly, due to nutrient starvation rather than to the action of factors secreted by mast cells. Our findings may provide insight into the effects on gene-expression events that occur in melanoma tumors under nutrient stress.

Mast cells are abundant in melanoma tumors, and studies suggest that they can be either detrimental or protective for melanoma growth. However, the underlying mechanisms are not fully understood. Here, we adopted a hanging drop-established spheroid system to investigate how mast cells can influence melanoma growth and phenotype in a 3-D context. In the presence of mast cells or mast cell-conditioned medium, melanoma spheroid growth was profoundly reduced. To address the underlying mechanism, we conducted a transcriptomic analysis, which revealed that mast cell-conditioned medium had extensive effects on the melanoma gene expression patterns. Pathway analyses revealed profound effects on the expression of genes related to amino acid and protein metabolism. The conditioned medium also induced an upregulated expression of cancer-related genes, including adhesion molecules implicated in metastatic spreading. In line with this, after transfer to a Matrigel extracellular matrix milieu, spheroids that had been developed in the presence of mast cell-conditioned medium displayed enhanced elevated growth and adhesive properties. However, when assessing for possible effects of nutrient starvation, i.e., reduced nutrient content in mast cell-conditioned medium, we found that the observed effects on melanoma spheroid growth potentially could be explained by such effects. Hence, it cannot be excluded that the observed phenotypic alterations of melanoma spheroids grown in the presence of mast cells or mast cell-conditioned media are, at least partly, due to nutrient starvation rather than to the action of factors secreted by mast cells. Instead, our findings may provide insight into the effects on gene expression events that occur in melanoma tumors under nutrient stress.

Background Mast cells (MCs) have a profound impact on allergic asthma. Under such conditions, MCs undergo degranulation, resulting in the release of exceptionally large amounts of MC-restricted proteases. However, the role of these proteases in asthma is only partially understood. Objectives We sought to test our hypothesis that MC proteases can influence the functionality of human lung fibroblasts (HLFs). Methods Primary HLFs were treated with MC chymase or tryptase, followed by assessment of parameters related to fibroblast function. Results HLFs underwent major morphologic changes in response to chymase, showing signs of cellular contraction, but were refractory to tryptase. However, no effects of chymase on HLF viability or proliferation were seen. Chymase, but not tryptase, had a major impact on the output of extracellular matrix–associated compounds from the HLFs, including degradation of fibronectin and collagen-1, and activation of pro–matrix metalloprotease 2. Further, chymase induced the release of various chemotactic factors from HLFs. In line with this, conditioned medium from chymase-treated HLFs showed chemotactic activity on neutrophils. Transcriptome analysis revealed that chymase induced a proinflammatory gene transcription profile in HLFs, whereas tryptase had minimal effects. Conclusions Chymase, but not tryptase, has a major impact on the phenotype of primary airway fibroblasts by modifying their output of extracellular matrix components and by inducing a proinflammatory phenotype.

To obtain a more detailed picture of macrophage (MΦ) biology, in the current study, we analyzed the transcriptome of mouse peritoneal MΦs by RNA-seq and PCR-based transcriptomics. The results show that peritoneal MΦs, based on mRNA content, under non-inflammatory conditions produce large amounts of a number of antimicrobial proteins such as lysozyme and several complement components. They were also found to be potent producers of several chemokines, including platelet factor 4 (PF4), Ccl6, Ccl9, Cxcl13, and Ccl24, and to express high levels of both TGF-β1 and TGF-β2. The liver is considered to be the main producer of most complement and coagulation components. However, we can now show that MΦs are also important sources of such compounds including C1qA, C1qB, C1qC, properdin, C4a, factor H, ficolin, and coagulation factor FV. In addition, FX, FVII, and complement factor B were expressed by the MΦs, altogether indicating that MΦs are important local players in both the complement and coagulation systems. For comparison, we analyzed human peripheral blood monocytes. We show that the human monocytes shared many characteristics with the mouse peritoneal MΦs but that there were also many major differences. Similar to the mouse peritoneal MΦs, the most highly expressed transcript in the monocytes was lysozyme, and high levels of both properdin and ficolin were observed. However, with regard to connective tissue components, such as fibronectin, lubricin, syndecan 3, and extracellular matrix protein 1, which were highly expressed by the peritoneal MΦs, the monocytes almost totally lacked transcripts. In contrast, monocytes expressed high levels of MHC Class II, whereas the peritoneal MΦs showed very low levels of these antigen-presenting molecules. Altogether, the present study provides a novel view of the phenotype of the major MΦ subpopulation in the mouse peritoneum and the large peritoneal MΦs and places the transcriptome profile of the peritoneal MΦs in a broader context, including a comparison of the peritoneal MΦ transcriptome with that of human peripheral blood monocytes and the liver.