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In this research we have used different cytokines and progesterone to enhance the immunomodulatory capacity of placental-derived stem cells (PLSCs) prior to their encapsulation. We assessed the effect of microencapsulation of the cells without (control) or after 3-day treatment with interferon gamma (INFγ), interleukin10 (IL-10), or progesterone (P4). Treated PLSCs demonstrated strong immunosuppressive effects on phytohemagglutinin (PHA)-activated peripheral blood mononuclear cells (PBMNCs). INFγ treatment resulted in the strongest immune inhibition among the treated groups. The treatments enhanced soluble human leukocyte antigen (sHLAG) secretion compared to control. The IL-10-treated group showed the highest effect on HLAG secretion compared to other groups. Alginate encapsulation of PLSCs did not affect cell viability, or sHLAG secretion. Also, after treatment the encapsulated PLSCs inhibited PHA-activated PBMNCs in the same manner as unencapsulated cells. We studied two groups of encapsulated PLSCs, one without perm-selective poly-L-ornithine (PLO)-coating and the other with PLO-coating, and measured levels of sHLAG secreted. We found no difference in sHLAG secretion between both groups. In summary, our data show that immunomodulatory function of the PLSC is not affected by encapsulation. These findings provide good promise for potential use of encapsulated PLSCs for immunomodulation treatment of disease by stem cell therapy.

Summary: Chronic allergic itch is a common symptom affecting millions of people and animals, but its pathogenesis is not fully explained. Herein, we show that periostin, abundantly expressed in the skin of patients with atopic dermatitis (AD), induces itch in mice, dogs, and monkeys. We identify the integrin αVβ3 expressed on a subset of sensory neurons as the periostin receptor. Using pharmacological and genetic approaches, we inhibited the function of neuronal integrin αVβ3, which significantly reduces periostin-induced itch in mice. Furthermore, we show that the cytokine TSLP, the application of AD-causing MC903 (calcipotriol), and house dust mites all induce periostin secretion. Finally, we establish that the JAK/STAT pathway is a key regulator of periostin secretion in keratinocytes. Altogether, our results identify a TSLP-periostin reciprocal activation loop that links the skin to the spinal cord via peripheral sensory neurons, and we characterize the non-canonical functional role of an integrin in itch. : Mishra et al. demonstrate periostin-induced itch in mice, dogs, and monkeys and identify the integrin αVβ3 as the periostin neuronal receptor. They find that keratinocytes release periostin in response to TSLP, thus identifying a possible reciprocal vicious circle implicating the cytokine TSLP and periostin in chronic allergic itch. Keywords: atopic dermatitis, AD, chronic allergic itch, integrin, JAK/STAT, keratinocytes, NPPB, periostin, sensory neuron, TRPV1, TRPA1, TSLP

Additional file 2 Contains supplementary figures, Figure S1–13. Format: PDF file.

Chronic allergic itch is a major symptom that affects millions of people and animals, but its pathogenesis is not fully explained. Here, we show that one of the mediators abundantly present in the skin of patients with atopic dermatitis, periostin, induces itch via sensory neurons in mice, dogs, and monkeys. We identified the periostin receptor, the integrin αvβ3 expressed on a subset of sensory neurons. Using pharmacological and genetic approaches, we inhibited the function of this neuronal integrin αvβ3, which significantly reduced periostin-mediated itch in mice. Furthermore, we showed that the cytokine TSLP, and the application of the AD-causing MC903 (calcipotriol) and house dust mites induced periostin secretion. Finally, we established that the JAK/STAT pathway is a key regulator of periostin secretion in keratinocytes. Overall, our results identified a TSLP-periostin reciprocal activation that links the skin to the central nervous system via peripheral sensory neurons, and we characterized the non-canonical functional role of a neuronal integrin in itch behavior.

This study aims to characterise the effect of fluid shear stress on human embryonic stem cell (hESC)-derived haematopoiesis and vasculogenesis by applying microfluidics, live cell imaging and single cell RNA sequencing. AGM-like haematopoietic cultures developed by Ng et al. were generated by manipulating WNT and TGFs signalling during mesoderm specification. The effect of pulsatile circulatory flow was studied in a microfluidic dynamic culture system, using SOX17 (Cherry)/RUNX1c(GFP) hESC line to read out arterial endothelial and haematopoietic differentiation, respectively. Microdevices generated cardiac-like, pulsatile flow in a circulatory culture system with a volume of 2–3µL. AGM-like haematopoietic development was observed by time-lapse imaging on chip for more than two weeks. We observed cells entering the circulation from the adherent layer, and the release of lightly tethered SOX17+ cells into the circulation. In parallel bulk differentiation culture using an orbital mixer to mimic the effect of wall shear stress, single cell RNA seq was performed on 8800 single cells at day 18 of culture. Control treatments were static culture and drug vehicle. Hierarchical cluster analysis identified 18 clusters belonging to erythroid-megakaryocytic, cardiovascular and myeloid differentiation pathways. We identified HOXA expressing mesenchymal cells in AGM cultures. Shear treatment promoted proliferative MYB, RUNX1, CD31 expressing blood progenitors, a reduced proportion of unipotent erythroid and megakaryocytic lineages, and increase numbers of myeloid and bipotent megakaryocyte-erythroid progenitors. The production of smooth muscle and cardiomyocytes was promoted by shear in non-AGM culture. This study demonstrates the feasibility of modelling human embryonic blood formation using microfluidic technology.

Figure S1. The result of mouse embryonic dental epithelium cell at E13.5 with only the mouse PPI background pathway data. Figure S2. The result of newborn mouse lens epithelium cell with only the mouse PPI background pathway data. Figure S3. The result of human PSC-derived ROR1+ cell (lens epithelium cell-like) with only the human PPI background pathway data. Figure S4. The result of newborn mouse lens fiber cell with only the mouse PPI background pathway data. Table S1. False positive rate calculation for SPAGI method of randomly assigns gene expression values of new born mouse lens epithelial cell and mouse tooth epithelial cell at embryonic day E13.5. Table S2. SPAGI test result for randomly assign gene expression values of new born mouse lens epithelial cell and mouse tooth epithelial cell at embryonic day E13.5. Table S3. Identification of known pathways by SPAGI and GO analysis methods. Table S4. Summary of known pathways identification by SPAGI and GO methods for randomly assigns genes of mouse lens epithelial cell. Table S5. Summary of known pathways identification by SPAGI and GO methods of randomly assigns genes for mouse tooth epithelial cell. Table S6. False positive rate of SPAGI and GO analysis method for known pathways. (PDF 658 kb)