Project description:The vitamin D receptor (VDR) regulates cell proliferation and differentiation including epidermal keratinocytes by modulating transcription of its target genes. We are investigating the role of VDR in epidermal stem cells and their progenies in the regeneration process of epidermis and hair in the skin. VDR null mice are utilized in which VDR is specifically deleted in keratin 14 (K14) expressing keratinocytes by Cre-lox strategy. The impact of VDR deletion was evaluated by comparison of VDR null mice with no cre littermate control mice. The VDR was abundantly expressed in potential epidermal stem cells including basal cells in interfollicular epidermis (IFE), and in CD34 expressing bulge keratinocytes in hair follicles. Gene expression profiles and subsequent pathway analysis of stem cell enriched keratinocyte populations revealed that the VDR deletion significantly suppressed β-catenin signaling as well as VDR signaling. The role of VDR in epidermal stem cells was studied during hair follicle cycling and wound healing processes. The epidermal stem cells were not appropriately stimulated by hair depilation, and did not reinitiate anagen in the hair follicles resulting in a failure of hair regrowth. In addition, the stem cells were not fully activated after full thickness wounds were generated in VDR null skin under a low calcium diet to suppress compensation pathways. Cell proliferation was not fully induced in potential stem cells located in both IFE and hair follicles near the wounding edges, and re-epithelialization rate was delayed in VDR null skin. Gene expression profiling of the wounded skin (3 days after injury) indicated that β-catenin signaling was not fully induced in VDR null skin comparable to that observed in β-catenin null mice. The β-catenin target genes including Axin2 and cell cycle regulators involved in epidermal stem cell function were not induced in the edges of the wound of VDR null skin. These results demonstrated that VDR plays an essential role in hair cycling and wound healing processes through regulation of β-catenin signaling in epidermal stem cells and their progenies. n=3 CON and KO (each sample contain RNA extracted from keratinocytes, which is isolated from VDR KO and littermate control skins excised from 3 mice)

Project description:The vitamin D receptor (VDR) regulates cell proliferation and differentiation including epidermal keratinocytes by modulating transcription of its target genes. We are investigating the role of VDR in epidermal stem cells and their progenies in the regeneration process of epidermis and hair in the skin. VDR null mice are utilized in which VDR is specifically deleted in keratin 14 (K14) expressing keratinocytes by Cre-lox strategy. The impact of VDR deletion was evaluated by comparison of VDR null mice with no cre littermate control mice. The VDR was abundantly expressed in potential epidermal stem cells including basal cells in interfollicular epidermis (IFE), and in CD34 expressing bulge keratinocytes in hair follicles. Gene expression profiles and subsequent pathway analysis of stem cell enriched keratinocyte populations revealed that the VDR deletion significantly suppressed β-catenin signaling as well as VDR signaling. The role of VDR in epidermal stem cells was studied during hair follicle cycling and wound healing processes. The epidermal stem cells were not appropriately stimulated by hair depilation, and did not reinitiate anagen in the hair follicles resulting in a failure of hair regrowth. In addition, the stem cells were not fully activated after full thickness wounds were generated in VDR null skin under a low calcium diet to suppress compensation pathways. Cell proliferation was not fully induced in potential stem cells located in both IFE and hair follicles near the wounding edges, and re-epithelialization rate was delayed in VDR null skin. Gene expression profiling of the wounded skin (3 days after injury) indicated that β-catenin signaling was not fully induced in VDR null skin comparable to that observed in β-catenin null mice. The β-catenin target genes including Axin2 and cell cycle regulators involved in epidermal stem cell function were not induced in the edges of the wound of VDR null skin. These results demonstrated that VDR plays an essential role in hair cycling and wound healing processes through regulation of β-catenin signaling in epidermal stem cells and their progenies.

Project description:Schistosomes are parasitic worms that can survive in the hostile environment of the human bloodstream where they appear refractory to both immune elimination and thrombus formation. We hypothesize that parasite migration in the bloodstream can stress the vascular endothelium causing this tissue to release chemicals alerting responsive host cells to the stress. Such chemicals are called damage associated molecular patterns (DAMPs) and among the most potent is the proinflammatory mediator, adenosine triphosphate (ATP). Furthermore, the ATP derivative ADP is a pro-thrombotic molecule that acts as a strong activator of platelets. Schistosomes are reported to possess at their host interactive tegumental surface a series of enzymes that could, like their homologs in mammals, degrade extracellular ATP and ADP. These are alkaline phosphatase (SmAP), phosphodiesterase (SmNPP-5) and ATP diphosphohydrolase (SmATPDase1). In this work we employ RNAi to knock down expression of the genes encoding these enzymes in the intravascular life stages of the parasite. We then compare the abilities of these parasites to degrade exogenously added ATP and ADP. We find that only SmATPDase1-suppressed parasites are significantly impaired in their ability to degrade these nucleotides. Suppression of SmAP or SmNPP-5 does not appreciably affect the worms' ability to catabolize ATP or ADP. These findings are confirmed by the functional characterization of the enzymatically active, full-length recombinant SmATPDase1 expressed in CHO-S cells. The enzyme is a true apyrase; SmATPDase1 degrades ATP and ADP in a cation dependent manner. Optimal activity is seen at alkaline pH. The Km of SmATPDase1 for ATP is 0.4 ± 0.02 mM and for ADP, 0.252 ± 0.02 mM. The results confirm the role of tegumental SmATPDase1 in the degradation of the exogenous pro-inflammatory and pro-thrombotic nucleotides ATP and ADP by live intravascular stages of the parasite. By degrading host inflammatory signals like ATP, and pro-thrombotic signals like ADP, these parasite enzymes may minimize host immune responses, inhibit blood coagulation and promote schistosome survival.

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Epidermal keratinocytes initiate wound healing and pro-inflammatory immune responses following percutaneous schistosome infection.

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