Effect of reflex and mechanical decreases in skin perfusion on thermal- and agonist-induced eccrine sweating in humans.

In humans, skin blood flux (SkBF) and eccrine sweating are tightly coupled, suggesting common neural control and regulation. This study was designed to separate these two sympathetic nervous system end-organ responses via nonadrenergic SkBF-decreasing mechanical perturbations during heightened sudomotor drive. We induced sweating physiologically via whole body heat stress using a high-density tube-lined suit (protocol 1; 2 women, 4 men), and pharmacologically via forearm intradermal microdialysis of two steady-state doses of a cholinergic agonist, pilocarpine (protocol 2; 4 women, 3 men). During sweating induction, we decreased SkBF via three mechanical perturbations: arm and leg dependency to engage the cutaneous venoarteriolar response (CVAR), limb venous occlusion to engage the CVAR and decrease perfusion pressure, and limb arterial occlusion to cause ischemia. In protocol 1, heat stress increased arm cutaneous vascular conductance and forearm sweat rate (capacitance hygrometry). During heat stress, despite decreases in SkBF during each of the acute (3 min) mechanical perturbations, eccrine sweat rate was unaffected. During heat stress with extended (10 min) ischemia, sweat rate decreased. In protocol 2, both pilocarpine doses (ED50 and EMAX) increased SkBF and sweat rate. Each mechanical perturbation resulted in decreased SkBF but minimal changes in eccrine sweat rate. Taken together, these data indicate that a wide range of acute decreases in SkBF do not appear to proportionally decrease either physiologically- or pharmacologically induced eccrine sweating in peripheral skin. This preservation of evaporative cooling despite acutely decreased SkBF could have consequential impacts for heat storage and balance during changes in body posture, limb position, or blood flow restrictive conditions.

Tape strips from early-onset pediatric atopic dermatitis highlight disease abnormalities in nonlesional skin.

BACKGROUND: Skin biopsies promote our understanding of atopic dermatitis/AD pathomechanisms in infants/toddlers with early-onset AD, but are not feasible in pediatric populations. Tape strips are an emerging, minimally invasive alternative, but global transcriptomic profiling in early pediatric AD is lacking. We aimed to provide global lesional and nonlesional skin profiles of infants/toddlers with recent-onset, moderate-to-severe AD using tape strips. METHODS: Sixteen tape strips were collected for RNA-seq profiling from 19 infants/toddlers (<5 years old; lesional and nonlesional) with early-onset moderate-to-severe AD (≤6 months) and 17 healthy controls. RESULTS: We identified 1829 differentially expressed genes/DEGs in lesional AD and 662 DEGs in nonlesional AD, vs healthy skin (fold-change ≥2, FDR <0.05), with 100% sample recovery. Both lesional and nonlesional skin showed significant dysregulations of Th2 (CCL17 and IL4R) and Th22/Th17 (IL36G, CCL20, and S100As)-related genes, largely lacking significant Th1-skewing. Significant down-regulation of terminal differentiation (FLG and FLG2), lipid synthesis/metabolism (ELOVL3 and FA2H), and tight junction (CLDN8) genes were primarily seen in lesional AD. Significant negative correlations were identified between Th2 measures and epidermal barrier gene-subsets and individual genes (FLG with IL-4R and CCL17; r < -0.4, P < .05). Significant correlations were also identified between clinical measures (body surface area/BSA, pruritus ADQ, and transepidermal water loss/TEWL) with immune and barrier mRNAs in lesional and/or nonlesional AD (FLG/FLG2 with TEWL; r < -0.4, P < .05). CONCLUSION: RNA-seq profiling using tape strips in early-onset pediatric AD captures immune and barrier alterations in both lesional and nonlesional skin. Tape strips provide insight into disease pathomechanisms and cutaneous disease activity.

Oxidant Signaling Mediated by Nox2 in Neutrophils Promotes Regenerative Myelopoiesis and Tissue Recovery following Ischemic Damage.

Ischemic tissue damage activates hematopoietic stem and progenitor cells (HSPCs) in the bone marrow (BM)-generating myeloid cells, and persistent HSPC activity may drive chronic inflammation and impair tissue recovery. Although increased reactive oxygen species in the BM regulate HSPC functions, their roles in myelopoiesis of activated HSPCs and subsequent tissue recovery during ischemic damage are not well understood. In this paper, we report that deletion of Nox2 NADPH oxidase in mice results in persistent elevations in BM HSPC activity and levels of inflammatory monocytes/macrophages in BM and ischemic tissue in a model of hindlimb ischemia. Ischemic tissue damage induces oxidants in BM such as elevations of hydrogen peroxide and oxidized phospholipids, which activate redox-sensitive Lyn kinase in a Nox2-dependent manner. Moreover, during tissue recovery after ischemic injury, this Nox2-ROS-Lyn kinase axis is induced by Nox2 in neutrophils that home to the BM, which inhibits HSPC activity and inflammatory monocyte generation and promotes tissue regeneration after ischemic damage. Thus, oxidant signaling in the BM mediated by Nox2 in neutrophils regulates myelopoiesis of HSPCs to promote regeneration of damaged tissue.

MicroCT angiography detects vascular formation and regression in skin wound healing.

Properly regulated angiogenesis and arteriogenesis are essential for effective wound healing. Tissue injury induces robust new vessel formation and subsequent vessel maturation, which involves vessel regression and remodeling. Although formation of functional vasculature is essential for healing, alterations in vascular structure over the time course of skin wound healing are not well understood. Here, using high-resolution ex vivo X-ray micro-computed tomography (microCT), we describe the vascular network during healing of skin excisional wounds with highly detailed three-dimensional (3D) reconstructed images and associated quantitative analysis. We found that relative vessel volume, surface area and branching number are significantly decreased in wounds from day 7 to days 14 and 21. Segmentation and skeletonization analysis of selected branches from high-resolution images as small as 2.5µm voxel size show that branching orders are decreased in the wound vessels during healing. In histological analysis, we found that the contrast agent fills mainly arterioles, but not small capillaries nor large veins. In summary, high-resolution microCT revealed dynamic alterations of vessel structures during wound healing. This technique may be useful as a key tool in the study of the formation and regression of wound vessels.

Macrophage PPARγ and impaired wound healing in type 2 diabetes.

Macrophages undergo a transition from pro-inflammatory to healing-associated phenotypes that is critical for efficient wound healing. However, the regulation of this transition during normal and impaired healing remains to be elucidated. In our studies, the switch in macrophage phenotypes during skin wound healing was associated with up-regulation of the peroxisome proliferator-activated receptor (PPAR)γ and its downstream targets, along with increased mitochondrial content. In the setting of diabetes, up-regulation of PPARγ activity was impaired by sustained expression of IL-1ß in both mouse and human wounds. In addition, experiments with myeloid-specific PPARγ knockout mice indicated that loss of PPARγ in macrophages is sufficient to prolong wound inflammation and delay healing. Furthermore, PPARγ agonists promoted a healing-associated macrophage phenotype both in vitro and in vivo, even in the diabetic wound environment. Importantly, topical administration of PPARγ agonists improved healing in diabetic mice, suggesting an appealing strategy for down-regulating inflammation and improving the healing of chronic wounds.

Sustained inflammasome activity in macrophages impairs wound healing in type 2 diabetic humans and mice.

The hypothesis of this study was that sustained activity of the Nod-like receptor protein (NLRP)-3 inflammasome in wounds of diabetic humans and mice contributes to the persistent inflammatory response and impaired healing characteristic of these wounds. Macrophages (Mp) isolated from wounds on diabetic humans and db/db mice exhibited sustained inflammasome activity associated with low level of expression of endogenous inflammasome inhibitors. Soluble factors in the biochemical milieu of these wounds are sufficient to activate the inflammasome, as wound-conditioned medium activates caspase-1 and induces release of interleukin (IL)-1ß and IL-18 in cultured Mp via a reactive oxygen species-mediated pathway. Importantly, inhibiting inflammasome activity in wounds of db/db mice using topical application of pharmacological inhibitors improved healing of these wounds, induced a switch from proinflammatory to healing-associated Mp phenotypes, and increased levels of prohealing growth factors. Furthermore, data generated from bone marrow-transfer experiments from NLRP-3 or caspase-1 knockout to db/db mice indicated that blocking inflammasome activity in bone marrow cells is sufficient to improve healing. Our findings indicate that sustained inflammasome activity in wound Mp contributes to impaired early healing responses of diabetic wounds and that the inflammasome may represent a new therapeutic target for improving healing in diabetic individuals.

Blocking interleukin-1ß induces a healing-associated wound macrophage phenotype and improves healing in type 2 diabetes.

Diabetes is associated with persistent inflammation and defective tissue repair responses. The hypothesis of this study was that interleukin (IL)-1ß is part of a proinflammatory positive feedback loop that sustains a persistent proinflammatory wound macrophage phenotype that contributes to impaired healing in diabetes. Macrophages isolated from wounds in diabetic humans and mice exhibited a proinflammatory phenotype, including expression and secretion of IL-1ß. The diabetic wound environment appears to be sufficient to induce these inflammatory phenomena because in vitro studies demonstrated that conditioned medium of both mouse and human wounds upregulates expression of proinflammatory genes and downregulates expression of prohealing factors in cultured macrophages. Furthermore, inhibiting the IL-1ß pathway using a neutralizing antibody and macrophages from IL-1 receptor knockout mice blocked the conditioned medium-induced upregulation of proinflammatory genes and downregulation of prohealing factors. Importantly, inhibiting the IL-1ß pathway in wounds of diabetic mice using a neutralizing antibody induced a switch from proinflammatory to healing-associated macrophage phenotypes, increased levels of wound growth factors, and improved healing of these wounds. Our findings indicate that targeting the IL-1ß pathway represents a new therapeutic approach for improving the healing of diabetic wounds.