Desmoglein 2 (Dsg2), a component of the desmosomal cell-cell adhesion structure, has been linked to invasion and metastasis in squamous cell carcinomas. However, it is unknown whether – and if so how – Dsg2 contributes to the malignant phenotype of keratinocytes. In this study, we addressed the consequences of Dsg2 overexpression under control of the involucrin promoter (Inv-Dsg2) in the epidermis of transgenic mice. These mice exhibited epidermal hyperkeratosis with slightly disrupted early and late differentiation markers, but intact epidermal barrier function. However, Inv-Dsg2 transgene expression was associated with extensive epidermal hyperplasia and increased keratinocyte proliferation in basal and suprabasal epidermal strata. Cultured Inv-Dsg2 keratinocytes showed enhanced cell survival in the anchorage-independent state that was critically dependent on EGF receptor activation and NF-κB activity. Consistent with the hyperproliferative and apoptosis-resistant phenotype of Inv-Dsg2 transgenic keratinocytes, we observed enhanced activation of multiple growth and survival pathways, including PI 3-kinase/AKT, MEK-MAPK, STAT3 and NF-κB, in the transgenic skin in situ. Finally, Inv-Dsg2 transgenic mice developed intraepidermal skin lesions resembling precancerous papillomas and were more susceptible to chemically induced carcinogenesis. In summary, overexpression of Dsg2 in epidermal keratinocytes deregulates multiple signaling pathways associated with increased growth rate, anchorage-independent cell survival, and the development of skin tumors in vivo.
Desmogleins play critical roles in cell adhesion and skin blistering diseases, as they are the target antigens of autoimmune antibodies and bacterial toxins. We recently cloned several novel members of the desmoglein gene family, bringing the number of desmogleins to four in the rat and human genomes and six in the mouse. Here, we have produced a monoclonal antibody to a cytoplasmic epitope of Dsg4, assessed its specificity and compared it to several existing Dsg1-3 antibodies. We also demonstrated cross-reactivity of commercially available and often used Dsg1 antibodies. Using these tools, we delineated the unique expression patterns of each desmoglein isoform in various human and mouse stratified squamous epithelia, including skin, hair, palm, and oral mucosa. Interestingly, in the epidermis, the expression of each desmoglein correlates with their gene arrangement in the cadherin locus. In human, Dsg4 was detected primarily in the granular and cornified cell layers of the epidermis, while present throughout all differentiated layers of the oral mucosa and palm, and in the matrix cells of anagen hair bulb. Similar pattern of expression for Dsg4 was observed in mouse, with the exception that it was expressed at significantly lower levels in the mouse epidermis. These results demonstrate the complexity of desmoglein gene expression and provide additional insights into the correlation between tissue expression patterns and disease phenotypes.
BackgroundMainland China experienced pandemic influenza H1N1 (2009) virus (pH1N1) with peak activity during November-December 2009. To understand the geographic extent, risk factors, and attack rate of pH1N1 infection in China we conducted a nationwide serological survey to determine the prevalence of antibodies to pH1N1.Methodology/Principal FindingsStored serum samples (n = 2,379) collected during 2006-2008 were used to estimate baseline serum reactogenicity to pH1N1. In January 2010, we used a multistage-stratified random sampling method to select 50,111 subjects who met eligibility criteria and collected serum samples and administered a standardized questionnaire. Antibody response to pH1N1 was measured using haemagglutination inhibition (HI) assay and the weighted seroprevalence was calculated using the Taylor series linearization method. Multivariable logistic regression analyses were used to examine risk factors for pH1N1 seropositivity. Baseline seroprevalence of pH1N1 antibody (HI titer ≥40) was 1.2%. The weighted seroprevalence of pH1N1 among the Chinese population was 21.5%(vaccinated: 62.0%; unvaccinated: 17.1%). Among unvaccinated participants, those aged 6-15 years (32.9%) and 16-24 years (30.3%) had higher seroprevalence compared with participants aged 25–59 years (10.7%) and ≥60 years (9.9%, P<0.0001). Children in kindergarten and students had higher odds of seropositivity than children in family care (OR: 1.36 and 2.05, respectively). We estimated that 207.7 million individuals (15.9%) experienced pH1N1 infection in China.Conclusions/SignificanceThe Chinese population had low pre-existing immunity to pH1N1 and experienced a relatively high attack rate in 2009 of this virus. We recommend routine control measures such as vaccination to reduce transmission and spread of seasonal and pandemic influenza viruses.
Objective Autograft microskin transplantation has been widely used as a skin graft therapy in full-thickness skin defect. However, skin grafting failure can lead to a pathological delay wound healing due to a poor vascularization bed. Considering the active role of adipose-derived stem cell (ADSC) in promoting angiogenesis, we intend to investigate the efficacy of autograft microskin combined with ADSC transplantation for facilitating wound healing in a full-thickness skin defect mouse model. Material and methods An in vivo full-thickness skin defect mouse model was used to evaluate the contribution of transplantation microskin and ADSC in wound healing. The angiogenesis was detected by immunohistochemistry staining. In vitro paracrine signaling pathway was evaluated by protein array and Gene Ontology, Kyoto Encyclopedia of Genes and Genomes pathway, and protein-protein interaction network analysis. Results Co-transplantation of microskin and ADSC potentiated the wound healing with better epithelization, smaller scar thickness, and higher angiogenesis (CD31) in the subcutaneous layer. We found both EGF and VEGF cytokines were secreted by microskin in vitro. Additionally, secretome proteomic analysis in a co-culture system of microskin and ADSC revealed that ADSC could secrete a wide range of important molecules to form a reacting network with microskin, including VEGF, IL-6, EGF, uPAR, MCP-3, G-CSF, and Tie-2, which most likely supported the angiogenesis effect as observed. Conclusion Overall, we concluded that the use of ADSC partially modulates microskin function and enhances wound healing by promoting angiogenesis in a full-thickness skin defect mouse model. Electronic supplementary material The online version of this article (10.1186/s13287-019-1389-4) contains supplementary material, which is available to authorized users.
Cellular function phenotype is regulated by various microRNAs (miRs), including miR-135a. However, how miR-135a is involved in the calcification in senescent vascular smooth muscle cells (VSMCs) is not clear yet. In the present study, we first identified the significantly altered miRNAs in VSMCs, then performed consecutive passage culture of VSMCs and analyzed the expression of miR-135a and calcification genes in the senescent phase. Next, the effects of the miR-135a inhibition on calcification and calcification genes were analyzed. The luciferase assay was used to validate the target protein of miR-135a. The western blotting was used to determine the effects of miR-135a on Krüppel-like factor 4 (KLF4) and signal transducer and activator of transcription 3 protein (STAT3) expression, as well as the relationship between KLF4 and STAT3. Finally, the quantified cellular calcification was measured to examine the involvement of miR-135a, KLF4 and STAT3 in VSMCs calcification. Our results showed that miR-135a was significantly altered in VSMCs. Cell calcification and calcification genes were greatly altered by miR-135a inhibition. KLF4 was validated as the target RNA of miR-135a. Expression of KLF4 and STAT3 were both significantly decreased by over expressed miR-135a, while the inhibition of miR-135a and KLF4 siRNA both decreased the STAT3 protein levels. Moreover, the inhibition of miR-135a dramatically increased the calcium concentration, but co-treatment with KLF4 or STAT3 siRNA both decreased the calcium concentration. The present study identified miR-135a as a potential osteogenic differentiation suppressor in senescent VSMCs and revealed that KLF4/STAT3 pathway, at least partially, was involved in the mechanism.
Severe burns are challenging to heal and result in significant death throughout the world. Adipose-derived mesenchymal stem cells (ADSCs) have emerged as a promising treatment for full-thickness burn healing but are impeded by their low viability and efficiency after grafting in vivo. Nitric oxide (NO) is beneficial in promoting stem cell bioactivity, but whether it can function effectively in vivo is still largely unknown. In this study, we bioprinted an efficient biological scaffold loaded with ADSCs and NO (3D-ADSCs/NO) to evaluate its biological efficacy in promoting severe burn wound healing. The integral 3D-ADSCs/NO hydrogel scaffolds were constructed via 3D bioprinting. Our results shown that 3D-ADSCs/NO can enhance the migration and angiogenesis of Human Umbilical Vein Endothelial Cells (HUVECs). Burn wound healing experiments in mice revealed that 3D-ADSCs/NO accelerated the wound healing by promoting faster epithelialization and collagen deposition. Notably, immunohistochemistry of CD31 suggested an increase in neovascularization, supported by the upregulation of vascular endothelial growth factor (VEGF) mRNA in ADSCs in the 3D biosystem. These findings indicated that 3D-ADSC/NO hydrogel scaffold can promote severe burn wound healing through increased neovascularization via the VEGF signalling pathway. This scaffold may be considered a promising strategy for healing severe burns.
Cell-cell adhesion mediated by desmosomes is crucial for maintaining proper epidermal structure and function, as evidenced by several severe and potentially fatal skin disorders involving impairment of desmosomal proteins. Pemphigus foliaceus (PF) and staphylococcal scalded skin syndrome (SSSS) are subcorneal blistering diseases resulting from loss of function of the desmosomal cadherin, desmoglein 1 (Dsg1). To further study the pathomechanism of these diseases and to assess the adhesive properties of Dsg2, we employed a recently established transgenic (Tg) mouse model expressing Dsg2 in the superficial epidermis. Neonatal Tg and wild type (WT) mice were injected with purified ETA or PF Ig. We showed that ectopic expression of Dsg2 reduced the extent of blister formation in response to both ETA and PF Ig. In response to PF Ig, we observed either a dramatic loss or a reorganization of Dsg1-α, Dsg1-β, and, to a lesser extent, Dsg1-γ, in WT mice. The Inv-Dsg2 Tg mice showed enhanced retention of Dsg1 at the cell-cell border. Collectively, our data support the role for Dsg2 in cell adhesion and suggest that ectopic superficial expression of Dsg2 can increase membrane preservation of Dsg1 and limit epidermal blister formation mediated by PF antibodies and exfoliative toxins.
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